WorldWideScience

Sample records for subnuclear targeting signals

  1. Targeted deficiency of the transcriptional activator Hnf1alpha alters subnuclear positioning of its genomic targets.

    Directory of Open Access Journals (Sweden)

    Reini F Luco

    2008-05-01

    Full Text Available DNA binding transcriptional activators play a central role in gene-selective regulation. In part, this is mediated by targeting local covalent modifications of histone tails. Transcriptional regulation has also been associated with the positioning of genes within the nucleus. We have now examined the role of a transcriptional activator in regulating the positioning of target genes. This was carried out with primary beta-cells and hepatocytes freshly isolated from mice lacking Hnf1alpha, an activator encoded by the most frequently mutated gene in human monogenic diabetes (MODY3. We show that in Hnf1a-/- cells inactive endogenous Hnf1alpha-target genes exhibit increased trimethylated histone H3-Lys27 and reduced methylated H3-Lys4. Inactive Hnf1alpha-targets in Hnf1a-/- cells are also preferentially located in peripheral subnuclear domains enriched in trimethylated H3-Lys27, whereas active targets in wild-type cells are positioned in more central domains enriched in methylated H3-Lys4 and RNA polymerase II. We demonstrate that this differential positioning involves the decondensation of target chromatin, and show that it is spatially restricted rather than a reflection of non-specific changes in the nuclear organization of Hnf1a-deficient cells. This study, therefore, provides genetic evidence that a single transcriptional activator can influence the subnuclear location of its endogenous genomic targets in primary cells, and links activator-dependent changes in local chromatin structure to the spatial organization of the genome. We have also revealed a defect in subnuclear gene positioning in a model of a human transcription factor disease.

  2. Subnuclear proteomics in colorectal cancer

    DEFF Research Database (Denmark)

    Albrethsen, Jakob; Knol, Jaco C; Piersma, Sander R

    2010-01-01

    Abnormalities in nuclear phenotype and chromosome structure are key features of cancer cells. Investigation of the protein determinants of nuclear subfractions in cancer may yield molecular insights into aberrant chromosome function and chromatin organization and in addition may yield biomarkers...... for early cancer detection. Here we evaluate a proteomics work flow for profiling protein constituents in subnuclear domains in colorectal cancer tissues and apply this work flow to a comparative analysis of the nuclear matrix fraction in colorectal adenoma and carcinoma tissue samples. First, we...... established the reproducibility of the entire work flow. In a reproducibility analysis of three nuclear matrix fractions independently isolated from the same colon tumor homogenate, 889 of 1,047 proteins (85%) were reproducibly identified at high confidence (minimally two peptides per protein at 99...

  3. International School of Subnuclear Physics 50th Course

    CERN Document Server

    What we would like LHC to give us; ISSP 2012

    2014-01-01

    This book is the proceedings of the International School of Subnuclear Physics, ISSP 2012, 50th Course — ERICE, 23 June 2013 — 2 July 2012. This course was devoted to the celebrations of the 50th Anniversary of the Subnuclear Physics School which was started in 1961 by Antonino Zichichi with John Bell at CERN and formally established in 1962 by Bell, Blackett, Weisskopf, Rabi and Zichichi in Geneva (CERN). The lectures covered the latest and most significant achievements in theoretical and in experimental subnuclear physics. Readership: Directed to experts and advanced-level students in the field of Theoretical and Experimental Subnuclear Physics.

  4. Subnuclear globules in simple mucinous glands of the gastrointestinal tract

    DEFF Research Database (Denmark)

    Holck, Susanne; Ingeholm, P.

    2007-01-01

    Pyloric and duodenal Brunner glands may be the site of a peculiar cytoplasmic change characterized by clear vacuoles or glassy eosinophilic globules, occupying a subnuclear position with displacement of the nuclei towards the lumenal surface. To our knowledge this previously documented,5,6 although...

  5. Targeting mutant NRAS signaling pathways in melanoma.

    Science.gov (United States)

    Vu, Ha Linh; Aplin, Andrew E

    2016-05-01

    Cutaneous melanoma is a devastating form of skin cancer and its incidence is increasing faster than any other preventable cancer in the United States. The mutant NRAS subset of melanoma is more aggressive and associated with poorer outcomes compared to non-NRAS mutant melanoma. The aggressive nature and complex molecular signaling conferred by this transformation has evaded clinically effective treatment options. This review examines the major downstream effectors of NRAS relevant in melanoma and the associated advances made in targeted therapies that focus on these effector pathways. We outline the history of MEK inhibition in mutant NRAS melanoma and recent advances with newer MEK inhibitors. Since MEK inhibitors will likely be optimized when combined with other targeted therapies, we focus on recently identified targets that can be used in combination with MEK inhibitors. Published by Elsevier Ltd.

  6. Subnuclear localization of proteins encoded by the oncogene v-myb and its cellular homolog c-myb.

    OpenAIRE

    Klempnauer, K H; Sippel, A E

    1986-01-01

    The retroviral transforming gene v-myb encodes a 45,000-Mr nuclear transforming protein (p45v-myb). p45v-myb is a truncated and mutated version of a 75,000-Mr protein encoded by the chicken c-myb gene (p75c-myb). Like its viral counterpart, p75c-myb is located in the cell nucleus. As a first step in identifying nuclear targets involved in cellular transformation by v-myb and in c-myb function, we determined the subnuclear locations of p45v-myb and p75c-myb. Approximately 80 to 90% of the tota...

  7. Targeting RAGE Signaling in Inflammatory Disease.

    Science.gov (United States)

    Hudson, Barry I; Lippman, Marc E

    2018-01-29

    The receptor for advanced glycation end-products (RAGE) is a multiligand pattern recognition receptor implicated in diverse chronic inflammatory states. RAGE binds and mediates the cellular response to a range of damage-associated molecular pattern molecules (DAMPs) including AGEs, HMGB1, S100s, and DNA. RAGE can also act as an innate immune sensor of microbial pathogen-associated molecular pattern molecules (PAMPs) including bacterial endotoxin, respiratory viruses, and microbial DNA. RAGE is expressed at low levels under normal physiology, but it is highly upregulated under chronic inflammation because of the accumulation of various RAGE ligands. Blocking RAGE signaling in cell and animal models has revealed that targeting RAGE impairs inflammation and progression of diabetic vascular complications, cardiovascular disease (CVD), and cancer progression and metastasis. The clinical relevance of RAGE in inflammatory disease is being demonstrated in emerging clinical trials of novel small-molecule RAGE inhibitors.

  8. Targeting Signaling Pathways in Epithelial Ovarian Cancer

    Directory of Open Access Journals (Sweden)

    Johannes Haybaeck

    2013-05-01

    Full Text Available Ovarian carcinoma (OC is the most lethal gynecological malignancy. Response to platinum-based chemotherapy is poor in some patients and, thus, current research is focusing on new therapy options. The various histological types of OC are characterized by distinctive molecular genetic alterations that are relevant for ovarian tumorigenesis. The understanding of these molecular pathways is essential for the development of novel therapeutic strategies. Purpose: We want to give an overview on the molecular genetic changes of the histopathological types of OC and their role as putative therapeutic targets. In Depth Review of Existing Data: In 2012, the vascular endothelial growth factor (VEGF inhibitor, bevacizumab, was approved for OC treatment. Bevacizumab has shown promising results as single agent and in combination with conventional chemotherapy, but its target is not distinctive when analyzed before treatment. At present, mammalian target of rapamycin (mTOR inhibitors, poly-ADP-ribose polymerase (PARP inhibitors and components of the EGFR pathway are in the focus of clinical research. Interestingly, some phytochemical substances show good synergistic effects when used in combination with chemotherapy. Conclusion: Ongoing studies of targeted agents in conjunction with chemotherapy will show whether there are alternative options to bevacizumab available for OC patients. Novel targets which can be assessed before therapy to predict efficacy are needed. The assessment of therapeutic targets is continuously improved by molecular pathological analyses on tumor tissue. A careful selection of patients for personalized treatment will help to reduce putative side effects and toxicity.

  9. The retinoid anticancer signal: mechanisms of target gene regulation

    OpenAIRE

    Liu, T.; Bohlken, A; Kuljaca, S; Lee, M.; Nguyen, T; S.; Smith; Cheung, B; Norris, M D; Haber, M; Holloway, A.J.; Bowtell, D D L; Marshall, G M

    2005-01-01

    Retinoids induce growth arrest, differentiation, and cell death in many cancer cell types. One factor determining the sensitivity or resistance to the retinoid anticancer signal is the transcriptional response of retinoid-regulated target genes in cancer cells. We used cDNA microarray to identify 31 retinoid-regulated target genes shared by two retinoid-sensitive neuroblastoma cell lines, and then sought to determine the relevance of the target gene responses to the retinoid anticancer signal...

  10. Targeting and localized signalling by small GTPases

    NARCIS (Netherlands)

    ten Klooster, Jean Paul; Hordijk, Peter L.

    2007-01-01

    Polarized cellular responses, for example, cell migration, require the co-ordinated assembly of signalling complexes at a particular subcellular location, such as the leading edge of cells. Small GTPases of the Ras superfamily play central roles in many (polarized) responses to growth factors,

  11. Tracking target signal strengths on a grid using sparsity

    NARCIS (Netherlands)

    Farahmand, S.; Giannakis, G.B.; Leus, G.J.T.; Tian, Z.

    2014-01-01

    Multi-target tracking is mainly challenged by the nonlinearity present in the measurement equation and the difficulty in fast and accurate data association. To overcome these challenges, the present paper introduces a grid-based model in which the state captures target signal strengths on a known

  12. ESTIMATION OF SIGNAL PARAMETERS FOR MULTIPLE TARGET LOCALIZATION

    Directory of Open Access Journals (Sweden)

    X. Susan Christina

    2014-12-01

    Full Text Available Target detection and localization is an active research area due to its importance in a wide range of application such as biomedical and military applications. In this paper, a novel method for the detection and estimation of signal parameters such as range and direction of arrival for multiple far- field target using wideband echo chirp signals is proposed. Sonar and radar are the active detection systems transmit well defined signals into a region of interest. A model preprocessing procedure is designed for the echo signal. The parameters estimation method for multiple targets is developed based on the Linear Canonical transform and the Fast Root MUSIC algorithm which is a high resolution DOA estimation method originally proposed for any arbitrary arrays to reduce the computational complexity in existing systems. The proposed method provides high accuracy of detection and resolution even in very low SNR values.

  13. Signal recognition partcle mediated protein targeting in Escherichia coli

    NARCIS (Netherlands)

    Valent, Q.A.

    2001-01-01

    The signal recognition particle (SRP) is a conserved ribonucleoprotein complex that binds to targeting sequences in nascent secretory and membrane proteins. The SRP guides these proteins to the cytoplasmic membrane in prokaryotes and the endoplasmic reticulum membrane in eukaryotes via an

  14. Probability distribution functions of echo signals from meteorological targets

    Science.gov (United States)

    Vasilyev, G. V.

    1975-01-01

    Simple expressions are obtained for the laws and moments of the probability distributions of averaged echo signals from meteorological targets at the output of a logarithmic radar receiver. Here, the distribution function is assumed to be represented in the form of an Edgeworth series.

  15. The Nuclear Protein Database (NPD): sub-nuclear localisation and functional annotation of the nuclear proteome

    Science.gov (United States)

    Dellaire, G.; Farrall, R.; Bickmore, W.A.

    2003-01-01

    The Nuclear Protein Database (NPD) is a curated database that contains information on more than 1300 vertebrate proteins that are thought, or are known, to localise to the cell nucleus. Each entry is annotated with information on predicted protein size and isoelectric point, as well as any repeats, motifs or domains within the protein sequence. In addition, information on the sub-nuclear localisation of each protein is provided and the biological and molecular functions are described using Gene Ontology (GO) terms. The database is searchable by keyword, protein name, sub-nuclear compartment and protein domain/motif. Links to other databases are provided (e.g. Entrez, SWISS-PROT, OMIM, PubMed, PubMed Central). Thus, NPD provides a gateway through which the nuclear proteome may be explored. The database can be accessed at http://npd.hgu.mrc.ac.uk and is updated monthly. PMID:12520015

  16. The Nuclear Protein Database (NPD): sub-nuclear localisation and functional annotation of the nuclear proteome.

    Science.gov (United States)

    Dellaire, G; Farrall, R; Bickmore, W A

    2003-01-01

    The Nuclear Protein Database (NPD) is a curated database that contains information on more than 1300 vertebrate proteins that are thought, or are known, to localise to the cell nucleus. Each entry is annotated with information on predicted protein size and isoelectric point, as well as any repeats, motifs or domains within the protein sequence. In addition, information on the sub-nuclear localisation of each protein is provided and the biological and molecular functions are described using Gene Ontology (GO) terms. The database is searchable by keyword, protein name, sub-nuclear compartment and protein domain/motif. Links to other databases are provided (e.g. Entrez, SWISS-PROT, OMIM, PubMed, PubMed Central). Thus, NPD provides a gateway through which the nuclear proteome may be explored. The database can be accessed at http://npd.hgu.mrc.ac.uk and is updated monthly.

  17. A novel putative miRNA target enhancer signal.

    Directory of Open Access Journals (Sweden)

    Thorsten Schmidt

    Full Text Available It is known that miRNA target sites are very short and the effect of miRNA-target site interaction alone appears as being unspecific. Recent experiments suggest further context signals involved in miRNA target site recognition and regulation. Here, we present a novel GC-rich RNA motif downstream of experimentally supported miRNA target sites in human mRNAs with no similarity to previously reported functional motifs. We demonstrate that the novel motif can be found in at least one third of all transcripts regulated by miRNAs. Furthermore, we show that motif occurrence and the frequency of miRNA target sites as well as the stability of their duplex structures correlate. The finding, that the novel motif is significantly associated with miRNA target sites, suggests a functional role of the motif in miRNA target site biology. Beyond, the novel motif has the impact to improve prediction of miRNA target sites significantly.

  18. Visual signals bias auditory targets in azimuth and depth.

    Science.gov (United States)

    Bowen, Amanda L; Ramachandran, Ramnarayan; Muday, Jeffrey A; Schirillo, James A

    2011-10-01

    In the psychophysical phenomenon visual bias, an accurately localized irrelevant signal, such as a light, impairs localization of a spatially discrepant target, such as a sound, when the two stimuli are perceived as unified. Many studies have demonstrated visual bias in azimuth, but none have tested directly or found this effect in depth. The current study was able to produce over 90% bias in azimuth and somewhat less (83%) bias in depth. A maximum likelihood estimate can predict bias by the variance in the localization of each unimodal signal in each dimension in space.

  19. GPR Signal Denoising and Target Extraction With the CEEMD Method

    KAUST Repository

    Li, Jing

    2015-04-17

    In this letter, we apply a time and frequency analysis method based on the complete ensemble empirical mode decomposition (CEEMD) method in ground-penetrating radar (GPR) signal processing. It decomposes the GPR signal into a sum of oscillatory components, with guaranteed positive and smoothly varying instantaneous frequencies. The key idea of this method relies on averaging the modes obtained by empirical mode decomposition (EMD) applied to several realizations of Gaussian white noise added to the original signal. It can solve the mode-mixing problem in the EMD method and improve the resolution of ensemble EMD (EEMD) when the signal has a low signal-to-noise ratio. First, we analyze the difference between the basic theory of EMD, EEMD, and CEEMD. Then, we compare the time and frequency analysis with Hilbert-Huang transform to test the results of different methods. The synthetic and real GPR data demonstrate that CEEMD promises higher spectral-spatial resolution than the other two EMD methods in GPR signal denoising and target extraction. Its decomposition is complete, with a numerically negligible error.

  20. Signaling pathways relevant to cognition-enhancing drug targets.

    Science.gov (United States)

    Ménard, Caroline; Gaudreau, Pierrette; Quirion, Rémi

    2015-01-01

    Aging is generally associated with a certain cognitive decline. However, individual differences exist. While age-related memory deficits can be observed in humans and rodents in the absence of pathological conditions, some individuals maintain intact cognitive functions up to an advanced age. The mechanisms underlying learning and memory processes involve the recruitment of multiple signaling pathways and gene expression, leading to adaptative neuronal plasticity and long-lasting changes in brain circuitry. This chapter summarizes the current understanding of how these signaling cascades could be modulated by cognition-enhancing agents favoring memory formation and successful aging. It focuses on data obtained in rodents, particularly in the rat as it is the most common animal model studied in this field. First, we will discuss the role of the excitatory neurotransmitter glutamate and its receptors, downstream signaling effectors [e.g., calcium/calmodulin-dependent protein kinase II (CaMKII), protein kinase C (PKC), extracellular signal-regulated kinases (ERK), mammalian target of rapamycin (mTOR), cAMP response element-binding protein (CREB)], associated immediate early gene (e.g., Homer 1a, Arc and Zif268), and growth factors [insulin-like growth factors (IGFs) and brain-derived neurotrophic factor (BDNF)] in synaptic plasticity and memory formation. Second, the impact of the cholinergic system and related modulators on memory will be briefly reviewed. Finally, since dynorphin neuropeptides have recently been associated with memory impairments in aging, it is proposed as an attractive target to develop novel cognition-enhancing agents.

  1. Missile-Borne SAR Raw Signal Simulation for Maneuvering Target

    Directory of Open Access Journals (Sweden)

    Weijie Xia

    2016-01-01

    Full Text Available SAR raw signal simulation under the case of maneuver and high-speed has been a challenging and urgent work recently. In this paper, a new method based on one-dimensional fast Fourier transform (1DFFT algorithm is presented for raw signal simulation of maneuvering target for missile-borne SAR. Firstly, SAR time-domain raw signal model is given and an effective Range Frequency Azimuth Time (RFAT algorithm based on 1DFFT is derived. In this algorithm, the “Stop and Go” (SaG model is adopted and the wide radar scattering characteristic of target is taken into account. Furthermore, the “Inner Pulse Motion” (IPM model is employed to deal with high-speed case. This new RFAT method can handle the maneuvering cases, high-speed cases, and bistatic radar cases, which are all possible in the missile-borne SAR. Besides, this raw signal simulation adopts the electromagnetic scattering calculation so that we do not need a scattering rate distribution map as the simulation input. Thus, the multiple electromagnetic reflections can be considered. Simulation examples prove the effectiveness of our method.

  2. Targeting glutamate signalling in depression: progress and prospects.

    Science.gov (United States)

    Murrough, James W; Abdallah, Chadi G; Mathew, Sanjay J

    2017-07-01

    Major depressive disorder (MDD) is severely disabling, and current treatments have limited efficacy. The glutamate N-methyl-D-aspartate receptor (NMDAR) antagonist ketamine was recently repurposed as a rapidly acting antidepressant, catalysing the vigorous investigation of glutamate-signalling modulators as novel therapeutic agents for depressive disorders. In this Review, we discuss the progress made in the development of such modulators for the treatment of depression, and examine recent preclinical and translational studies that have investigated the mechanisms of action of glutamate-targeting antidepressants. Fundamental questions remain regarding the future prospects of this line of drug development, including questions concerning safety and tolerability, efficacy, dose-response relationships and therapeutic mechanisms.

  3. An ensemble method for predicting subnuclear localizations from primary protein structures.

    Directory of Open Access Journals (Sweden)

    Guo Sheng Han

    Full Text Available BACKGROUND: Predicting protein subnuclear localization is a challenging problem. Some previous works based on non-sequence information including Gene Ontology annotations and kernel fusion have respective limitations. The aim of this work is twofold: one is to propose a novel individual feature extraction method; another is to develop an ensemble method to improve prediction performance using comprehensive information represented in the form of high dimensional feature vector obtained by 11 feature extraction methods. METHODOLOGY/PRINCIPAL FINDINGS: A novel two-stage multiclass support vector machine is proposed to predict protein subnuclear localizations. It only considers those feature extraction methods based on amino acid classifications and physicochemical properties. In order to speed up our system, an automatic search method for the kernel parameter is used. The prediction performance of our method is evaluated on four datasets: Lei dataset, multi-localization dataset, SNL9 dataset and a new independent dataset. The overall accuracy of prediction for 6 localizations on Lei dataset is 75.2% and that for 9 localizations on SNL9 dataset is 72.1% in the leave-one-out cross validation, 71.7% for the multi-localization dataset and 69.8% for the new independent dataset, respectively. Comparisons with those existing methods show that our method performs better for both single-localization and multi-localization proteins and achieves more balanced sensitivities and specificities on large-size and small-size subcellular localizations. The overall accuracy improvements are 4.0% and 4.7% for single-localization proteins and 6.5% for multi-localization proteins. The reliability and stability of our classification model are further confirmed by permutation analysis. CONCLUSIONS: It can be concluded that our method is effective and valuable for predicting protein subnuclear localizations. A web server has been designed to implement the proposed method

  4. Induction of Synthetic Lethality by Natural Compounds Targeting Cancer Signaling.

    Science.gov (United States)

    Farrand, Lee; Byun, Sanguine

    2017-11-16

    Despite the breakthroughs that have been achieved, significant unmet needs relating to the inadequate efficacy and toxicity of currently-available cancer therapies remain. Kinase inhibitors are a class of agents that target signaling factors responsible for the survival of malignant cells, and may address at least some of these issues. The concept of synthetic lethality provides a potential solution to counteract pathway redundancies, and refers to situations in which a mutation in one of two particular genes alone permits cell survival, while simultaneous mutation in both results in cell death. When exploited in the context of cancer therapy, pathways that are uniquely upregulated in cancer cells become selective targets, with reduced off-target toxicity toward their healthy counterparts. Natural compounds represent a large and readily-accessible library of bioactive structures that can be screened for synthetically lethal interactions by testing for the inhibition of kinases relevant to cancer cell survival. In this review, we discuss the concept of synthetic lethality and focus on scenarios in which natural compounds that target kinases may be applied to tip the balance in favor of cancer cell death during therapeutic challenge. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  5. Glycation & the RAGE axis: targeting signal transduction through DIAPH1.

    Science.gov (United States)

    Shekhtman, Alexander; Ramasamy, Ravichandran; Schmidt, Ann Marie

    2017-02-01

    The consequences of chronic disease are vast and unremitting; hence, understanding the pathogenic mechanisms mediating such disorders holds promise to identify therapeutics and diminish the consequences. The ligands of the receptor for advanced glycation end products (RAGE) accumulate in chronic diseases, particularly those characterized by inflammation and metabolic dysfunction. Although first discovered and reported as a receptor for advanced glycation end products (AGEs), the expansion of the repertoire of RAGE ligands implicates the receptor in diverse milieus, such as autoimmunity, chronic inflammation, obesity, diabetes, and neurodegeneration. Areas covered: This review summarizes current knowledge regarding the ligand families of RAGE and data from human subjects and animal models on the role of the RAGE axis in chronic diseases. The recent discovery that the cytoplasmic domain of RAGE binds to the formin homology 1 (FH1) domain, DIAPH1, and that this interaction is essential for RAGE ligand-stimulated signal transduction, is discussed. Finally, we review therapeutic opportunities targeting the RAGE axis as a means to mitigate chronic diseases. Expert commentary: With the aging of the population and the epidemic of cardiometabolic disease, therapeutic strategies to target molecular pathways that contribute to the sequelae of these chronic diseases are urgently needed. In this review, we propose that the ligand/RAGE axis and its signaling nexus is a key factor in the pathogenesis of chronic disease and that therapeutic interruption of this pathway may improve quality and duration of life.

  6. Lysophosphatidic acid targets vascular and oncogenic pathways via RAGE signaling

    Science.gov (United States)

    Touré, Fatouma; Chitayat, Seth; Pei, Renjun; Song, Fei; Li, Qing; Zhang, Jinghua; Rosario, Rosa; Ramasamy, Ravichandran; Chazin, Walter J.

    2012-01-01

    The endogenous phospholipid lysophosphatidic acid (LPA) regulates fundamental cellular processes such as proliferation, survival, motility, and invasion implicated in homeostatic and pathological conditions. Hence, delineation of the full range of molecular mechanisms by which LPA exerts its broad effects is essential. We report avid binding of LPA to the receptor for advanced glycation end products (RAGE), a member of the immunoglobulin superfamily, and mapping of the LPA binding site on this receptor. In vitro, RAGE was required for LPA-mediated signal transduction in vascular smooth muscle cells and C6 glioma cells, as well as proliferation and migration. In vivo, the administration of soluble RAGE or genetic deletion of RAGE mitigated LPA-stimulated vascular Akt signaling, autotaxin/LPA-driven phosphorylation of Akt and cyclin D1 in the mammary tissue of transgenic mice vulnerable to carcinogenesis, and ovarian tumor implantation and development. These findings identify novel roles for RAGE as a conduit for LPA signaling and suggest targeting LPA–RAGE interaction as a therapeutic strategy to modify the pathological actions of LPA. PMID:23209312

  7. Beta Adrenergic Signaling: A Targetable Regulator of Angiosarcoma and Hemangiosarcoma

    Directory of Open Access Journals (Sweden)

    Erin B. Dickerson

    2015-09-01

    Full Text Available Human angiosarcomas and canine hemangiosarcomas are highly aggressive cancers thought to arise from cells of vascular origin. The pathological features, morphological organization, and clinical behavior of canine hemangiosarcomas are virtually indistinct from those of human angiosarcomas. Overall survival with current standard-of-care approaches remains dismal for both humans and dogs, and each is likely to succumb to their disease within a short duration. While angiosarcomas in humans are extremely rare, limiting their study and treatment options, canine hemangiosarcomas occur frequently. Therefore, studies of these sarcomas in dogs can be used to advance treatment approaches for both patient groups. Emerging data suggest that angiosarcomas and hemangiosarcomas utilize beta adrenergic signaling to drive their progression by regulating the tumor cell niche and fine-tuning cellular responses within the tumor microenvironment. These discoveries indicate that inhibition of beta adrenergic signaling could serve as an Achilles heel for these tumors and emphasize the need to design therapeutic strategies that target tumor cell and stromal cell constituents. In this review, we summarize recent discoveries and present new hypotheses regarding the roles of beta adrenergic signaling in angiosarcomas and hemangiosarcomas. Because the use of beta adrenergic receptor antagonists is well established in human and veterinary medicine, beta blockade could provide an immediate adjunct therapy for treatment along with a tangible opportunity to improve upon the outcomes of both humans and dogs with these diseases.

  8. Beta Adrenergic Signaling: A Targetable Regulator of Angiosarcoma and Hemangiosarcoma

    Science.gov (United States)

    Dickerson, Erin B.; Bryan, Brad A.

    2015-01-01

    Human angiosarcomas and canine hemangiosarcomas are highly aggressive cancers thought to arise from cells of vascular origin. The pathological features, morphological organization, and clinical behavior of canine hemangiosarcomas are virtually indistinct from those of human angiosarcomas. Overall survival with current standard-of-care approaches remains dismal for both humans and dogs, and each is likely to succumb to their disease within a short duration. While angiosarcomas in humans are extremely rare, limiting their study and treatment options, canine hemangiosarcomas occur frequently. Therefore, studies of these sarcomas in dogs can be used to advance treatment approaches for both patient groups. Emerging data suggest that angiosarcomas and hemangiosarcomas utilize beta adrenergic signaling to drive their progression by regulating the tumor cell niche and fine-tuning cellular responses within the tumor microenvironment. These discoveries indicate that inhibition of beta adrenergic signaling could serve as an Achilles heel for these tumors and emphasize the need to design therapeutic strategies that target tumor cell and stromal cell constituents. In this review, we summarize recent discoveries and present new hypotheses regarding the roles of beta adrenergic signaling in angiosarcomas and hemangiosarcomas. Because the use of beta adrenergic receptor antagonists is well established in human and veterinary medicine, beta blockade could provide an immediate adjunct therapy for treatment along with a tangible opportunity to improve upon the outcomes of both humans and dogs with these diseases. PMID:29061946

  9. FGF signaling inhibitor, SPRY4, is evolutionarily conserved target of WNT signaling pathway in progenitor cells.

    Science.gov (United States)

    Katoh, Yuriko; Katoh, Masaru

    2006-03-01

    WNT, FGF and Hedgehog signaling pathways network together during embryogenesis, tissue regeneration, and carcinogenesis. FGF16, FGF18, and FGF20 genes are targets of WNT-mediated TCF/LEF-beta-catenin-BCL9/BCL9L-PYGO transcriptional complex. SPROUTY (SPRY) and SPRED family genes encode inhibitors for receptor tyrosine kinase signaling cascades, such as those of FGF receptor family members and EGF receptor family members. Here, transcriptional regulation of SPRY1, SPRY2, SPRY3, SPRY4, SPRED1, SPRED2, and SPRED3 genes by WNT/beta-catenin signaling cascade was investigated by using bioinformatics and human intelligence (humint). Because double TCF/LEF-binding sites were identified within the 5'-promoter region of human SPRY4 gene, comparative genomics analyses on SPRY4 orthologs were further performed. SPRY4-FGF1 locus at human chromosome 5q31.3 and FGF2-NUDT6-SPATA5-SPRY1 locus at human chromosome 4q27-q28.1 were paralogous regions within the human genome. Chimpanzee SPRY4 gene was identified within NW_107083.1 genome sequence. Human, chimpanzee, rat and mouse SPRY4 orthologs, consisting of three exons, were well conserved. SPRY4 gene was identified as the evolutionarily conserved target of WNT/beta-catenin signaling pathway based on the conservation of double TCF/LEF-binding sites within 5'-promoter region of mammalian SPRY4 orthologs. Human SPRY4 mRNA was expressed in embryonic stem (ES) cells, brain, pancreatic islet, colon cancer, head and neck tumor, melanoma, and pancreatic cancer. WNT signaling activation in progenitor cells leads to the growth regulation of progenitor cells themselves through SPRY4 induction, and also to the growth stimulation of proliferating cells through FGF secretion. Epigenetic silencing and loss-of-function mutations of SPRY4 gene in progenitor cells could lead to carcinogenesis. SPRY4 is the pharmacogenomics target in the fields of oncology and regenerative medicine.

  10. Protein Sub-Nuclear Localization Based on Effective Fusion Representations and Dimension Reduction Algorithm LDA

    Directory of Open Access Journals (Sweden)

    Shunfang Wang

    2015-12-01

    Full Text Available An effective representation of a protein sequence plays a crucial role in protein sub-nuclear localization. The existing representations, such as dipeptide composition (DipC, pseudo-amino acid composition (PseAAC and position specific scoring matrix (PSSM, are insufficient to represent protein sequence due to their single perspectives. Thus, this paper proposes two fusion feature representations of DipPSSM and PseAAPSSM to integrate PSSM with DipC and PseAAC, respectively. When constructing each fusion representation, we introduce the balance factors to value the importance of its components. The optimal values of the balance factors are sought by genetic algorithm. Due to the high dimensionality of the proposed representations, linear discriminant analysis (LDA is used to find its important low dimensional structure, which is essential for classification and location prediction. The numerical experiments on two public datasets with KNN classifier and cross-validation tests showed that in terms of the common indexes of sensitivity, specificity, accuracy and MCC, the proposed fusing representations outperform the traditional representations in protein sub-nuclear localization, and the representation treated by LDA outperforms the untreated one.

  11. Protein Sub-Nuclear Localization Based on Effective Fusion Representations and Dimension Reduction Algorithm LDA.

    Science.gov (United States)

    Wang, Shunfang; Liu, Shuhui

    2015-12-19

    An effective representation of a protein sequence plays a crucial role in protein sub-nuclear localization. The existing representations, such as dipeptide composition (DipC), pseudo-amino acid composition (PseAAC) and position specific scoring matrix (PSSM), are insufficient to represent protein sequence due to their single perspectives. Thus, this paper proposes two fusion feature representations of DipPSSM and PseAAPSSM to integrate PSSM with DipC and PseAAC, respectively. When constructing each fusion representation, we introduce the balance factors to value the importance of its components. The optimal values of the balance factors are sought by genetic algorithm. Due to the high dimensionality of the proposed representations, linear discriminant analysis (LDA) is used to find its important low dimensional structure, which is essential for classification and location prediction. The numerical experiments on two public datasets with KNN classifier and cross-validation tests showed that in terms of the common indexes of sensitivity, specificity, accuracy and MCC, the proposed fusing representations outperform the traditional representations in protein sub-nuclear localization, and the representation treated by LDA outperforms the untreated one.

  12. Leptin signaling molecular actions and drug target in hepatocellular carcinoma

    Directory of Open Access Journals (Sweden)

    Jiang N

    2014-11-01

    Full Text Available Nan Jiang,1,* Rongtong Sun,2,* Qing Sun3 1Shandong University School of Medicine, Jinan, Shandong Province, People’s Republic of China; 2Weihai Municipal Hospital, Weihai, Shandong Province, People’s Republic of China; 3Department of Pathology, QianFoShan Hospital Affiliated to Shandong University, Jinan, Shandong Province, People’s Republic of China *These authors contributed equally to this work Abstract: Previous reports indicate that over 13 different tumors, including hepatocellular carcinoma (HCC, are related to obesity. Obesity-associated inflammatory, metabolic, and endocrine mediators, as well as the functioning of the gut microbiota, are suspected to contribute to tumorigenesis. In obese people, proinflammatory cytokines/chemokines including tumor necrosis factor-alpha, interleukin (IL-1 and IL-6, insulin and insulin-like growth factors, adipokines, plasminogen activator inhibitor-1, adiponectin, and leptin are found to play crucial roles in the initiation and development of cancer. The cytokines induced by leptin in adipose tissue or tumor cells have been intensely studied. Leptin-induced signaling pathways are critical for biological functions such as adiposity, energy balance, endocrine function, immune reaction, and angiogenesis as well as oncogenesis. Leptin is an activator of cell proliferation and anti-apoptosis in several cell types, and an inducer of cancer stem cells; its critical roles in tumorigenesis are based on its oncogenic, mitogenic, proinflammatory, and pro-angiogenic actions. This review provides an update of the pathological effects of leptin signaling with special emphasis on potential molecular mechanisms and therapeutic targeting, which could potentially be used in future clinical settings. In addition, leptin-induced angiogenic ability and molecular mechanisms in HCC are discussed. The stringent binding affinity of leptin and its receptor Ob-R, as well as the highly upregulated expression of both

  13. Endocannabinoid signaling in female reproductive events: a potential therapeutic target?

    Science.gov (United States)

    Maccarrone, Mauro

    2015-01-01

    Nearly 30 years after the discovery in 1964 of the psychoactive ingredient of cannabis (Cannabis sativa), Δ(9)-tetrahydrocannabinol, its endogenous counterparts were discovered and collectively termed endocannabinoids (eCBs): N-arachidonoylethanolamine (anandamide) in 1992 and 2-arachidonoylglycerol in 1995. Since then, intense research has identified additional eCBs and an ensemble of proteins that bind, synthesize and degrade them, the so-called eCB system. Altogether, these new compounds have been recognized as key mediators of several aspects of human pathophysiology, and in particular of female fertility. Here, the main features of the eCB system are presented, in order to put in a better perspective the relevance of eCB signaling in virtually all steps of human reproduction and to highlight emerging hopes that elements of this system might indeed become novel targets to combat fertility problems.

  14. Regression of Pathological Cardiac Hypertrophy: Signaling Pathways and Therapeutic Targets

    Science.gov (United States)

    Hou, Jianglong; Kang, Y. James

    2012-01-01

    Pathological cardiac hypertrophy is a key risk factor for heart failure. It is associated with increased interstitial fibrosis, cell death and cardiac dysfunction. The progression of pathological cardiac hypertrophy has long been considered as irreversible. However, recent clinical observations and experimental studies have produced evidence showing the reversal of pathological cardiac hypertrophy. Left ventricle assist devices used in heart failure patients for bridging to transplantation not only improve peripheral circulation but also often cause reverse remodeling of the geometry and recovery of the function of the heart. Dietary supplementation with physiologically relevant levels of copper can reverse pathological cardiac hypertrophy in mice. Angiogenesis is essential and vascular endothelial growth factor (VEGF) is a constitutive factor for the regression. The action of VEGF is mediated by VEGF receptor-1, whose activation is linked to cyclic GMP-dependent protein kinase-1 (PKG-1) signaling pathways, and inhibition of cyclic GMP degradation leads to regression of pathological cardiac hypertrophy. Most of these pathways are regulated by hypoxia-inducible factor. Potential therapeutic targets for promoting the regression include: promotion of angiogenesis, selective enhancement of VEGF receptor-1 signaling pathways, stimulation of PKG-1 pathways, and sustention of hypoxia-inducible factor transcriptional activity. More exciting insights into the regression of pathological cardiac hypertrophy are emerging. The time of translating the concept of regression of pathological cardiac hypertrophy to clinical practice is coming. PMID:22750195

  15. Signal Transduction and Molecular Targets of Selected Flavonoids

    Science.gov (United States)

    Bode, Ann M.

    2013-01-01

    Abstract Significance: Diet exerts a major influence on the risk for developing cancer and heart disease. Food factors such as flavonoids are alleged to protect cells from premature aging and disease by shielding DNA, proteins, and lipids from oxidative damage. Recent Advances: Our work has focused on clarifying the effects of dietary components on cancer cell proliferation and tumor growth, discovering mechanisms to explain the effects, and identifying the specific molecular targets of these compounds. Our strategy for identifying specific molecular targets of phytochemicals involves the use of supercomputer technology combined with protein crystallography, molecular biology, and experimental laboratory verification. Critical Issues: One of the greatest challenges for scientists is to reduce the accumulation of distortion and half truths reported in the popular media regarding the health benefits of certain foods or food supplements. The use of these is not new, but interest has increased dramatically because of perceived health benefits that are presumably acquired without unpleasant side effects. Flavonoids are touted to exert many beneficial effects in vitro. However, whether they can produce these effects in vivo is disputed. Future Directions: The World Health Organization indicates that one third of all cancer deaths are preventable and that diet is closely linked to prevention. Based on this idea and epidemiological findings, attention has centered on dietary phytochemicals as an effective intervention in cancer development. However, an unequivocal link between diet and cancer has not been established. Thus, identifying cancer preventive dietary agents with specific molecular targets is essential to move forward toward successful cancer prevention. Antioxid. Redox Signal. 19, 163–180. PMID:23458437

  16. GABAergic signaling as therapeutic target for Autism Spectrum Disorders

    Directory of Open Access Journals (Sweden)

    Giada eCellot

    2014-07-01

    Full Text Available GABA, the main inhibitory neurotransmitter in the adult brain, early in postnatal life exerts a depolarizing and excitatory action. This depends on accumulation of chloride inside the cell via the cation-chloride importer NKCC1, being the expression of the chloride exporter KCC2 very low at birth. The developmentally regulated expression of KCC2 results in extrusion of chloride with age and a shift of GABA from the depolarizing to the hyperpolarizing direction. The depolarizing action of GABA leads to intracellular calcium rise through voltage-dependent calcium channels and/or NMDA receptors. GABA-mediated calcium signals regulate a variety of developmental processes from cell proliferation migration, differentiation, synapse maturation and neuronal wiring. Therefore, it is not surprising that some forms of neuro-developmental disorders such as Autism Spectrum Disorders (ASDs are associated with alterations of GABAergic signaling and impairment of the excitatory/inhibitory balance in selective neuronal circuits. In this review we will discuss how changes of GABAA-mediated neurotransmission affect several forms of ASDs including the Fragile X, the Angelman and Rett syndromes. Then, we will describe various animal models of ASDs with GABAergic dysfunctions, highlighting their behavioral deficits and the possibility to rescue them by targeting selective components of the GABAergic synapse. In particular, we will discuss how in some cases, reverting the polarity of GABA responses from the depolarizing to the hyperpolarizing direction with the diuretic bumetanide, a selective blocker of NKCC1, may have beneficial effects on ASDs, thus opening new therapeutic perspectives for the treatment of these devastating disorders.

  17. Dynamical self-consistent description of exotic structures in nuclear matter at subnuclear densities

    CERN Document Server

    de la Mota, Virginia; Figerou, Sébastien

    2010-01-01

    We investigate the occurrence of exotic structures in nuclear matter at subnuclear densities within the framework of the dywan model. This approach, developed ab initio for the description of nuclear collisions, is a microscopic dynamical approach in which the numerical treatment makes use of wavelet representation techniques. Before tackling the effects of multi-particle correlations on the overall dynamics, we focused the present work on the study of cold matter within a pure mean field description. Starting from inhomogeneous initial conditions provided by an arrangement of nuclei located on an initial crystalline lattice, the exotic structures result from the dynamical self-consistent evolution. The nuclear system can freely self-organize, it can modify or even break the lattice structure and the initial symmetries of matter distribution. This approach goes beyond the Wigner-Seitz approximation and no assumption of final shapes of matter is made. In this framework, different effects, as the sensitivity of...

  18. Targeting signaling pathways in acute lymphoblastic leukemia: new insights.

    Science.gov (United States)

    Harrison, Christine J

    2013-01-01

    The genetics of acute lymphoblastic leukemia are becoming well understood and the incidence of individual chromosomal abnormalities varies considerably with age. Cytogenetics provide reliable risk stratification for treatment: high hyperdiploidy and ETV6-RUNX1 are good risk, whereas BCR-ABL1, MLL rearrangements, and hypodiploidy are poor risk. Nevertheless, some patients within the good- and intermediate-risk groups will unpredictably relapse. With advancing technologies in array-based approaches (single nucleotide polymorphism arrays) and next-generation sequencing to study the genome, increasing numbers of new genetic changes are being discovered. These include deletions of B-cell differentiation and cell cycle control genes, as well as mutations of genes in key signaling pathways. Their associations and interactions with established cytogenetic subgroups and with each other are becoming elucidated. Whether they have a link to outcome is the most important factor for refinement of risk factors in relation to clinical trials. For several newly identified abnormalities, including intrachromosomal amplification of chromosome 21 (iAMP21), that are associated with a poor prognosis with standard therapy, appropriately modified treatment has significantly improved outcome. After the successful use of tyrosine kinase inhibitors in the treatment of BCR-ABL1-positive acute lymphoblastic leukemia, patients with alternative ABL1 translocations and rearrangements involving PDGFRB may benefit from treatment with tyrosine kinase inhibitors. Other aberrations, for example, CRLF2 overexpression and JAK2 mutations, are also providing potential novel therapeutic targets with the prospect of reduced toxicity.

  19. Visualization by BiFC of different C/EBP{beta} dimers and their interaction with HP1{alpha} reveals a differential subnuclear distribution of complexes in living cells

    Energy Technology Data Exchange (ETDEWEB)

    Susperreguy, Sebastian; Prendes, Luciana P.; Desbats, Maria A.; Charo, Nancy L. [Instituto de Biologia y Medicina Experimental, CONICET, Buenos Aires (Argentina); Brown, Karen [Chromosome Biology Group, Imperial College of London, London (United Kingdom); MacDougald, Ormond A. [Dept. of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI (United States); Kerppola, Tom [Dept. of Biological Chemistry, University of Michigan Medical School, Ann Arbor, MI (United States); Schwartz, Jessica [Dept. of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI (United States); Piwien-Pilipuk, Graciela, E-mail: gpiwien@conicet.gov.ar [Instituto de Biologia y Medicina Experimental, CONICET, Buenos Aires (Argentina)

    2011-04-01

    How the co-ordinated events of gene activation and silencing during cellular differentiation are influenced by spatial organization of the cell nucleus is still poorly understood. Little is known about the molecular mechanisms controlling subnuclear distribution of transcription factors, and their interplay with nuclear proteins that shape chromatin structure. Here we show that C/EBP{beta} not only associates with pericentromeric heterochromatin but also interacts with the nucleoskeleton upon induction of adipocyte differentiation of 3T3-L1 cells. Different C/EBP{beta} dimers localize in different nuclear domains. Using BiFC in living cells, we show that LAP (Liver Activating Protein) homodimers localize in euchromatin and heterochromatin. In contrast, LIP (Liver Inhibitory Protein) homodimers localize exclusively in heterochromatin. Importantly, their differential subnuclear distribution mirrors the site for interaction with HP1{alpha}. HP1{alpha} inhibits LAP transcriptional capacity and occupies the promoter of the C/EBP{beta}-dependent gene c/ebp{alpha} in 3T3-L1 preadipocytes. When adipogenesis is induced, HP1{alpha} binding decreases from c/ebp{alpha} promoter, allowing transcription. Thus, the equilibrium among different pools of C/EBP{beta} associated with chromatin or nucleoskeleton, and dynamic changes in their interaction with HP1{alpha}, play key roles in the regulation of C/EBP target genes during adipogenesis.

  20. Extended model of raw data signals for space-time adaptive processing and moving target indicators

    NARCIS (Netherlands)

    Lidický, L.; Hoogeboom, P.

    2006-01-01

    This paper presents a multi-channel synthetic aperture radar (SAR) signal model. Its purpose is to study nonstationary raw SAR signals in moving target indication (MTI) systems. This is opposed to the traditional approach in which only stationary (harmonic) signals are considered. The principal

  1. Notch signaling as a therapeutic target for breast cancer treatment?

    Science.gov (United States)

    Han, Jianxun; Hendzel, Michael J; Allalunis-Turner, Joan

    2011-05-31

    Aberrant Notch signaling can induce mammary gland carcinoma in transgenic mice, and high expressions of Notch receptors and ligands have been linked to poor clinical outcomes in human patients with breast cancer. This suggests that inhibition of Notch signaling may be beneficial for breast cancer treatment. In this review, we critically evaluate the evidence that supports or challenges the hypothesis that inhibition of Notch signaling would be advantageous in breast cancer management. We find that there are many remaining uncertainties that must be addressed experimentally if we are to exploit inhibition of Notch signaling as a treatment approach in breast cancer. Nonetheless, Notch inhibition, in combination with other therapies, is a promising avenue for future management of breast cancer. Furthermore, since aberrant Notch4 activity can induce mammary gland carcinoma in the absence of RBPjκ, a better understanding of the components of RBPjκ-independent oncogenic Notch signaling pathways and their contribution to Notch-induced tumorigenesis would facilitate the deployment of Notch inhibition strategies for effective treatment of breast cancer.

  2. Variation of the Nuclear, Subnuclear and Chromosomal Flavanol Deposition in Hemlock and Rye

    Directory of Open Access Journals (Sweden)

    Jürgen Polster

    2007-07-01

    Full Text Available Nuclei of hemlock (Tsuga canadensis and Tsuga canadensis var. nana wereinvestigated for the presence of flavanols. Histochemical staining with p-dimethylaminocinnamaldehyde proved to be a highly valuable method yielding a brightblue flavanol coloration for nuclei. There was a significant variation in flavanol deposition(1 among nuclei, (2 at the subnuclear level and also (3 along the chromosomes duringmitosis. The presence of flavanols in nucleoli could not be established probably becausethey were too small, measuring less than 1 μm in diameter. In contrast to Tsuga, the cellsand nuclei of rootlets from rye (Secale cereale were totally devoid of natural flavanols.However, externally added flavanols, catechin and epicatechin, were bound to the ryenuclei, while the rather large nucleoli failed to associate with the flavanols. The strong sinkactivity of nucleoplasm and chromosomes for flavanols in Tsuga and Secale indicates aprocess which is apparently widespread even in distantly related plant species. Variationsin chromatin-associated flavanols could to some extent be induced byacetylation/deacetylation of histones, as confirmed in the present study by means of UV-VIS spectroscopic titrations of histone sulphate and chemically acetylated histone sulphate.

  3. Variation of the Nuclear, Subnuclear and Chromosomal Flavanol Deposition in Hemlock and Rye

    Science.gov (United States)

    Feucht, Walter; Dithmar, Heike; Polster, Jürgen

    2007-01-01

    Nuclei of hemlock (Tsuga canadensis and Tsuga canadensis var. nana) were investigated for the presence of flavanols. Histochemical staining with p-dimethylaminocinnamaldehyde proved to be a highly valuable method yielding a bright blue flavanol coloration for nuclei. There was a significant variation in flavanol deposition (1) among nuclei, (2) at the subnuclear level and also (3) along the chromosomes during mitosis. The presence of flavanols in nucleoli could not be established probably because they were too small, measuring less than 1 μm in diameter. In contrast to Tsuga, the cells and nuclei of rootlets from rye (Secale cereale) were totally devoid of natural flavanols. However, externally added flavanols, catechin and epicatechin, were bound to the rye nuclei, while the rather large nucleoli failed to associate with the flavanols. The strong sink activity of nucleoplasm and chromosomes for flavanols in Tsuga and Secale indicates a process which is apparently widespread even in distantly related plant species. Variations in chromatin-associated flavanols could to some extent be induced by acetylation/deacetylation of histones, as confirmed in the present study by means of UVVIS spectroscopic titrations of histone sulphate and chemically acetylated histone sulphate.

  4. Target Doppler Estimation Using Wideband Frequency Modulated Signals

    NARCIS (Netherlands)

    Doisy, Y.; Deruaz, L.; Beerens, S.P.; Been, R.

    2000-01-01

    The topic of this paper is the design and performance analysis of wideband sonar waveforms capable of estimating both target range and Döppler using as few replicas in the processing as possible. First, it is shown that for conventional Döppler sensitive waveforms, for which the Döppler and delay

  5. Mitochondria targeting by environmental stressors: Implications for redox cellular signaling.

    Science.gov (United States)

    Blajszczak, Chuck; Bonini, Marcelo G

    2017-11-01

    Mitochondria are cellular powerhouses as well as metabolic and signaling hubs regulating diverse cellular functions, from basic physiology to phenotypic fate determination. It is widely accepted that reactive oxygen species (ROS) generated in mitochondria participate in the regulation of cellular signaling, and that some mitochondria chronically operate at a high ROS baseline. However, it is not completely understood how mitochondria adapt to persistently high ROS states and to environmental stressors that disturb the redox balance. Here we will review some of the current concepts regarding how mitochondria resist oxidative damage, how they are replaced when excessive oxidative damage compromises function, and the effect of environmental toxicants (i.e. heavy metals) on the regulation of mitochondrial ROS (mtROS) production and subsequent impact. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Direct targets of pSTAT5 signalling in erythropoiesis.

    Directory of Open Access Journals (Sweden)

    Kevin R Gillinder

    Full Text Available Erythropoietin (EPO acts through the dimeric erythropoietin receptor to stimulate proliferation, survival, differentiation and enucleation of erythroid progenitor cells. We undertook two complimentary approaches to find EPO-dependent pSTAT5 target genes in murine erythroid cells: RNA-seq of newly transcribed (4sU-labelled RNA, and ChIP-seq for pSTAT5 30 minutes after EPO stimulation. We found 302 pSTAT5-occupied sites: ~15% of these reside in promoters while the rest reside within intronic enhancers or intergenic regions, some >100kb from the nearest TSS. The majority of pSTAT5 peaks contain a central palindromic GAS element, TTCYXRGAA. There was significant enrichment for GATA motifs and CACCC-box motifs within the neighbourhood of pSTAT5-bound peaks, and GATA1 and/or KLF1 co-occupancy at many sites. Using 4sU-RNA-seq we determined the EPO-induced transcriptome and validated differentially expressed genes using dynamic CAGE data and qRT-PCR. We identified known direct pSTAT5 target genes such as Bcl2l1, Pim1 and Cish, and many new targets likely to be involved in driving erythroid cell differentiation including those involved in mRNA splicing (Rbm25, epigenetic regulation (Suv420h2, and EpoR turnover (Clint1/EpsinR. Some of these new EpoR-JAK2-pSTAT5 target genes could be used as biomarkers for monitoring disease activity in polycythaemia vera, and for monitoring responses to JAK inhibitors.

  7. High-Risk Human Papillomavirus Targets Crossroads in Immune Signaling

    Science.gov (United States)

    Tummers, Bart; Van Der Burg, Sjoerd H.

    2015-01-01

    Persistent infections with a high-risk type human papillomavirus (hrHPV) can progress to cancer. High-risk HPVs infect keratinocytes (KCs) and successfully suppress host immunity for up to two years despite the fact that KCs are well equipped to detect and initiate immune responses to invading pathogens. Viral persistence is achieved by active interference with KCs innate and adaptive immune mechanisms. To this end hrHPV utilizes proteins encoded by its viral genome, as well as exploits cellular proteins to interfere with signaling of innate and adaptive immune pathways. This results in impairment of interferon and pro-inflammatory cytokine production and subsequent immune cell attraction, as well as resistance to incoming signals from the immune system. Furthermore, hrHPV avoids the killing of infected cells by interfering with antigen presentation to antigen-specific cytotoxic T lymphocytes. Thus, hrHPV has evolved multiple mechanisms to avoid detection and clearance by both the innate and adaptive immune system, the molecular mechanisms of which will be dealt with in detail in this review. PMID:26008697

  8. High-Risk Human Papillomavirus Targets Crossroads in Immune Signaling

    Directory of Open Access Journals (Sweden)

    Bart Tummers

    2015-05-01

    Full Text Available Persistent infections with a high-risk type human papillomavirus (hrHPV can progress to cancer. High-risk HPVs infect keratinocytes (KCs and successfully suppress host immunity for up to two years despite the fact that KCs are well equipped to detect and initiate immune responses to invading pathogens. Viral persistence is achieved by active interference with KCs innate and adaptive immune mechanisms. To this end hrHPV utilizes proteins encoded by its viral genome, as well as exploits cellular proteins to interfere with signaling of innate and adaptive immune pathways. This results in impairment of interferon and pro-inflammatory cytokine production and subsequent immune cell attraction, as well as resistance to incoming signals from the immune system. Furthermore, hrHPV avoids the killing of infected cells by interfering with antigen presentation to antigen-specific cytotoxic T lymphocytes. Thus, hrHPV has evolved multiple mechanisms to avoid detection and clearance by both the innate and adaptive immune system, the molecular mechanisms of which will be dealt with in detail in this review.

  9. High-risk human papillomavirus targets crossroads in immune signaling.

    Science.gov (United States)

    Tummers, Bart; Burg, Sjoerd H Van Der

    2015-05-21

    Persistent infections with a high-risk type human papillomavirus (hrHPV) can progress to cancer. High-risk HPVs infect keratinocytes (KCs) and successfully suppress host immunity for up to two years despite the fact that KCs are well equipped to detect and initiate immune responses to invading pathogens. Viral persistence is achieved by active interference with KCs innate and adaptive immune mechanisms. To this end hrHPV utilizes proteins encoded by its viral genome, as well as exploits cellular proteins to interfere with signaling of innate and adaptive immune pathways. This results in impairment of interferon and pro-inflammatory cytokine production and subsequent immune cell attraction, as well as resistance to incoming signals from the immune system. Furthermore, hrHPV avoids the killing of infected cells by interfering with antigen presentation to antigen-specific cytotoxic T lymphocytes. Thus, hrHPV has evolved multiple mechanisms to avoid detection and clearance by both the innate and adaptive immune system, the molecular mechanisms of which will be dealt with in detail in this review.

  10. Combination therapy approaches to target insulin-like growth factor receptor signaling in breast cancer.

    Science.gov (United States)

    Ochnik, Aleksandra M; Baxter, Robert C

    2016-11-01

    Insulin-like growth factor receptor (IGF1R) signaling as a therapeutic target has been widely studied and clinically tested. Despite the vast amount of literature supporting the biological role of IGF1R in breast cancer, effective clinical translation in targeting its activity as a cancer therapy has not been successful. The intrinsic complexity of cancer cell signaling mediated by many tyrosine kinase growth factor receptors that work together to modulate each other and intracellular downstream mediators in the cell highlights that studying IGF1R expression and activity as a prognostic factor and therapeutic target in isolation is certainly associated with problems. This review discusses the current literature and clinical trials associated with IGF-1 signaling and attempts to look at new ways of designing novel IGF1R-directed breast cancer therapy approaches to target its activity 
and/or intracellular downstream signaling pathways in IGF1R-expressing breast cancers. © 2016 Society for Endocrinology.

  11. A Review: Phytochemicals Targeting JAK/STAT Signaling and IDO Expression in Cancer.

    Science.gov (United States)

    Arumuggam, Niroshaathevi; Bhowmick, Neil A; Rupasinghe, H P Vasantha

    2015-06-01

    Cancer remains a major health problem worldwide. Among many other factors, two regulatory defects that are present in most cancer cells are constitutive activation of Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling pathway and the induction of indoleamine 2, 3-dioxygenase (IDO), an enzyme that catalyzes tryptophan degradation, through JAK/STAT signaling. Cytokine signaling activates STAT proteins in regulating cell proliferation, differentiation, and survival through modulation of target genes. Many phytochemicals can inhibit both JAK/STAT signaling and IDO expression in antigen-presenting cells by targeting different pathways. Some of the promising phytochemicals that are discussed in this review include resveratrol, cucurbitacin, curcumin, (-)-epigallocatechin gallate, and others. It is now evident that phytochemicals play key roles in inhibition of tumor proliferation and development and provide novel means for therapeutic targeting of cancer. Copyright © 2015 John Wiley & Sons, Ltd.

  12. Leptin signaling as a therapeutic target of obesity.

    Science.gov (United States)

    Sáinz, Neira; González-Navarro, Carlos J; Martínez, J Alfredo; Moreno-Aliaga, Maria J

    2015-07-01

    Leptin is a hormone with a key role in food intake and body weight homeostasis. Congenital leptin deficiency (CLD) is a rare disease that causes hyperphagia and early severe obesity. However, common obesity conditions are associated with hyperleptinemia and leptin resistance. The main signaling pathways activated by leptin as well as the mechanisms underlying the regulatory actions of leptin on food intake and on lipid and glucose metabolism are reviewed. The potential mechanisms involving leptin resistance and the main regulatory hormonal and nutritional factors controlling leptin production/functions are also analyzed. The pathophysiology of leptin in human obesity, and especially the trials analyzing effects of leptin replacement therapy in patients with CLD or in subjects with common obesity and in post-obese weight-reduced subjects are also summarized. The use of drugs or specific bioactive food components with anti-inflammatory properties to reduce the inflammatory state associated with obesity, especially at the hypothalamus, may help to overcome leptin resistance. Research should also be focused on investigating dietary strategies, food supplements or drugs capable of avoiding or reversing the leptin fall during weight management, in order to promote sustained body weight lowering and weight loss maintenance.

  13. Targeting FLT3 Signaling in Childhood Acute Myeloid Leukemia

    Directory of Open Access Journals (Sweden)

    Amy N. Sexauer

    2017-11-01

    Full Text Available Acute myeloid leukemia (AML is the second most common leukemia of childhood and is associated with high rates of chemotherapy resistance and relapse. Clinical outcomes for children with AML treated with maximally intensive multi-agent chemotherapy lag far behind those of children with the more common acute lymphoblastic leukemia, demonstrating continued need for new therapeutic approaches to decrease relapse risk and improve long-term survival. Mutations in the FMS-like tyrosine kinase-3 receptor gene (FLT3 occur in approximately 25% of children and adults with AML and are associated with particularly poor prognoses. Identification and development of targeted FLT3 inhibitors represents a major precision medicine paradigm shift in the treatment of patients with AML. While further development of many first-generation FLT3 inhibitors was hampered by limited potency and significant toxicity due to effects upon other kinases, the more selective second- and third-generation FLT3 inhibitors have demonstrated excellent tolerability and remarkable efficacy in the relapsed/refractory and now de novo FLT3-mutated AML settings. While these newest and most promising inhibitors have largely been studied in the adult population, pediatric investigation of FLT3 inhibitors with chemotherapy is relatively recently ongoing or planned. Successful development of FLT3 inhibitor-based therapies will be essential to improve outcomes in children with this high-risk subtype of AML.

  14. Activation of Smurf E3 ligase promoted by smoothened regulates hedgehog signaling through targeting patched turnover.

    Directory of Open Access Journals (Sweden)

    Shoujun Huang

    2013-11-01

    Full Text Available Hedgehog signaling plays conserved roles in controlling embryonic development; its dysregulation has been implicated in many human diseases including cancers. Hedgehog signaling has an unusual reception system consisting of two transmembrane proteins, Patched receptor and Smoothened signal transducer. Although activation of Smoothened and its downstream signal transduction have been intensively studied, less is known about how Patched receptor is regulated, and particularly how this regulation contributes to appropriate Hedgehog signal transduction. Here we identified a novel role of Smurf E3 ligase in regulating Hedgehog signaling by controlling Patched ubiquitination and turnover. Moreover, we showed that Smurf-mediated Patched ubiquitination depends on Smo activity in wing discs. Mechanistically, we found that Smo interacts with Smurf and promotes it to mediate Patched ubiquitination by targeting the K1261 site in Ptc. The further mathematic modeling analysis reveals that a bidirectional control of activation of Smo involving Smurf and Patched is important for signal-receiving cells to precisely interpret external signals, thereby maintaining Hedgehog signaling reliability. Finally, our data revealed an evolutionarily conserved role of Smurf proteins in controlling Hh signaling by targeting Ptc during development.

  15. Synergistic target combination prediction from curated signaling networks: Machine learning meets systems biology and pharmacology.

    Science.gov (United States)

    Chua, Huey Eng; Bhowmick, Sourav S; Tucker-Kellogg, Lisa

    2017-10-01

    Given a signaling network, the target combination prediction problem aims to predict efficacious and safe target combinations for combination therapy. State-of-the-art in silico methods use Monte Carlo simulated annealing (mcsa) to modify a candidate solution stochastically, and use the Metropolis criterion to accept or reject the proposed modifications. However, such stochastic modifications ignore the impact of the choice of targets and their activities on the combination's therapeutic effect and off-target effects, which directly affect the solution quality. In this paper, we present mascot, a method that addresses this limitation by leveraging two additional heuristic criteria to minimize off-target effects and achieve synergy for candidate modification. Specifically, off-target effects measure the unintended response of a signaling network to the target combination and is often associated with toxicity. Synergy occurs when a pair of targets exerts effects that are greater than the sum of their individual effects, and is generally a beneficial strategy for maximizing effect while minimizing toxicity. mascot leverages on a machine learning-based target prioritization method which prioritizes potential targets in a given disease-associated network to select more effective targets (better therapeutic effect and/or lower off-target effects); and on Loewe additivity theory from pharmacology which assesses the non-additive effects in a combination drug treatment to select synergistic target activities. Our experimental study on two disease-related signaling networks demonstrates the superiority of mascot in comparison to existing approaches. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Identification of Potential Drug Targets in Cancer Signaling Pathways using Stochastic Logical Models.

    Science.gov (United States)

    Zhu, Peican; Aliabadi, Hamidreza Montazeri; Uludağ, Hasan; Han, Jie

    2016-03-18

    The investigation of vulnerable components in a signaling pathway can contribute to development of drug therapy addressing aberrations in that pathway. Here, an original signaling pathway is derived from the published literature on breast cancer models. New stochastic logical models are then developed to analyze the vulnerability of the components in multiple signalling sub-pathways involved in this signaling cascade. The computational results are consistent with the experimental results, where the selected proteins were silenced using specific siRNAs and the viability of the cells were analyzed 72 hours after silencing. The genes elF4E and NFkB are found to have nearly no effect on the relative cell viability and the genes JAK2, Stat3, S6K, JUN, FOS, Myc, and Mcl1 are effective candidates to influence the relative cell growth. The vulnerabilities of some targets such as Myc and S6K are found to vary significantly depending on the weights of the sub-pathways; this will be indicative of the chosen target to require customization for therapy. When these targets are utilized, the response of breast cancers from different patients will be highly variable because of the known heterogeneities in signaling pathways among the patients. The targets whose vulnerabilities are invariably high might be more universally acceptable targets.

  17. Nuclease-free target recycling signal amplification for ultrasensitive multiplexing DNA biosensing.

    Science.gov (United States)

    Zhao, Zhihan; Chen, Shixing; Wang, Jianbang; Su, Jing; Xu, Jiaqiang; Mathur, Sanjay; Fan, Chunhai; Song, Shiping

    2017-08-15

    Ultrasensitive biosensing technologies without gene amplification held great promise for direct detection of DNA. Herein we report a novel biosensing method, combining target recycling signal-amplification strategy and a homemade electrochemical device. Especially, the target recycling was achieved by a strand displacement process, no needing the help of any nucleases. In the presence of target DNA, the recycling system could be activated to generate a cascade of assembly steps with three hairpin DNA segments. Each recycling process were accompanied by a disassembly step that the last hairpin DNA segment displaces target DNA from the complex at the end of each circulation, freeing targets to activate the self-assembly of more trefoil DNA structures. This biosensing method could detect target DNA at aM level and can distinguish target DNA from interfering DNAs, demonstrating its high sensitivity and high selectivity. Importantly, the biosensing method could work well with serum samples. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Sensitivity analysis of intracellular signaling pathway kinetics predicts targets for stem cell fate control.

    Directory of Open Access Journals (Sweden)

    Alborz Mahdavi

    2007-07-01

    Full Text Available Directing stem cell fate requires knowledge of how signaling networks integrate temporally and spatially segregated stimuli. We developed and validated a computational model of signal transducer and activator of transcription-3 (Stat3 pathway kinetics, a signaling network involved in embryonic stem cell (ESC self-renewal. Our analysis identified novel pathway responses; for example, overexpression of the receptor glycoprotein-130 results in reduced pathway activation and increased ESC differentiation. We used a systematic in silico screen to identify novel targets and protein interactions involved in Stat3 activation. Our analysis demonstrates that signaling activation and desensitization (the inability to respond to ligand restimulation is regulated by balancing the activation state of a distributed set of parameters including nuclear export of Stat3, nuclear phosphatase activity, inhibition by suppressor of cytokine signaling, and receptor trafficking. This knowledge was used to devise a temporally modulated ligand delivery strategy that maximizes signaling activation and leads to enhanced ESC self-renewal.

  19. The cornerstone K-RAS mutation in pancreatic adenocarcinoma: From cell signaling network, target genes, biological processes to therapeutic targeting.

    Science.gov (United States)

    Jonckheere, Nicolas; Vasseur, Romain; Van Seuningen, Isabelle

    2017-03-01

    RAS belongs to the super family of small G proteins and plays crucial roles in signal transduction from membrane receptors in the cell. Mutations of K-RAS oncogene lead to an accumulation of GTP-bound proteins that maintains an active conformation. In the pancreatic ductal adenocarcinoma (PDAC), one of the most deadly cancers in occidental countries, mutations of the K-RAS oncogene are nearly systematic (>90%). Moreover, K-RAS mutation is the earliest genetic alteration occurring during pancreatic carcinogenetic sequence. In this review, we discuss the central role of K-RAS mutations and their tremendous diversity of biological properties by the interconnected regulation of signaling pathways (MAPKs, NF-κB, PI3K, Ral…). In pancreatic ductal adenocarcinoma, transcriptome analysis and preclinical animal models showed that K-RAS mutation alters biological behavior of PDAC cells (promoting proliferation, migration and invasion, evading growth suppressors, regulating mucin pattern, and miRNA expression). K-RAS also impacts tumor microenvironment and PDAC metabolism reprogramming. Finally we discuss therapeutic targeting strategies of K-RAS that have been developed without significant clinical success so far. As K-RAS is considered as the undruggable target, targeting its multiple effectors and target genes should be considered as potential alternatives. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Advancement of Wnt signal pathway and the target of breast cancer

    Directory of Open Access Journals (Sweden)

    Liang Quan

    2016-01-01

    Full Text Available Wnt/β-catenin signaling has been proved to play an important role in the development and promotion of cancer metastasis. The activation of Wnt signals can lead to duplicating, updating, metastasizing and relapsing. The Wnt signaling pathway is mainly divided into the Wnt/β-catenin pathway and the Wnt/calcium pathway. A better understanding of all the diverse functions of Wnt and their molecular mechanisms has evoked prevailing interest in identifying additional targets related to the Wnt /β-catenin pathways in breast cancer. A number of new target, related to Wnt /β-catenin pathways have been identified in recent years, including NOP14, BKCa channels, Emilin2, WISP, MicroRNAs, NRBP1, TRAF4, and Wntless. In this review, we will introduce the new targets related to the Wnt /β-catenin pathways in breast cancer.

  1. Targeting Ras signaling in AML: RALB is a small GTPase with big potential.

    Science.gov (United States)

    Pomeroy, Emily J; Eckfeldt, Craig E

    2017-07-06

    Acute myeloid leukemia (AML) is a devastating malignancy for which novel treatment approaches are desperately needed. Ras signaling is an attractive therapeutic target for AML because a large proportion of AMLs have mutations in NRAS, KRAS, or genes that activate Ras signaling, and key Ras effectors are activated in virtually all AML patient samples. This has inspired efforts to develop Ras-targeted treatment strategies for AML. Due to the inherent difficulty and disappointing efficacy of targeting Ras proteins directly, many have focused on inhibiting Ras effector pathways. Inhibiting the major oncogenic Ras effectors, the mitogen-activated protein kinase (MAPK) and/or phosphatidylinositiol-3-kinase (PI3K) pathways, has generally demonstrated modest efficacy for AML. While this may be in part related to functional redundancy between these pathways, it is now clear that other Ras effectors have key oncogenic roles. Specifically, the Ras-like (Ral) GTPases have emerged as critical mediators of Ras-driven transformation and AML cell survival. Our group recently uncovered a critical role for RALB signaling in leukemic cell survival and a potential mediator of relapse following Ras-targeted therapy in AML. Furthermore, we found that RALB signaling is hyperactivated in AML patient samples, and inhibiting RALB has potent anti-leukemic activity in preclinical AML models. While key questions remain regarding the importance of RALB signaling across the genetically diverse spectrum of AML, the specific mechanism(s) that promotes leukemic cell survival downstream of RALB, and how to pharmacologically target RALB signaling effectively - RALB has emerged as a critical Ras effector and potential therapeutic target for AML.

  2. HER2-family signalling mechanisms, clinical implications and targeting in breast cancer.

    OpenAIRE

    Elster, N; Collins, Denis M; Toomey, Sinead; Crown, John; Eustace, Alex J; Hennessy, Bryan T.

    2015-01-01

    Approximately 20 % of human breast cancers (BC) overexpress HER2 protein, and HER2-positivity is associated with a worse prognosis. Although HER2-targeted therapies have significantly improved outcomes for HER2-positive BC patients, resistance to trastuzumab-based therapy remains a clinical problem. In order to better understand resistance to HER2-targeted therapies in HER2-positive BC, it is necessary to examine HER family signalling as a whole. An extensive literature search was carried out...

  3. Using Chou's pseudo amino acid composition based on approximate entropy and an ensemble of AdaBoost classifiers to predict protein subnuclear location.

    Science.gov (United States)

    Jiang, Xiaoying; Wei, Rong; Zhao, Yanjun; Zhang, Tongliang

    2008-05-01

    The knowledge of subnuclear localization in eukaryotic cells is essential for understanding the life function of nucleus. Developing prediction methods and tools for proteins subnuclear localization become important research fields in protein science for special characteristics in cell nuclear. In this study, a novel approach has been proposed to predict protein subnuclear localization. Sample of protein is represented by Pseudo Amino Acid (PseAA) composition based on approximate entropy (ApEn) concept, which reflects the complexity of time series. A novel ensemble classifier is designed incorporating three AdaBoost classifiers. The base classifier algorithms in three AdaBoost are decision stumps, fuzzy K nearest neighbors classifier, and radial basis-support vector machines, respectively. Different PseAA compositions are used as input data of different AdaBoost classifier in ensemble. Genetic algorithm is used to optimize the dimension and weight factor of PseAA composition. Two datasets often used in published works are used to validate the performance of the proposed approach. The obtained results of Jackknife cross-validation test are higher and more balance than them of other methods on same datasets. The promising results indicate that the proposed approach is effective and practical. It might become a useful tool in protein subnuclear localization. The software in Matlab and supplementary materials are available freely by contacting the corresponding author.

  4. Silica Nanoparticles Target a Wnt Signal Transducer for Degradation and Impair Embryonic Development in Zebrafish.

    Science.gov (United States)

    Yi, Hongyang; Wang, Zhuyao; Li, Xiaojiao; Yin, Min; Wang, Lihua; Aldalbahi, Ali; El-Sayed, Nahed Nasser; Wang, Hui; Chen, Nan; Fan, Chunhai; Song, Haiyun

    2016-01-01

    Many types of biocompatible nanomaterials have proven of low cytotoxicity and hold great promise for various applications in nanomedicine. Whereas they generally do not cause apparent organ toxicity or tissue damage in adult animals, it is yet to determine their biological consequences in more general contexts. In this study, we investigate how silica nanoparticles (NPs) affect cellular activities and functions under several physiological or pathological conditions. Although silica NPs are generally regarded as "inert" nanocarriers and widely employed in biomedical studies, we find that they actively affect Wnt signaling in various types of cell lines, diminishing its anti-adipogenic effect in preadipocytes and pro-invasive effect in breast cancer cells, and more significantly, impair Wnt-regulated embryonic development in Zebrafish. We further demonstrate that intracellular silica NPs block Wnt signal transduction in a way resembling signaling molecules. Specifically, silica NPs target the Dvl protein, a key component of Wnt signaling cascade, for lysosomal degradation. As Wnt signaling play significant roles in embryonic development and adipogenesis, the observed physiological effects beyond toxicity imply potential risk of obesity, or developmental defects in somitogenesis and osteogenesis upon exposure to silica NPs. In addition, given the clinical implications of Wnt signaling in tumorigenesis and cancer metastasis, our work also establishes for the first time a molecular link between nanomaterials and the Wnt signaling pathway, which opens new door for novel applications of unmodified silica NPs in targeted therapy for cancers and other critical illness.

  5. Suppressor of cytokine Signaling-3 inhibits interleukin-1 signaling by targeting the TRAF-6/TAK1 complex

    DEFF Research Database (Denmark)

    Frobøse, Helle; Rønn, Sif Groth; Heding, Peter E

    2006-01-01

    -activated kinase 1, TNF receptor-associated factor (TRAF)6, and TGFbeta-activated kinase (TAK)1, but not when the MAP3K MAPK/ERK kinase kinase-1 is used instead of TAK1, indicating that the target for SOCS-3 is the TRAF6/TAK1 signaling complex. By coimmunoprecipitation, it was shown that SOCS-3 inhibited...... the association between TRAF6 and TAK1 and that SOCS-3 coimmunoprecipitated with TAK1 and TRAF6. Furthermore, SOCS-3 inhibited the IL-1-induced catalytic activity of TAK1. Because ubiquitination of TRAF6 is required for activation of TAK1, we analyzed the role of SOCS-3 on TRAF6 ubiquitination and found that SOCS......-3 inhibited ubiquitin modification of TRAF6. These results indicate that SOCS-3 inhibits IL-1 signal transduction by inhibiting ubiquitination of TRAF6, thus preventing association and activation of TAK1....

  6. Targeting GPCR-Gβγ-GRK2 signaling as a novel strategy for treating cardiorenal pathologies.

    Science.gov (United States)

    Rudomanova, Valeria; Blaxall, Burns C

    2017-08-01

    The pathologic crosstalk between the heart and kidney is known as cardiorenal syndrome (CRS). While the specific mechanisms underlying this crosstalk remain poorly understood, CRS is associated with exacerbated dysfunction of either or both organs and reduced survival. Maladaptive fibrotic remodeling is a key component of both heart and kidney failure pathogenesis and progression. G-protein coupled receptor (GPCR) signaling is a crucial regulator of cardiovascular and renal function. Chronic/pathologic GPCR signaling elicits the interaction of the G-protein Gβγ subunit with GPCR kinase 2 (GRK2), targeting the receptor for internalization, scaffolding to pathologic signals, and receptor degradation. Targeting this pathologic Gβγ-GRK2 interaction has been suggested as a possible strategy for the treatment of HF. In the current review, we discuss recent updates in understanding the role of GPCR-Gβγ-GRK2 signaling as a crucial mediator of maladaptive organ remodeling detected in HF and kidney dysfunction, with specific attention to small molecule-mediated inhibition of pathologic Gβγ-GRK2 interactions. Further, we explore the potential of GPCR-Gβγ-GRK2 signaling as a possible therapeutic target for cardiorenal pathologies. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. MicroRNA-145 suppresses hepatocellular carcinoma by targeting IRS1 and its downstream Akt signaling

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yelin [Department of Anesthesiology, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou (China); Key Laboratory of Combined Multi-organ Transplantation, Ministry of Public Health, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou (China); Hu, Chen; Cheng, Jun [Department of Surgery, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou (China); Key Laboratory of Combined Multi-organ Transplantation, Ministry of Public Health, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou (China); Chen, Binquan [Department of Anesthesiology, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou (China); Ke, Qinghong; Lv, Zhen; Wu, Jian [Department of Surgery, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou (China); Key Laboratory of Combined Multi-organ Transplantation, Ministry of Public Health, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou (China); Zhou, Yanfeng, E-mail: zyfhdj@yahoo.com [Department of Anesthesiology, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou (China)

    2014-04-18

    Highlights: • MiR-145 expression is down-regulated in HCC tissues and inversely related with IRS1 levels. • MiR-145 directly targets IRS1 in HCC cells. • Restored expression of miR-145 suppressed HCC cell proliferation and growth. • MiR-145 induced IRS1 under-expression potentially reduced downstream AKT signaling. - Abstract: Accumulating evidences have proved that dysregulation of microRNAs (miRNAs) is involved in cancer initiation and progression. In this study, we showed that miRNA-145 level was significantly decreased in hepatocellular cancer (HCC) tissues and cell lines, and its low expression was inversely associated with the abundance of insulin receptor substrate 1 (IRS1), a key mediator in oncogenic insulin-like growth factor (IGF) signaling. We verified IRS1 as a direct target of miR-145 using Western blotting and luciferase reporter assay. Further, the restoration of miR-145 in HCC cell lines suppressed cancer cell growth, owing to down-regulated IRS1 expression and its downstream Akt/FOXO1 signaling. Our results demonstrated that miR-145 could inhibit HCC through targeting IRS1 and its downstream signaling, implicating the loss of miR-145 regulation may be a potential molecular mechanism causing aberrant oncogenic signaling in HCC.

  8. Identification of novel plant peroxisomal targeting signals by a combination of machine learning methods and in vivo subcellular targeting analyses.

    Science.gov (United States)

    Lingner, Thomas; Kataya, Amr R; Antonicelli, Gerardo E; Benichou, Aline; Nilssen, Kjersti; Chen, Xiong-Yan; Siemsen, Tanja; Morgenstern, Burkhard; Meinicke, Peter; Reumann, Sigrun

    2011-04-01

    In the postgenomic era, accurate prediction tools are essential for identification of the proteomes of cell organelles. Prediction methods have been developed for peroxisome-targeted proteins in animals and fungi but are missing specifically for plants. For development of a predictor for plant proteins carrying peroxisome targeting signals type 1 (PTS1), we assembled more than 2500 homologous plant sequences, mainly from EST databases. We applied a discriminative machine learning approach to derive two different prediction methods, both of which showed high prediction accuracy and recognized specific targeting-enhancing patterns in the regions upstream of the PTS1 tripeptides. Upon application of these methods to the Arabidopsis thaliana genome, 392 gene models were predicted to be peroxisome targeted. These predictions were extensively tested in vivo, resulting in a high experimental verification rate of Arabidopsis proteins previously not known to be peroxisomal. The prediction methods were able to correctly infer novel PTS1 tripeptides, which even included novel residues. Twenty-three newly predicted PTS1 tripeptides were experimentally confirmed, and a high variability of the plant PTS1 motif was discovered. These prediction methods will be instrumental in identifying low-abundance and stress-inducible peroxisomal proteins and defining the entire peroxisomal proteome of Arabidopsis and agronomically important crop plants.

  9. Nuclear marginalization of host cell chromatin associated with expansion of two discrete virus-induced subnuclear compartments during baculovirus infection.

    Science.gov (United States)

    Nagamine, Toshihiro; Kawasaki, Yu; Abe, Atsushi; Matsumoto, Shogo

    2008-07-01

    Chromatin structure is strictly regulated during the cell cycle. DNA viruses occasionally disturb the spatial organization of the host cell chromatin due to formation of the viral DNA replication compartment. To examine chromatin behavior in baculovirus-infected cells, we constructed recombinant plasmids expressing fluorescent protein-tagged histone H4 molecules and visualized the intracellular localization of chromatin by their transient expression in live infected cells. Similar to other DNA viruses, the baculovirus Bombyx mori nucleopolyhedrovirus induced marginal relocation of chromatin within the nuclei of BmN cells, simultaneously with expansion of the viral DNA replication compartment, the virogenic stroma (VS). In the late stage of infection, however, the peristromal region (PR), another virus-induced subnuclear compartment, was also excluded from the chromatin-localizing area. Provided that late-gene products such as PR proteins (e.g., envelope proteins of the occlusion-derived virus) were expressed, blockage of viral DNA synthesis failed to inhibit chromatin relocation, despite abrogation of VS expansion. Instead, chromatin became marginalized concomitantly with PR expansion, suggesting that the PR contributes directly to chromatin replacement. In addition, chromatin was excluded from relatively large subnuclear structures that were induced in uninfected cells by cotransfection with four baculovirus genes, ie1, lef3, p143, and hr. Omission of any of the four genes, however, failed to result in formation of the large structures or chromatin exclusion. This correlation between compartmentalization and chromatin exclusion suggests the possibility that a chromatin-exclusive property of viral molecules, at least in part, supports nuclear compartmentalization of virus-infected cells.

  10. Nuclear Marginalization of Host Cell Chromatin Associated with Expansion of Two Discrete Virus-Induced Subnuclear Compartments during Baculovirus Infection▿

    Science.gov (United States)

    Nagamine, Toshihiro; Kawasaki, Yu; Abe, Atsushi; Matsumoto, Shogo

    2008-01-01

    Chromatin structure is strictly regulated during the cell cycle. DNA viruses occasionally disturb the spatial organization of the host cell chromatin due to formation of the viral DNA replication compartment. To examine chromatin behavior in baculovirus-infected cells, we constructed recombinant plasmids expressing fluorescent protein-tagged histone H4 molecules and visualized the intracellular localization of chromatin by their transient expression in live infected cells. Similar to other DNA viruses, the baculovirus Bombyx mori nucleopolyhedrovirus induced marginal relocation of chromatin within the nuclei of BmN cells, simultaneously with expansion of the viral DNA replication compartment, the virogenic stroma (VS). In the late stage of infection, however, the peristromal region (PR), another virus-induced subnuclear compartment, was also excluded from the chromatin-localizing area. Provided that late-gene products such as PR proteins (e.g., envelope proteins of the occlusion-derived virus) were expressed, blockage of viral DNA synthesis failed to inhibit chromatin relocation, despite abrogation of VS expansion. Instead, chromatin became marginalized concomitantly with PR expansion, suggesting that the PR contributes directly to chromatin replacement. In addition, chromatin was excluded from relatively large subnuclear structures that were induced in uninfected cells by cotransfection with four baculovirus genes, ie1, lef3, p143, and hr. Omission of any of the four genes, however, failed to result in formation of the large structures or chromatin exclusion. This correlation between compartmentalization and chromatin exclusion suggests the possibility that a chromatin-exclusive property of viral molecules, at least in part, supports nuclear compartmentalization of virus-infected cells. PMID:18434402

  11. Tissue-specific target analysis of disease-associated microRNAs in human signaling pathways.

    Directory of Open Access Journals (Sweden)

    Andreas Kowarsch

    Full Text Available MicroRNAs are a large class of post-transcriptional regulators that bind to the 3' untranslated region of messenger RNAs. They play a critical role in many cellular processes and have been linked to the control of signal transduction pathways. Recent studies indicate that microRNAs can function as tumor suppressors or even as oncogenes when aberrantly expressed. For more general insights of disease-associated microRNAs, we analyzed their impact on human signaling pathways from two perspectives. On a global scale, we found a core set of signaling pathways with enriched tissue-specific microRNA targets across diseases. The function of these pathways reflects the affinity of microRNAs to regulate cellular processes associated with apoptosis, proliferation or development. Comparing cancer and non-cancer related microRNAs, we found no significant differences between both groups. To unveil the interaction and regulation of microRNAs on signaling pathways locally, we analyzed the cellular location and process type of disease-associated microRNA targets and proteins. While disease-associated proteins are highly enriched in extracellular components of the pathway, microRNA targets are preferentially located in the nucleus. Moreover, targets of disease-associated microRNAs preferentially exhibit an inhibitory effect within the pathways in contrast to disease proteins. Our analysis provides systematic insights into the interaction of disease-associated microRNAs and signaling pathways and uncovers differences in cellular locations and process types of microRNA targets and disease-associated proteins.

  12. Signal integration: a framework for understanding the efficacy of therapeutics targeting the human EGFR family

    Science.gov (United States)

    Shepard, H. Michael; Brdlik, Cathleen M.; Schreiber, Hans

    2008-01-01

    The human EGFR (HER) family is essential for communication between many epithelial cancer cell types and the tumor microenvironment. Therapeutics targeting the HER family have demonstrated clinical success in the treatment of diverse epithelial cancers. Here we propose that the success of HER family–targeted monoclonal antibodies in cancer results from their ability to interfere with HER family consolidation of signals initiated by a multitude of other receptor systems. Ligand/receptor systems that initiate these signals include cytokine receptors, chemokine receptors, TLRs, GPCRs, and integrins. We further extrapolate that improvements in cancer therapeutics targeting the HER family are likely to incorporate mechanisms that block or reverse stromal support of malignant progression by isolating the HER family from autocrine and stromal influences. PMID:18982164

  13. Impact of targeting insulin-like growth factor signaling in head and neck cancers.

    Science.gov (United States)

    Limesand, Kirsten H; Chibly, Alejandro Martinez; Fribley, Andrew

    2013-10-01

    The IGF system has been shown to have either negative or negligible impact on clinical outcomes of tumor development depending on specific tumor sites or stages. This review focuses on the clinical impact of IGF signaling in head and neck cancer, the effects of IGF targeted therapies, and the multi-dimensional role of IRS 1/2 signaling as a potential mechanism in resistance to targeted therapies. Similar to other tumor sites, both negative and positive correlations between levels of IGF-1/IGF-1-R and clinical outcomes in head and neck cancer have been reported. In addition, utilization of IGF targeted therapies has not demonstrated significant clinical benefit; therefore the prognostic impact of the IGF system on head and neck cancer remains uncertain. Copyright © 2013 Elsevier Ltd. All rights reserved.

  14. Microbial effectors target multiple steps in the salicylic acid production and signaling pathway

    Directory of Open Access Journals (Sweden)

    Shigeyuki eTanaka

    2015-05-01

    Full Text Available Microbes attempting to colonize plants are recognized through the plant immune surveillance system. This leads to a complex array of global as well as specific defense responses, which are often associated with plant cell death and subsequent arrest of the invader. The responses also entail complex changes in phytohormone signaling pathways. Among these, salicylic acid signaling is an important pathway because of its ability to trigger plant cell death. As biotrophic and hemibiotrophic pathogens need to invade living plant tissue to cause disease, they have evolved efficient strategies to downregulate salicylic acid signaling by virulence effectors, which can be proteins or secondary metabolites. Here we review the strategies prokaryotic pathogens have developed to target salicylic acid biosynthesis and signaling, and contrast this with recent insights into how plant pathogenic eukaryotic fungi and oomycetes accomplish the same goal.

  15. Multiresolution Signal Processing Techniques for Ground Moving Target Detection Using Airborne Radar

    Directory of Open Access Journals (Sweden)

    Bergin Jameson S

    2006-01-01

    Full Text Available Synthetic aperture radar (SAR exploits very high spatial resolution via temporal integration and ownship motion to reduce the background clutter power in a given resolution cell to allow detection of nonmoving targets. Ground moving target indicator (GMTI radar, on the other hand, employs much lower-resolution processing but exploits relative differences in the space-time response between moving targets and clutter for detection. Therefore, SAR and GMTI represent two different temporal processing resolution scales which have typically been optimized and demonstrated independently to work well for detecting either stationary (in the case of SAR or exo-clutter (in the case of GMTI targets. Based on this multiresolution interpretation of airborne radar data processing, there appears to be an opportunity to develop detection techniques that attempt to optimize the signal processing resolution scale (e.g., length of temporal integration to match the dynamics of a target of interest. This paper investigates signal processing techniques that exploit long CPIs to improve the detection performance of very slow-moving targets.

  16. To fingolimod and beyond: The rich pipeline of drug candidates that target S1P signaling.

    Science.gov (United States)

    Chew, Wee Siong; Wang, Wei; Herr, Deron R

    2016-11-01

    Sphingosine 1-phosphate (S1P) is an extracellular lipid signaling molecule that acts as a selective, high-affinity ligand for a family of five G protein-coupled receptors. This signaling system was first identified twenty years ago, and has since been shown to regulate a diverse range of physiological processes and disease states, such as cardiovascular development, immune function, hypoxic responses, and cancer. The therapeutic potential of targeting this system took center stage when it was demonstrated that the immune modulator, fingolimod (FTY720/Gilenya), exerts it lymphopenic effect by acting on S1P receptors, primarily on S1P receptor 1 (S1P1). In 2010, fingolimod became the first oral medication approved for the treatment of multiple sclerosis (MS). Since then, second-generation S1P receptor modulators have been under development in an effort to provide improved safety and efficacy profiles for MS, and to broaden their use to other autoimmune indications. Beyond the development of S1P1-modulators, there has been considerable effort in targeting other components of the S1P signaling pathway for the treatment of other diseases, such as cardiovascular disease, sepsis, and cancer. This manuscript provides an overview of the clinical and preclinical development of drugs targeting S1P signaling. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Targeting Calcium Signaling Induces Epigenetic Reactivation of Tumor Suppressor Genes in Cancer.

    Science.gov (United States)

    Raynal, Noël J-M; Lee, Justin T; Wang, Youjun; Beaudry, Annie; Madireddi, Priyanka; Garriga, Judith; Malouf, Gabriel G; Dumont, Sarah; Dettman, Elisha J; Gharibyan, Vazganush; Ahmed, Saira; Chung, Woonbok; Childers, Wayne E; Abou-Gharbia, Magid; Henry, Ryan A; Andrews, Andrew J; Jelinek, Jaroslav; Cui, Ying; Baylin, Stephen B; Gill, Donald L; Issa, Jean-Pierre J

    2016-03-15

    Targeting epigenetic pathways is a promising approach for cancer therapy. Here, we report on the unexpected finding that targeting calcium signaling can reverse epigenetic silencing of tumor suppressor genes (TSG). In a screen for drugs that reactivate silenced gene expression in colon cancer cells, we found three classical epigenetic targeted drugs (DNA methylation and histone deacetylase inhibitors) and 11 other drugs that induced methylated and silenced CpG island promoters driving a reporter gene (GFP) as well as endogenous TSGs in multiple cancer cell lines. These newly identified drugs, most prominently cardiac glycosides, did not change DNA methylation locally or histone modifications globally. Instead, all 11 drugs altered calcium signaling and triggered calcium-calmodulin kinase (CamK) activity, leading to MeCP2 nuclear exclusion. Blocking CamK activity abolished gene reactivation and cancer cell killing by these drugs, showing that triggering calcium fluxes is an essential component of their epigenetic mechanism of action. Our data identify calcium signaling as a new pathway that can be targeted to reactivate TSGs in cancer. ©2015 American Association for Cancer Research.

  18. Block design enhances classification of 3D reach targets from electroencephalographic signals.

    Science.gov (United States)

    Sosnik, Ronen; Tadipatri, Vijay Aditya; Tewfik, Ahmed H; Pellizzer, Giuseppe

    2016-08-04

    To date, decoding accuracy of actual or imagined pointing movements to targets in 3D space from electroencephalographic (EEG) signals has remained modest. The reason may pertain to the fact that these movements activate essentially the same neural networks. In this study, we aimed at testing whether repetitive pointing movements to each of the targets promotes the development of segregated neural patterns, resulting in enhanced decoding accuracy. Six human subjects generated slow or fast repetitive pointing movements with their right dominant arm to one of five targets distributed in 3D space, followed by repetitive imagery of movements to the same target or to a different target. Nine naive subjects generated both repetitive and non-repetitive slow actual movements to each of the five targets to test the effect of block design on decoding accuracy. In order to assure that base line drift and low frequency motion artifacts do not contaminate the data, the data were high-pass filtered in 4-30Hz, leaving out the delta and gamma band. For the repetitive trials, the model decoded target location with 81% accuracy, which is significantly higher than chance level. The average decoding rate of target location was only 30% for the non-repetitive trials, which is not significantly different than chance level. A subset of electrodes, mainly over the contralateral sensorimotor areas, was found to provide most of the discriminative features for all tested conditions. Time proximity between trained and tested blocks was found to enhance decoding accuracy of target location both by target non-specific and specific mechanisms. Our findings suggest that movement repetition promotes the development of distinct neural patterns, presumably by the formation of target-specific kinesthetic memory. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

  19. p27: a barometer of signaling deregulation and potential predictor of response to targeted therapies.

    Science.gov (United States)

    Wander, Seth A; Zhao, Dekuang; Slingerland, Joyce M

    2011-01-01

    Phosphorylation of the cyclin-dependent kinase inhibitor p27 by upstream mitogenic signaling pathways regulates its stability, localization, and biological function. In human cancers, loss of the antiproliferative action of p27 can arise through reduced protein levels and/or cytoplasmic mislocalization, leading to increased cell proliferation and/or cell migration, respectively. Reduced p27 expression levels and p27 mislocalization have potential prognostic and therapeutic implications in various types of human cancers. This review highlights mechanisms of functional deregulation of p27 by oncogenic signaling that provide an important molecular rationale for pathway targeting in cancer treatment. ©2010 AACR.

  20. Understanding and Targeting the Wnt/β-Catenin Signaling Pathway in Chronic Leukemia

    Directory of Open Access Journals (Sweden)

    S. Thanendrarajan

    2011-01-01

    Full Text Available It has been revealed that the Wnt/β-catenin signaling pathway plays an important role in the development of solid tumors and hematological malignancies, particularly in B-cell neoplasia and leukemia. In the last decade there have been made experimental approaches targeting the Wnt pathway in chronic leukemia. In this paper we provide an overview about the current state of knowledge regarding the Wnt/β-catenin signaling pathway in chronic leukemia with special focus on therapeutic options and strategies.

  1. The Probing Radio Signal Polarization Effect on Separation Efficiency of Surface Target Response

    Directory of Open Access Journals (Sweden)

    A. N. Pinchuk

    2015-01-01

    Full Text Available The aim of the study was a quantitative analysis of the level of interference with radar monitoring characteristics of surface targets, caused by the scattered electromagnetic field, arising due to the interaction between radio waves and sea surface, which is a study aspect a radiooceanography encompasses. Backscatter signal, arising from the interaction of radio waves and sea surface, extends in a direction opposite the probing radar signal of spread marine and coastal radar stations.With radar sounding of sea surface at high incidence angles of radio waves, a basic physical mechanism to form the received signal is resonant (Bragg scattering, and at small incidence angles of radio waves it is quasi-specular reflection. Consequently, the energy of electromagnetic radiation, backscattered by the sea surface, depends on the type of wave polarization: for horizontal polarization it is less than for vertical one.The paper presents a mathematical model, which describes dependence of interference level caused by interaction between radio waves and sea surface, on the radio wave polarization for the case when the same polarization is used to sent-out and receive a radio wave.To determine the noise reduction to be achievable with radar monitoring the surface targets by selecting the polarization of the probing radar signal, a signal/noise ratio is analyzed for its different polarizations.It is shown that in order to reduce the noise level caused by the interaction between radio waves and sea surface, it is possible to use the differences in the level of scattered radio signals of different polarization: with horizontally-polarized radar operation at incidence angles of 75°- 85° a signal/noise ratio is by 20-35 dB higher than that of vertically- polarized one.

  2. Nicking endonuclease and target recycles signal amplification assisted quantum dots for fluorescence detection of DNA

    Energy Technology Data Exchange (ETDEWEB)

    Niu Shuyan; Li Quanyi; Qu Lijing; Wang Wei [Key Lab of Eco-chemical Engineering, Ministry of Education, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042 (China)

    2010-11-08

    An ultrasensitive fluorescence detection method for DNA based on nicking endonuclease (NEase) and target recycles assisted with CdTe quantum dots (QDs) is reported. In the detection system, when the target DNA is present, it hybridizes with a linker strand to from a duplex, in which the NEase recognizes specific nucleotide sequences and cleaves the linker strand. After nicking, the fragments of the linker strand spontaneously dissociate from the target DNA and another linker strand hybridizes to the target to trigger another strand-scission cycle. On the other hand, when the target was absent, no duplex is formed and no fragment of linker strand is produced. Then CdTe QDs and magnetic beads (MBs), which were all modified with DNA sequences complementary to that of the linker strands are added to the solution to detect the presence of a target DNA. The signal was generated through the difference in Foerster resonance energy transfer (FRET) between the MB and CdTe QDs. This method indicates that one target DNA leads to cleavage of hundreds of linker DNA, increasing detection sensitivity by nearly three orders of magnitude. This method should be applicable whenever there is a requirement to detect a specific DNA sequence and can also be used for multicomponent detection.

  3. cGMP signaling as a target for the prevention and treatment of breast cancer.

    Science.gov (United States)

    Windham, Perrin F; Tinsley, Heather N

    2015-04-01

    One in eight women in the United States will be diagnosed with invasive breast cancer in her lifetime. Advances in therapeutic strategies, diagnosis, and improved awareness have resulted in a significant reduction in breast cancer related mortality. However, there is a continued need for more effective and less toxic drugs for both the prevention and the treatment of breast cancer in order to see a continued decline in the morbidity and mortality associated with this disease. Recent studies suggest that the cGMP signaling pathway may be aberrantly regulated in breast cancer. As such, this pathway may serve as a source of novel targets for future breast cancer drug discovery efforts. This review provides an overview of cGMP signaling in normal physiology and in breast cancer as well as current strategies being investigated for targeting this pathway in breast cancer. Copyright © 2014 Elsevier Ltd. All rights reserved.

  4. Lithium inhibits tumorigenic potential of PDA cells through targeting hedgehog-GLI signaling pathway.

    Directory of Open Access Journals (Sweden)

    Zhonglu Peng

    Full Text Available Hedgehog signaling pathway plays a critical role in the initiation and development of pancreatic ductal adenocarcinoma (PDA and represents an attractive target for PDA treatment. Lithium, a clinical mood stabilizer for mental disorders, potently inhibits the activity of glycogen synthase kinase 3β (GSK3β that promotes the ubiquitin-dependent proteasome degradation of GLI1, an important downstream component of hedgehog signaling. Herein, we report that lithium inhibits cell proliferation, blocks G1/S cell-cycle progression, induces cell apoptosis and suppresses tumorigenic potential of PDA cells through down-regulation of the expression and activity of GLI1. Moreover, lithium synergistically enhances the anti-cancer effect of gemcitabine. These findings further our knowledge of mechanisms of action for lithium and provide a potentially new therapeutic strategy for PDA through targeting GLI1.

  5. Potential for targeting dopamine/DARPP-32 signaling in neuropsychiatric and neurodegenerative disorders.

    Science.gov (United States)

    Nishi, Akinori; Shuto, Takahide

    2017-03-01

    Alterations in dopamine neurotransmission has been implicated in pathophysiology of neuropsychiatric and neurodegenerative disorders, and DARPP-32 plays a pivotal role in dopamine neurotransmission. DARPP-32 likely influences dopamine-mediated behaviors in animal models of neuropsychiatric and neurodegenerative disorders and therapeutic effects of pharmacological treatment. Areas covered: We will review animal studies on the biochemical and behavioral roles of DARPP-32 in drug addiction, schizophrenia and Parkinson's disease. In general, under physiological and pathophysiological conditions, DARPP-32 in D1 receptor expressing (D1R) -medium spiny neurons (MSNs) promotes dopamine/D1 receptor/PKA signaling, whereas DARPP-32 in D2 receptor expressing (D2R)-MSNs counteracts dopamine/D2 receptor signaling. However, the function of DARPP-32 is differentially regulated in acute and chronic phases of drug addiction; DARPP-32 enhances D1 receptor/PKA signaling in the acute phase, whereas DARPP-32 suppresses D1 receptor/PKA signaling in the chronic phase through homeostatic mechanisms. Therefore, DARPP-32 plays a bidirectional role in dopamine neurotransmission, depending on the cell type and experimental conditions, and is involved in dopamine-related behavioral abnormalities. Expert opinion: DARPP-32 differentially regulates dopamine signaling in D1R- and D2R-MSNs, and a shift of balance between D1R- and D2R-MSN function is associated with behavioral abnormalities. An adjustment of this imbalance is achieved by therapeutic approaches targeting DARPP-32-related signaling molecules.

  6. Nitric oxide signaling in human ovarian cancer: A potential therapeutic target.

    Science.gov (United States)

    El-Sehemy, Ahmed; Postovit, Lynne-Marie; Fu, YangXin

    2016-04-01

    Ovarian cancer is the leading cause of death due to gynecologic malignancies worldwide. Current therapy regimens are ineffective to treat advanced ovarian cancers, presenting a need to develop novel therapeutic strategies. Nitric oxide (NO) is a multifunctional gaseous molecule that is generated by cancer, stromal and endothelial cells and plays a multifaceted role in cancer biology through multiple mechanisms. Accumulating evidence suggests that NO signaling is involved in multiple aspects of ovarian cancer, including growth, apoptosis, cancer-stromal cell interaction, angiogenesis and response to chemotherapy. This review will discuss the experimental and clinical evidence of the involvement of NO signaling in ovarian cancer and the therapeutic potential of targeting NO signaling in ovarian cancer. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. Non-genomic actions of aldosterone: From receptors and signals to membrane targets.

    LENUS (Irish Health Repository)

    2012-02-01

    In tissues which express the mineralocorticoid receptor (MR), aldosterone modulates the expression of membrane targets such as the subunits of the epithelial Na(+) channel, in combination with important signalling intermediates such as serum and glucocorticoid-regulated kinase-1. In addition, the rapid \\'non-genomic\\' activation of protein kinases and secondary messenger signalling cascades has also been detected in aldosterone-sensitive tissues of the nephron, distal colon and cardiovascular system. These rapid actions are variously described as being coupled to MR or to an as yet unidentified, membrane-associated aldosterone receptor. The rapidly activated signalling cascades add a level of fine-tuning to the activity of aldosterone-responsive membrane transporters and also modulate the aldosterone-induced changes in gene expression through receptor and transcription factor phosphorylation.

  8. Non-genomic actions of aldosterone: From receptors and signals to membrane targets.

    LENUS (Irish Health Repository)

    Dooley, Ruth

    2011-07-26

    In tissues which express the mineralocorticoid receptor (MR), aldosterone modulates the expression of membrane targets such as the subunits of the epithelial Na(+) channel, in combination with important signalling intermediates such as serum and glucocorticoid-regulated kinase-1. In addition, the rapid \\'non-genomic\\' activation of protein kinases and secondary messenger signalling cascades has also been detected in aldosterone-sensitive tissues of the nephron, distal colon and cardiovascular system. These rapid actions are variously described as being coupled to MR or to an as yet unidentified, membrane-associated aldosterone receptor. The rapidly activated signalling cascades add a level of fine-tuning to the activity of aldosterone-responsive membrane transporters and also modulate the aldosterone-induced changes in gene expression through receptor and transcription factor phosphorylation.

  9. Methylglyoxal activates the target of rapamycin complex 2-protein kinase C signaling pathway in Saccharomyces cerevisiae.

    Science.gov (United States)

    Nomura, Wataru; Inoue, Yoshiharu

    2015-04-01

    Methylglyoxal is a typical 2-oxoaldehyde derived from glycolysis. We show here that methylglyoxal activates the Pkc1-Mpk1 mitogen-activated protein (MAP) kinase cascade in a target of rapamycin complex 2 (TORC2)-dependent manner in the budding yeast Saccharomyces cerevisiae. We demonstrate that TORC2 phosphorylates Pkc1 at Thr(1125) and Ser(1143). Methylglyoxal enhanced the phosphorylation of Pkc1 at Ser(1143), which transmitted the signal to the downstream Mpk1 MAP kinase cascade. We found that the phosphorylation status of Pkc1(T1125) affected the phosphorylation of Pkc1 at Ser(1143), in addition to its protein levels. Methylglyoxal activated mammalian TORC2 signaling, which, in turn, phosphorylated Akt at Ser(473). Our results suggest that methylglyoxal is a conserved initiator of TORC2 signaling among eukaryotes. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  10. Monte Carlo Simulation of the Echo Signals from Low-Flying Targets for Airborne Radar

    Directory of Open Access Journals (Sweden)

    Mingyuan Man

    2014-01-01

    Full Text Available A demonstrated hybrid method based on the combination of half-space physical optics method (PO, graphical-electromagnetic computing (GRECO, and Monte Carlo method on echo signals from low-flying targets based on actual environment for airborne radar is presented in this paper. The half-space physical optics method , combined with the graphical-electromagnetic computing (GRECO method to eliminate the shadow regions quickly and rebuild the target automatically, is employed to calculate the radar cross section (RCS of the conductive targets in half space fast and accurately. The direct echo is computed based on the radar equation. The reflected paths from sea or ground surface cause multipath effects. In order to accurately obtain the echo signals, the phase factors are modified for fluctuations in multipath, and the statistical average value of the echo signals is obtained using the Monte Carlo method. A typical simulation is performed, and the numerical results show the accuracy of the proposed method.

  11. Molecular alterations in signal pathways of melanoma and new personalized treatment strategies: Targeting of Notch

    Directory of Open Access Journals (Sweden)

    Julija Mozūraitienė

    2015-01-01

    Full Text Available Despite modern achievements in therapy of malignant melanomas new treatment strategies are welcomed in clinics for survival of patients. Now it is supposed that personalized molecular therapies for each patient are needed concerning a specificity of molecular alterations in patient's tumors. In human melanoma, Notch signaling interacts with other pathways, including MAPK, PI3K-AKT, NF-kB, and p53. This article discusses mutated genes and leading aberrant signal pathways in human melanoma which are of interest concerning to their perspective for personalized treatment strategies in melanoma. We speculate that E3 ubiquitin ligases MDM2 and MDM4 can be attractive therapeutic target for p53 and Notch signaling pathways in malignant melanoma by using small molecule inhibitors. It is possible that restoration of p53-MDM2-NUMB complexes in melanoma can restore wild type p53 function and positively modulate Notch pathway. In this review we summarize recent data about novel US Food and Drug Administration approved target drugs for metastatic melanoma treatment, and suppose model for treatment strategy by targeting Notch.

  12. L-type calcium channel targeting and local signalling in cardiac myocytes.

    Science.gov (United States)

    Shaw, Robin M; Colecraft, Henry M

    2013-05-01

    In the heart, Ca(2+) influx via Ca(V)1.2 L-type calcium channels (LTCCs) is a multi-functional signal that triggers muscle contraction, controls action potential duration, and regulates gene expression. The use of LTCC Ca(2+) as a multi-dimensional signalling molecule in the heart is complicated by several aspects of cardiac physiology. Cytosolic Ca(2+) continuously cycles between ~100 nM and ~1 μM with each heartbeat due to Ca(2+) linked signalling from LTCCs to ryanodine receptors. This rapid cycling raises the question as to how cardiac myocytes distinguish the Ca(2+) fluxes originating through L-type channels that are dedicated to contraction from Ca(2+) fluxes originating from other L-type channels that are used for non-contraction-related signalling. In general, disparate Ca(2+) sources in cardiac myocytes such as current through differently localized LTCCs as well as from IP3 receptors can signal selectively to Ca(2+)-dependent effectors in local microdomains that can be impervious to the cytoplasmic Ca(2+) transients that drive contraction. A particular challenge for diversified signalling via cardiac LTCCs is that they are voltage-gated and, therefore, open and presumably flood their microdomains with Ca(2+) with each action potential. Thus spatial localization of Cav1.2 channels to different types of microdomains of the ventricular cardiomyocyte membrane as well as the existence of particular macromolecular complexes in each Cav1.2 microdomain are important to effect different types of Cav1.2 signalling. In this review we examine aspects of Cav1.2 structure, targeting and signalling in two specialized membrane microdomains--transverse tubules and caveolae.

  13. Cellular Signaling Pathway Alterations and Potential Targeted Therapies for Medullary Thyroid Carcinoma

    Directory of Open Access Journals (Sweden)

    Serena Giunti

    2013-01-01

    Full Text Available Parafollicular C-cell-derived medullary thyroid cancer (MTC comprises 3% to 4% of all thyroid cancers. While cytotoxic treatments have been shown to have limited efficacy, targeted molecular therapies that inhibit rearranged during transfection (RET and other tyrosine kinase receptors that are mainly involved in angiogenesis have shown great promise in the treatment of metastatic or locally advanced MTC. Multi-tyrosine kinase inhibitors such as vandetanib, which is already approved for the treatment of progressive MTC, and cabozantinib have shown distinct advantages with regard to rates of disease response and control. However, these types of tyrosine kinase inhibitor compounds are able to concurrently block several types of targets, which limits the understanding of RET as a specific target. Moreover, important resistances to tyrosine kinase inhibitors can occur, which limit the long-term efficacy of these treatments. Deregulated cellular signaling pathways and genetic alterations in MTC, particularly the activation of the RAS/mammalian target of rapamycin (mTOR cascades and RET crosstalk signaling, are now emerging as novel and potentially promising therapeutic treatments for aggressive MTC.

  14. The complexity of targeting EGFR signalling in cancer: from expression to turnover.

    Science.gov (United States)

    Sebastian, Sinto; Settleman, Jeffrey; Reshkin, Stephan J; Azzariti, Amalia; Bellizzi, Antonia; Paradiso, Angelo

    2006-08-01

    The epidermal growth factor receptor (ErbB1 or EGFR) has been found to be altered in a variety of human cancers. A number of agents targeting these receptors, including specific antibodies directed against the ligand-binding domain of the receptor and small molecules that inhibit kinase activity are either in clinical trials or are already approved for clinical treatment. However, identifying patients that are likely to respond to such treatments has been challenging. As a consequence, it still remains important to identify additional alterations of the tumor cell that contribute to the response to EGFR-targeted agents. While EGFR-mediated signalling pathways have been well established, there is still a rather limited understanding of how intracellular protein-protein interactions, ubiquitination, endocytosis and subsequent degradation of EGFR contribute to the determination of sensitivity to EGFR targeting agents and are emerging areas of investigation. This review primarily focuses on the basic signal transduction pathways mediated through activated membrane bound and/or endosomal EGFR and emphasizes the need to co-target additional proteins that function either upstream or downstream of EGFR to improve cancer therapy.

  15. An autocrine Wnt5a-Ror signaling loop mediates sympathetic target innervation.

    Science.gov (United States)

    Ryu, Yun Kyoung; Collins, Sarah Ellen; Ho, Hsin-Yi Henry; Zhao, Haiqing; Kuruvilla, Rejji

    2013-05-01

    During nervous system development, axon branching at nerve terminals is an essential step in the formation of functional connections between neurons and target cells. It is known that target tissues exert control of terminal arborization through secretion of trophic factors. However, whether the in-growing axons themselves produce diffusible cues to instruct target innervation remains unclear. Here, we use conditional mutant mice to show that Wnt5a derived from sympathetic neurons is required for their target innervation in vivo. Conditional deletion of Wnt5a resulted in specific deficits in the extension and arborization of sympathetic fibers in their final target fields, while no defects were observed in the overall tissue patterning, proliferation, migration or differentiation of neuronal progenitors. Using compartmentalized neuronal cultures, we further demonstrate that the Ror receptor tyrosine kinases are required locally in sympathetic axons to mediate Wnt5a-dependent branching. Thus, our study suggests an autocrine Wnt5a-Ror signaling pathway that directs sympathetic axon branching during target innervation. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. Redox Signaling as a Therapeutic Target to Inhibit Myofibroblast Activation in Degenerative Fibrotic Disease

    Directory of Open Access Journals (Sweden)

    Natalie Sampson

    2014-01-01

    Full Text Available Degenerative fibrotic diseases encompass numerous systemic and organ-specific disorders. Despite their associated significant morbidity and mortality, there is currently no effective antifibrotic treatment. Fibrosis is characterized by the development and persistence of myofibroblasts, whose unregulated deposition of extracellular matrix components disrupts signaling cascades and normal tissue architecture leading to organ failure and death. The profibrotic cytokine transforming growth factor beta (TGFβ is considered the foremost inducer of fibrosis, driving myofibroblast differentiation in diverse tissues. This review summarizes recent in vitro and in vivo data demonstrating that TGFβ-induced myofibroblast differentiation is driven by a prooxidant shift in redox homeostasis. Elevated NADPH oxidase 4 (NOX4-derived hydrogen peroxide (H2O2 supported by concomitant decreases in nitric oxide (NO signaling and reactive oxygen species scavengers are central factors in the molecular pathogenesis of fibrosis in numerous tissues and organs. Moreover, complex interplay between NOX4-derived H2O2 and NO signaling regulates myofibroblast differentiation. Restoring redox homeostasis via antioxidants or NOX4 inactivation as well as by enhancing NO signaling via activation of soluble guanylyl cyclases or inhibition of phosphodiesterases can inhibit and reverse myofibroblast differentiation. Thus, dysregulated redox signaling represents a potential therapeutic target for the treatment of wide variety of different degenerative fibrotic disorders.

  17. Elevated immune-inflammatory signaling in mood disorders: a new therapeutic target?

    Science.gov (United States)

    McNamara, Robert K; Lotrich, Francis E

    2012-09-01

    Converging translational evidence has implicated elevated immune-inflammatory signaling activity in the pathoetiology of mood disorders, including major depressive disorder and bipolar disorder. This is supported in part by cross-sectional evidence for increased levels of proinflammatory eicosanoids, cytokines and acute-phase proteins during mood episodes, and prospective longitudinal evidence for the emergence of mood symptoms in response to chronic immune-inflammatory activation. In addition, mood-stabilizer and atypical antipsychotic medications downregulate initial components of the immune-inflammatory signaling pathway, and adjunctive treatment with anti-inflammatory agents augment the therapeutic efficacy of antidepressant, mood stabilizer and atypical antipsychotic medications. Potential pathogenic mechanisms linked with elevated immune-inflammatory signaling include perturbations in central serotonin neurotransmission and progressive white matter pathology. Both heritable genetic factors and environmental factors including dietary fatty-acid composition may act in concert to sustain elevated immune-inflammatory signaling. Collectively, these data suggest that elevated immune-inflammatory signaling is a mechanism that is relevant to the pathoetiology of mood disorders, and may therefore represent a new therapeutic target for the development of more effective treatments.

  18. Androgen receptor signaling pathways as a target for breast cancer treatment.

    Science.gov (United States)

    Pietri, Elisabetta; Conteduca, Vincenza; Andreis, Daniele; Massa, Ilaria; Melegari, Elisabetta; Sarti, Samanta; Cecconetto, Lorenzo; Schirone, Alessio; Bravaccini, Sara; Serra, Patrizia; Fedeli, Anna; Maltoni, Roberta; Amadori, Dino; De Giorgi, Ugo; Rocca, Andrea

    2016-10-01

    The androgen receptor (AR) is a ligand-dependent transcription factor, and its effects on breast range from physiological pubertal development and age-related modifications to cancer onset and proliferation. The prevalence of AR in early breast cancer is around 60%, and AR is more frequently expressed in ER-positive than in ER-negative tumors. We offer an overview of AR signaling pathways in different breast cancer subtypes, providing evidence that its oncogenic role is likely to be different in distinct biological and clinical scenarios. In particular, in ER-positive breast cancer, AR signaling often antagonizes the growth stimulatory effect of ER signaling; in triple-negative breast cancer (TNBC), AR seems to drive tumor progression (at least in luminal AR subtype of TNBC with a gene expression profile mimicking luminal subtypes despite being negative to ER and enriched in AR expression); in HER2-positive breast cancer, in the absence of ER expression, AR signaling has a proliferative role. These data represent the rationale for AR-targeting treatment as a potentially new target therapy in breast cancer subset using androgen agonists in some AR-positive/ER-positive tumors, AR antagonists in triple-negative/AR-positive tumors and in combination with anti-HER2 agents or with other signaling pathways inhibitors (including PI3K/MYC/ERK) in HER2-positive/AR-positive tumors. Only the ongoing and future prospective clinical trials will allow us to establish which agents are the best option in every specific condition, keeping in mind that there is evidence of opposite androgens and AR agonist/antagonist drug effects on cell proliferation particularly in AR-positive/ER-positive tumors. © 2016 Society for Endocrinology.

  19. Campylobacter jejuni induces colitis through activation of mammalian target of rapamycin signaling.

    Science.gov (United States)

    Sun, Xiaolun; Threadgill, Deborah; Jobin, Christian

    2012-01-01

    Campylobacter jejuni is the worldwide leading cause of bacterial-induced enteritis. The molecular and cellular events that lead to campylobacteriosis are poorly understood. We identify mammalian target of rapamycin (mTOR) as a signaling pathway that leads to C jejuni-induced intestinal inflammation. Germ-free (control) or conventionally derived Il10(-/-) mice that express enhanced green fluorescent protein (EGFP) under the control of nuclear factor κB (Il10(-/-); NF-κB(EGFP) mice) were infected with C jejuni (10(9) colony-forming units/mouse) for 12 days; their responses were determined using histologic, semiquantitative reverse-transcription polymerase chain reaction, fluorescence in situ hybridization, transmission electron microscopy, and tissue culture analyses. mTOR signaling was blocked by daily intraperitoneal injections of the pharmacologic inhibitor rapamycin (1.5 mg/kg). CD4(+) T cells were depleted by intraperitoneal injections of antibodies against CD4 (0.5 mg/mouse every 3 days). Bacterial survival in splenocytes was measured using a gentamycin killing assay. C jejuni induced intestinal inflammation, which correlated with activation of mTOR signaling and neutrophil infiltration. The inflamed intestines of these mice had increased levels of interleukin-1β, Cxcl2, interleukin-17a, and EGFP; C jejuni localized to colons and extraintestinal tissues of infected Il10(-/-); NF-κB(EGFP) mice compared with controls. Rapamycin, administered before or after introduction of C jejuni, blocked C jejuni-induced intestinal inflammation and bacterial accumulation. LC3II processing and killing of C jejuni were increased in splenocytes incubated with rapamycin compared with controls. mTOR signaling mediates C jejuni-induced colitis in Il10(-/-) mice, independently of T-cell activation. Factors involved in mTOR signaling might be therapeutic targets for campylobacteriosis. Copyright © 2012 AGA Institute. Published by Elsevier Inc. All rights reserved.

  20. Novel Signal-Enhancing Approaches for Optical Detection of Nucleic Acids—Going beyond Target Amplification

    Directory of Open Access Journals (Sweden)

    Laura Miotke

    2015-09-01

    Full Text Available Detection of low-abundance nucleic acids is a challenging task, which over the last two decades has been solved using enzymatic target amplification. Enzymatic synthesis enhances the signal so that diverse, scientifically and clinically relevant molecules can be identified and studied, including cancer DNA, viral nucleic acids, and regulatory RNAs. However, using enzymes increases the detection time and cost, not to mention the high risk of mistakes with amplification and data alignment. These limitations have stimulated a growing interest in enzyme-free methods within researchers and industry. In this review we discuss recent advances in signal-enhancing approaches aimed at nucleic acid diagnostics that do not require target amplification. Regardless of enzyme usage, signal enhancement is crucial for the reliable detection of nucleic acids at low concentrations. We pay special attention to novel nanomaterials, fluorescence microscopy, and technical advances in detectors for optical assessment. We summarize sensitivity parameters of the currently available assays and devices which makes this review relevant to the broad spectrum of researchers working in fields from biophysics, to engineering, to synthetic biology and bioorganic chemistry.

  1. Targeting autocrine HB-EGF signaling with specific ADAM12 inhibition using recombinant ADAM12 prodomain

    Science.gov (United States)

    Miller, Miles A.; Moss, Marcia L.; Powell, Gary; Petrovich, Robert; Edwards, Lori; Meyer, Aaron S.; Griffith, Linda G.; Lauffenburger, Douglas A.

    2015-01-01

    Dysregulation of ErbB-family signaling underlies numerous pathologies and has been therapeutically targeted through inhibiting ErbB-receptors themselves or their cognate ligands. For the latter, “decoy” antibodies have been developed to sequester ligands including heparin-binding epidermal growth factor (HB-EGF); however, demonstrating sufficient efficacy has been difficult. Here, we hypothesized that this strategy depends on properties such as ligand-receptor binding affinity, which varies widely across the known ErbB-family ligands. Guided by computational modeling, we found that high-affinity ligands such as HB-EGF are more difficult to target with decoy antibodies compared to low-affinity ligands such as amphiregulin (AREG). To address this issue, we developed an alternative method for inhibiting HB-EGF activity by targeting its cleavage from the cell surface. In a model of the invasive disease endometriosis, we identified A Disintegrin and Metalloproteinase 12 (ADAM12) as a protease implicated in HB-EGF shedding. We designed a specific inhibitor of ADAM12 based on its recombinant prodomain (PA12), which selectively inhibits ADAM12 but not ADAM10 or ADAM17. In endometriotic cells, PA12 significantly reduced HB-EGF shedding and resultant cellular migration. Overall, specific inhibition of ligand shedding represents a possible alternative to decoy antibodies, especially for ligands such as HB-EGF that exhibit high binding affinity and localized signaling. PMID:26477568

  2. Two Direct Targets of Cytokinin Signaling Regulate Symbiotic Nodulation in Medicago truncatula[W][OA

    Science.gov (United States)

    Ariel, Federico; Brault-Hernandez, Marianne; Laffont, Carole; Huault, Emeline; Brault, Mathias; Plet, Julie; Moison, Michael; Blanchet, Sandrine; Ichanté, Jean Laurent; Chabaud, Mireille; Carrere, Sébastien; Crespi, Martin; Chan, Raquel L.; Frugier, Florian

    2012-01-01

    Cytokinin regulates many aspects of plant development, and in legume crops, this phytohormone is necessary and sufficient for symbiotic nodule organogenesis, allowing them to fix atmospheric nitrogen. To identify direct links between cytokinins and nodule organogenesis, we determined a consensus sequence bound in vitro by a transcription factor (TF) acting in cytokinin signaling, the nodule-enhanced Medicago truncatula Mt RR1 response regulator (RR). Among genes rapidly regulated by cytokinins and containing this so-called RR binding site (RRBS) in their promoters, we found the nodulation-related Type-A RR Mt RR4 and the Nodulation Signaling Pathway 2 (NSP2) TF. Site-directed mutagenesis revealed that RRBS cis-elements in the RR4 and NSP2 promoters are essential for expression during nodule development and for cytokinin induction. Furthermore, a microRNA targeting NSP2 (miR171 h) is also rapidly induced by cytokinins and then shows an expression pattern anticorrelated with NSP2. Other primary targets regulated by cytokinins depending on the Cytokinin Response1 (CRE1) receptor were a cytokinin oxidase/dehydrogenase (CKX1) and a basic Helix-Loop-Helix TF (bHLH476). RNA interference constructs as well as insertion of a Tnt1 retrotransposon in the bHLH gene led to reduced nodulation. Hence, we identified two TFs, NSP2 and bHLH476, as direct cytokinin targets acting at the convergence of phytohormonal and symbiotic cues. PMID:23023168

  3. Targeting cancer stem cells and signaling pathways by phytochemicals: Novel approach for breast cancer therapy.

    Science.gov (United States)

    Dandawate, Prasad R; Subramaniam, Dharmalingam; Jensen, Roy A; Anant, Shrikant

    2016-10-01

    Breast cancer is the most common form of cancer diagnosed in women worldwide and the second leading cause of cancer-related deaths in the USA. Despite the development of newer diagnostic methods, selective as well as targeted chemotherapies and their combinations, surgery, hormonal therapy, radiotherapy, breast cancer recurrence, metastasis and drug resistance are still the major problems for breast cancer. Emerging evidence suggest the existence of cancer stem cells (CSCs), a population of cells with the capacity to self-renew, differentiate and be capable of initiating and sustaining tumor growth. In addition, CSCs are believed to be responsible for cancer recurrence, anticancer drug resistance, and metastasis. Hence, compounds targeting breast CSCs may be better therapeutic agents for treating breast cancer and control recurrence and metastasis. Naturally occurring compounds, mainly phytochemicals have gained immense attention in recent times because of their wide safety profile, ability to target heterogeneous populations of cancer cells as well as CSCs, and their key signaling pathways. Therefore, in the present review article, we summarize our current understanding of breast CSCs and their signaling pathways, and the phytochemicals that affect these cells including curcumin, resveratrol, tea polyphenols (epigallocatechin-3-gallate, epigallocatechin), sulforaphane, genistein, indole-3-carbinol, 3, 3'-di-indolylmethane, vitamin E, retinoic acid, quercetin, parthenolide, triptolide, 6-shogaol, pterostilbene, isoliquiritigenin, celastrol, and koenimbin. These phytochemicals may serve as novel therapeutic agents for breast cancer treatment and future leads for drug development. Copyright © 2016. Published by Elsevier Ltd.

  4. Two direct targets of cytokinin signaling regulate symbiotic nodulation in Medicago truncatula.

    Science.gov (United States)

    Ariel, Federico; Brault-Hernandez, Marianne; Laffont, Carole; Huault, Emeline; Brault, Mathias; Plet, Julie; Moison, Michael; Blanchet, Sandrine; Ichanté, Jean Laurent; Chabaud, Mireille; Carrere, Sébastien; Crespi, Martin; Chan, Raquel L; Frugier, Florian

    2012-09-01

    Cytokinin regulates many aspects of plant development, and in legume crops, this phytohormone is necessary and sufficient for symbiotic nodule organogenesis, allowing them to fix atmospheric nitrogen. To identify direct links between cytokinins and nodule organogenesis, we determined a consensus sequence bound in vitro by a transcription factor (TF) acting in cytokinin signaling, the nodule-enhanced Medicago truncatula Mt RR1 response regulator (RR). Among genes rapidly regulated by cytokinins and containing this so-called RR binding site (RRBS) in their promoters, we found the nodulation-related Type-A RR Mt RR4 and the Nodulation Signaling Pathway 2 (NSP2) TF. Site-directed mutagenesis revealed that RRBS cis-elements in the RR4 and NSP2 promoters are essential for expression during nodule development and for cytokinin induction. Furthermore, a microRNA targeting NSP2 (miR171 h) is also rapidly induced by cytokinins and then shows an expression pattern anticorrelated with NSP2. Other primary targets regulated by cytokinins depending on the Cytokinin Response1 (CRE1) receptor were a cytokinin oxidase/dehydrogenase (CKX1) and a basic Helix-Loop-Helix TF (bHLH476). RNA interference constructs as well as insertion of a Tnt1 retrotransposon in the bHLH gene led to reduced nodulation. Hence, we identified two TFs, NSP2 and bHLH476, as direct cytokinin targets acting at the convergence of phytohormonal and symbiotic cues.

  5. ANP-NPRA signaling pathway--a potential therapeutic target for the treatment of malignancy.

    Science.gov (United States)

    Zhao, Zhilong; Zhang, Jia; Li, Min; Yang, Ya; Sun, Kai; Wang, Jiansheng

    2013-01-01

    It was well established that the atrial natriuretic peptide (ANP)/natriuretic peptide receptor-A (NPRA) signaling pathway controls natriuretic, diuretic, vasorelaxant, and anti-proliferative responses in the regulation of the human cardiovascular system by previous studies. Yet in recent years, more and more evidence has shown that the ANP/NPRA signaling pathway plays an important role in human cancer. For example, NPRA is abundantly expressed on tumorigenic mouse and human prostate cancer (PCa) cells, but not in nontumorigenic prostate epithelial cells and down-regulation of NPRA-induced apoptosis in PCa cells. Dexamethasone can increase the expression of ANP markedly, and that is the reason why dexamethasone is the cornerstone in the treatment of multiple myeloma. NPRA deficiency can substantially protect C57BL/6 mice from lung, skin, and ovarian cancers. These results strongly suggest ANP and NPRA may play an anti-cancer and carcinogenesis role, respectively, and this signaling pathway could be a more potent target for cancer therapy. In light of these new insights, this review will summarize the structures, functions, and their regulation by cell signaling, and their different impacts on tumors.

  6. Targeting the Microglial Signaling Pathways: New Insights in the Modulation of Neuropathic Pain

    Science.gov (United States)

    Popiolek-Barczyk, Katarzyna; Mika, Joanna

    2016-01-01

    The microglia, once thought only to be supporting cells of the central nervous system (CNS), are now recognized to play essential roles in many pathologies. Many studies within the last decades indicated that the neuro-immune interaction underlies the generation and maintenance of neuropathic pain. Through a large number of receptors and signaling pathways, the microglial cells communicate with neurons, astrocytes and other cells, including those of the immune system. A disturbance or loss of CNS homeostasis causes rapid responses of the microglia, which undergo a multistage activation process. The activated microglia change their cell shapes and gene expression profiles, which induce proliferation, migration, and the production of pro- or antinociceptive factors. The cells release a large number of mediators that can act in a manner detrimental or beneficial to the surrounding cells and can indirectly alter the nociceptive signals. This review discusses the most important microglial intracellular signaling cascades (MAPKs, NF-κB, JAK/STAT, PI3K/Akt) that are essential for neuropathic pain development and maintenance. Our objective was to identify new molecular targets that may result in the development of powerful tools to control the signaling associated with neuropathic pain. PMID:27281131

  7. [Cell signaling pathways interaction in cellular proliferation: Potential target for therapeutic interventionism].

    Science.gov (United States)

    Valdespino-Gómez, Víctor Manuel; Valdespino-Castillo, Patricia Margarita; Valdespino-Castillo, Víctor Edmundo

    2015-01-01

    Nowadays, cellular physiology is best understood by analysing their interacting molecular components. Proteins are the major components of the cells. Different proteins are organised in the form of functional clusters, pathways or networks. These molecules are ordered in clusters of receptor molecules of extracellular signals, transducers, sensors and biological response effectors. The identification of these intracellular signaling pathways in different cellular types has required a long journey of experimental work. More than 300 intracellular signaling pathways have been identified in human cells. They participate in cell homeostasis processes for structural and functional maintenance. Some of them participate simultaneously or in a nearly-consecutive progression to generate a cellular phenotypic change. In this review, an analysis is performed on the main intracellular signaling pathways that take part in the cellular proliferation process, and the potential use of some components of these pathways as target for therapeutic interventionism are also underlined. Copyright © 2015 Academia Mexicana de Cirugía A.C. Published by Masson Doyma México S.A. All rights reserved.

  8. Targeting Sonic Hedgehog Signaling by Compounds and Derivatives from Natural Products

    Directory of Open Access Journals (Sweden)

    Yu-Chuen Huang

    2013-01-01

    Full Text Available Cancer stem cells (CSCs are a major cause of cancer treatment failure, relapse, and drug resistance and are known to be responsible for cancer cell invasion and metastasis. The Sonic hedgehog (Shh signaling pathway is crucial to embryonic development. Intriguingly, the aberrant activation of the Shh pathway plays critical roles in developing CSCs and leads to angiogenesis, migration, invasion, and metastasis. Natural compounds and chemical structure modified derivatives from complementary and alternative medicine have received increasing attention as cancer chemopreventives, and their antitumor effects have been demonstrated both in vitro and in vivo. However, reports for their bioactivity against CSCs and specifically targeting Shh signaling remain limited. In this review, we summarize investigations of the compounds cyclopamine, curcumin, epigallocatechin-3-gallate, genistein, resveratrol, zerumbone, norcantharidin, and arsenic trioxide, with a focus on Shh signaling blockade. Given that Shh signaling antagonism has been clinically proven as effective strategy against CSCs, this review may be exploitable for development of novel anticancer agents from complementary and alternative medicine.

  9. The Role of Mammalian Target of Rapamycin (mTOR) in Insulin Signaling.

    Science.gov (United States)

    Yoon, Mee-Sup

    2017-10-27

    The mammalian target of rapamycin (mTOR) is a serine/threonine kinase that controls a wide spectrum of cellular processes, including cell growth, differentiation, and metabolism. mTOR forms two distinct multiprotein complexes known as mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2), which are characterized by the presence of raptor and rictor, respectively. mTOR controls insulin signaling by regulating several downstream components such as growth factor receptor-bound protein 10 (Grb10), insulin receptor substrate (IRS-1), F-box/WD repeat-containing protein 8 (Fbw8), and insulin like growth factor 1 receptor/insulin receptor (IGF-IR/IR). In addition, mTORC1 and mTORC2 regulate each other through a feedback loop to control cell growth. This review outlines the current understanding of mTOR regulation in insulin signaling in the context of whole body metabolism.

  10. Targeting CB2-GPR55 receptor heteromers modulates cancer cell signaling.

    Science.gov (United States)

    Moreno, Estefanía; Andradas, Clara; Medrano, Mireia; Caffarel, María M; Pérez-Gómez, Eduardo; Blasco-Benito, Sandra; Gómez-Cañas, María; Pazos, M Ruth; Irving, Andrew J; Lluís, Carme; Canela, Enric I; Fernández-Ruiz, Javier; Guzmán, Manuel; McCormick, Peter J; Sánchez, Cristina

    2014-08-08

    The G protein-coupled receptors CB2 (CB2R) and GPR55 are overexpressed in cancer cells and human tumors. Because a modulation of GPR55 activity by cannabinoids has been suggested, we analyzed whether this receptor participates in cannabinoid effects on cancer cells. Here we show that CB2R and GPR55 form heteromers in cancer cells, that these structures possess unique signaling properties, and that modulation of these heteromers can modify the antitumoral activity of cannabinoids in vivo. These findings unveil the existence of previously unknown signaling platforms that help explain the complex behavior of cannabinoids and may constitute new targets for therapeutic intervention in oncology. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. The interaction between signal regulatory protein alpha (SIRPα) and CD47: structure, function, and therapeutic target.

    Science.gov (United States)

    Barclay, A Neil; Van den Berg, Timo K

    2014-01-01

    CD47 is a broadly expressed membrane protein that interacts with the myeloid inhibitory immunoreceptor SIRPα (also termed CD172a or SHPS-1). SIRPα is the prototypic member of the SIRP paired receptor family of closely related SIRP proteins. Engagement of SIRPα by CD47 provides a downregulatory signal that inhibits host cell phagocytosis, and CD47 therefore functions as a "don't-eat-me" signal. Here, we discuss recent structural analysis of CD47-SIRPα interactions and implications of this for the function and evolution of SIRPα and paired receptors in general. Furthermore, we review the proposed roles of CD47-SIRPα interactions in phagocytosis, (auto)immunity, and host defense, as well as its potential significance as a therapeutic target in cancer and inflammation and for improving graft survival in xenotransplantation.

  12. Scopolamine rapidly increases mammalian target of rapamycin complex 1 signaling, synaptogenesis, and antidepressant behavioral responses.

    Science.gov (United States)

    Voleti, Bhavya; Navarria, Andrea; Liu, Rong-Jian; Banasr, Mounira; Li, Nanxin; Terwilliger, Rose; Sanacora, Gerard; Eid, Tore; Aghajanian, George; Duman, Ronald S

    2013-11-15

    Clinical studies report that scopolamine, an acetylcholine muscarinic receptor antagonist, produces rapid antidepressant effects in depressed patients, but the mechanisms underlying the therapeutic response have not been determined. The present study examines the role of the mammalian target of rapamycin complex 1 (mTORC1) and synaptogenesis, which have been implicated in the rapid actions of N-methyl-D-aspartate receptor antagonists. The influence of scopolamine on mTORC1 signaling was determined by analysis of the phosphorylated and activated forms of mTORC1 signaling proteins in the prefrontal cortex (PFC). The numbers and function of spine synapses were analyzed by whole cell patch clamp recording and two-photon image analysis of PFC neurons. The actions of scopolamine were examined in the forced swim test in the absence or presence of selective mTORC1 and glutamate receptor inhibitors. The results demonstrate that a single, low dose of scopolamine rapidly increases mTORC1 signaling and the number and function of spine synapses in layer V pyramidal neurons in the PFC. Scopolamine administration also produces an antidepressant response in the forced swim test that is blocked by pretreatment with the mTORC1 inhibitor or by a glutamate alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor antagonist. Taken together, the results demonstrate that the antidepressant actions of scopolamine require mTORC1 signaling and are associated with increased glutamate transmission, and synaptogenesis, similar to N-methyl-D-aspartate receptor antagonists. These findings provide novel targets for safer and more efficacious rapid-acting antidepressant agents. © 2013 Society of Biological Psychiatry.

  13. TAK1 MAPKKK mediates TGF-β signaling by targeting SnoN oncoprotein for degradation

    Science.gov (United States)

    Kajino, Taisuke; Omori, Emily; Ishii, Shunsuke; Matsumoto, Kunihiro; Ninomiya-Tsuji, Jun

    2007-01-01

    Transforming growth factor-β (TGF-β) regulates a variety of physiologic processes through essential intracellular mediators Smads. The SnoN oncoprotein is an inhibitor of TGF-β signaling. SnoN recruits transcriptional repressor complex to block Smad-dependent transcriptional activation of TGF-β-responsive genes. Following TGF-β stimulation, SnoN is rapidly degraded, thereby allowing the activation of TGF-β target genes. Here, we report the role of TAK1 as a SnoN protein kinase. TAK1 interacts with and phosphorylates SnoN, and this phosphorylation regulates the stability of SnoN. Inactivation of TAK1 prevents TGF-β-induced SnoN degradation, and impairs induction of the TGF-β-responsive genes. These data suggest that TAK1 modulates TGF-β dependent cellular responses by targeting SnoN for degradation. PMID:17276978

  14. Not just an antibiotic target: Exploring the role of type I signal peptidase in bacterial virulence.

    Science.gov (United States)

    Walsh, Shawn I; Craney, Arryn; Romesberg, Floyd E

    2016-12-15

    The looming antibiotic crisis has prompted the development of new strategies towards fighting infection. Traditional antibiotics target bacterial processes essential for viability, whereas proposed antivirulence approaches rely on the inhibition of factors that are required only for the initiation and propagation of infection within a host. Although antivirulence compounds have yet to prove their efficacy in the clinic, bacterial signal peptidase I (SPase) represents an attractive target in that SPase inhibitors exhibit broad-spectrum antibiotic activity, but even at sub-MIC doses also impair the secretion of essential virulence factors. The potential consequences of SPase inhibition on bacterial virulence have not been thoroughly examined, and are explored within this review. In addition, we review growing evidence that SPase has relevant biological functions outside of mediating secretion, and discuss how the inhibition of these functions may be clinically significant. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Targeting gastrin-releasing peptide suppresses neuroblastoma progression via upregulation of PTEN signaling.

    Directory of Open Access Journals (Sweden)

    Pritha Paul

    Full Text Available We have previously demonstrated the role of gastrin-releasing peptide (GRP as an autocrine growth factor for neuroblastoma. Here, we report that GRP silencing regulates cell signaling involved in the invasion-metastasis cascade. Using a doxycycline inducible system, we demonstrate that GRP silencing decreased anchorage-independent growth, inhibited migration and neuroblastoma cell-mediated angiogenesis in vitro, and suppressed metastasis in vivo. Targeted inhibition of GRP decreased the mRNA levels of oncogenes responsible for neuroblastoma progression. We also identified PTEN/AKT signaling as a key mediator of the tumorigenic properties of GRP in neuroblastoma cells. Interestingly, PTEN overexpression decreased GRP-mediated migration and angiogenesis; a novel role for this, otherwise, understated tumor suppressor in neuroblastoma. Furthermore, activation of AKT (pAKT positively correlated with neuroblastoma progression in an in vivo tumor-metastasis model. PTEN expression was slightly decreased in metastatic lesions. A similar phenomenon was observed in human neuroblastoma sections, where, early-stage localized tumors had a higher PTEN expression relative to pAKT; however, an inverse expression pattern was observed in liver lesions. Taken together, our results argue for a dual purpose of targeting GRP in neuroblastoma--1 decreasing expression of critical oncogenes involved in tumor progression, and 2 enhancing activation of tumor suppressor genes to treat aggressive, advanced-stage disease.

  16. Targeting notch signaling pathway in cancer: clinical development advances and challenges.

    Science.gov (United States)

    Takebe, Naoko; Nguyen, Dat; Yang, Sherry X

    2014-02-01

    Notch signaling plays an important role in development and cell fate determination, and it is deregulated in human hematologic malignancies and solid tumors. This review includes a brief introduction of the relevant pathophysiology of Notch signaling pathway and primarily focuses on the clinical development of promising agents that either obstruct Notch receptor cleavages such as γ-secretase inhibitors (GSIs) or interfere with the Notch ligand-receptor interaction by monoclonal antibodies (mAbs). Antitumor activity by GSIs and mAbs administered as single agent in early phases of clinical trials has been observed in advanced or metastatic thyroid cancer, non-small cell lung cancer, intracranial tumors, sarcoma or desmoid tumors, colorectal cancer with neuroendocrine features, melanoma and ovarian cancer. A number of mechanism-based adverse events particularly gastrointestinal toxicities emerged and mitigation strategies are developed after testing multiple GSIs and Notch targeting mAbs. We also discuss pharmacodynamic biomarkers in conjunction with methods of assessment of the molecular target inhibition validation. Biomarkers of efficacy or benefit may be of importance for a successful development of this class of drugs. Published by Elsevier Inc.

  17. Lipid raft-mediated Akt signaling as a therapeutic target in mantle cell lymphoma

    Science.gov (United States)

    Reis-Sobreiro, M; Roué, G; Moros, A; Gajate, C; de la Iglesia-Vicente, J; Colomer, D; Mollinedo, F

    2013-01-01

    Recent evidence shows that lipid raft membrane domains modulate both cell survival and death. Here, we have found that the phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathway is present in the lipid rafts of mantle cell lymphoma (MCL) cells, and this location seems to be critical for full activation and MCL cell survival. The antitumor lipids (ATLs) edelfosine and perifosine target rafts, and we found that ATLs exerted in vitro and in vivo antitumor activity against MCL cells by displacing Akt as well as key regulatory kinases p-PDK1 (phosphatidylinositol-dependent protein kinase 1), PI3K and mTOR (mammalian TOR) from lipid rafts. This raft reorganization led to Akt dephosphorylation, while proapoptotic Fas/CD95 death receptor was recruited into rafts. Raft integrity was critical for Ser473 Akt phosphorylation. ATL-induced apoptosis appeared to correlate with the basal Akt phosphorylation status in MCL cell lines and primary cultures, and could be potentiated by the PI3K inhibitor wortmannin, or inhibited by the Akt activator pervanadate. Classical Akt inhibitors induced apoptosis in MCL cells. Microenvironmental stimuli, such as CD40 ligation or stromal cell contact, did not prevent ATL-induced apoptosis in MCL cell lines and patient-derived cells. These results highlight the role of raft-mediated PI3K/Akt signaling in MCL cell survival and chemotherapy, thus becoming a new target for MCL treatment. PMID:23727661

  18. Candida albicans Heat Shock Proteins and Hsps-Associated Signaling Pathways as Potential Antifungal Targets

    Science.gov (United States)

    Gong, Ying; Li, Tao; Yu, Cuixiang; Sun, Shujuan

    2017-01-01

    In recent decades, the incidence of invasive fungal infections has increased notably. Candida albicans (C. albicans), a common opportunistic fungal pathogen that dwells on human mucosal surfaces, can cause fungal infections, especially in immunocompromised and high-risk surgical patients. In addition, the wide use of antifungal agents has likely contributed to resistance of C. albicans to traditional antifungal drugs, increasing the difficulty of treatment. Thus, it is urgent to identify novel antifungal drugs to cope with C. albicans infections. Heat shock proteins (Hsps) exist in most organisms and are expressed in response to thermal stress. In C. albicans, Hsps control basic physiological activities or virulence via interaction with a variety of diverse regulators of cellular signaling pathways. Moreover, it has been demonstrated that Hsps confer drug resistance to C. albicans. Many studies have shown that disrupting the normal functions of C. albicans Hsps inhibits fungal growth or reverses the tolerance of C. albicans to traditional antifungal drugs. Here, we review known functions of the diverse Hsp family, Hsp-associated intracellular signaling pathways and potential antifungal targets based on these pathways in C. albicans. We hope this review will aid in revealing potential new roles of C. albicans Hsps in addition to canonical heat stress adaptions and provide more insight into identifying potential novel antifungal targets. PMID:29312897

  19. Functions of paracrine PDGF signaling in the proangiogenic tumor stroma revealed by pharmacological targeting.

    Directory of Open Access Journals (Sweden)

    Kristian Pietras

    2008-01-01

    Full Text Available BACKGROUND: Important support functions, including promotion of tumor growth, angiogenesis, and invasion, have been attributed to the different cell types populating the tumor stroma, i.e., endothelial cells, cancer-associated fibroblasts, pericytes, and infiltrating inflammatory cells. Fibroblasts have long been recognized inside carcinomas and are increasingly implicated as functional participants. The stroma is prominent in cervical carcinoma, and distinguishable from nonmalignant tissue, suggestive of altered (tumor-promoting functions. We postulated that pharmacological targeting of putative stromal support functions, in particular those of cancer-associated fibroblasts, could have therapeutic utility, and sought to assess the possibility in a pre-clinical setting. METHODS AND FINDINGS: We used a genetically engineered mouse model of cervical carcinogenesis to investigate platelet-derived growth factor (PDGF receptor signaling in cancer-associated fibroblasts and pericytes. Pharmacological blockade of PDGF receptor signaling with the clinically approved kinase inhibitor imatinib slowed progression of premalignant cervical lesions in this model, and impaired the growth of preexisting invasive carcinomas. Inhibition of stromal PDGF receptors reduced proliferation and angiogenesis in cervical lesions through a mechanism involving suppression of expression of the angiogenic factor fibroblast growth factor 2 (FGF-2 and the epithelial cell growth factor FGF-7 by cancer-associated fibroblasts. Treatment with neutralizing antibodies to the PDGF receptors recapitulated these effects. A ligand trap for the FGFs impaired the angiogenic phenotype similarly to imatinib. Thus PDGF ligands expressed by cancerous epithelia evidently stimulate PDGFR-expressing stroma to up-regulate FGFs, promoting angiogenesis and epithelial proliferation, elements of a multicellular signaling network that elicits functional capabilities in the tumor microenvironment

  20. Targeting FLT3-ITD signaling mediates ceramide-dependent mitophagy and attenuates drug resistance in AML.

    Science.gov (United States)

    Dany, Mohammed; Gencer, Salih; Nganga, Rose; Thomas, Raquela J; Oleinik, Natalia; Baron, Kyla D; Szulc, Zdzislaw M; Ruvolo, Peter; Kornblau, Steven; Andreeff, Michael; Ogretmen, Besim

    2016-10-13

    Signaling pathways regulated by mutant Fms-like tyrosine kinase 3 (FLT3)-internal tandem duplication (ITD), which mediate resistance to acute myeloid leukemia (AML) cell death, are poorly understood. Here, we reveal that pro-cell death lipid ceramide generation is suppressed by FLT3-ITD signaling. Molecular or pharmacologic inhibition of FLT3-ITD reactivated ceramide synthesis, selectively inducing mitophagy and AML cell death. Mechanistically, FLT3-ITD targeting induced ceramide accumulation on the outer mitochondrial membrane, which then directly bound autophagy-inducing light chain 3 (LC3), involving its I35 and F52 residues, to recruit autophagosomes for execution of lethal mitophagy. Short hairpin RNA (shRNA)-mediated knockdown of LC3 prevented AML cell death in response to FLT3-ITD inhibition by crenolanib, which was restored by wild-type (WT)-LC3, but not mutants of LC3 with altered ceramide binding (I35A-LC3 or F52A-LC3). Mitochondrial ceramide accumulation and lethal mitophagy induction in response to FLT3-ITD targeting was mediated by dynamin-related protein 1 (Drp1) activation via inhibition of protein kinase A-regulated S637 phosphorylation, resulting in mitochondrial fission. Inhibition of Drp1 prevented ceramide-dependent lethal mitophagy, and reconstitution of WT-Drp1 or phospho-null S637A-Drp1 but not its inactive phospho-mimic mutant (S637D-Drp1), restored mitochondrial fission and mitophagy in response to crenolanib in FLT3-ITD+ AML cells expressing stable shRNA against endogenous Drp1. Moreover, activating FLT3-ITD signaling in crenolanib-resistant AML cells suppressed ceramide-dependent mitophagy and prevented cell death. FLT3-ITD+ AML drug resistance is attenuated by LCL-461, a mitochondria-targeted ceramide analog drug, in vivo, which also induced lethal mitophagy in human AML blasts with clinically relevant FLT3 mutations. Thus, these data reveal a novel mechanism which regulates AML cell death by ceramide-dependent mitophagy in response to

  1. Natural compounds targeting major cell signaling pathways: a novel paradigm for osteosarcoma therapy.

    Science.gov (United States)

    Angulo, Pablo; Kaushik, Gaurav; Subramaniam, Dharmalingam; Dandawate, Prasad; Neville, Kathleen; Chastain, Katherine; Anant, Shrikant

    2017-01-07

    Osteosarcoma is the most common primary bone cancer affecting children and adolescents worldwide. Despite an incidence of three cases per million annually, it accounts for an inordinate amount of morbidity and mortality. While the use of chemotherapy (cisplatin, doxorubicin, and methotrexate) in the last century initially resulted in marginal improvement in survival over surgery alone, survival has not improved further in the past four decades. Patients with metastatic osteosarcoma have an especially poor prognosis, with only 30% overall survival. Hence, there is a substantial need for new therapies. The inability to control the metastatic progression of this localized cancer stems from a lack of complete knowledge of the biology of osteosarcoma. Consequently, there has been an aggressive undertaking of scientific investigation of various signaling pathways that could be instrumental in understanding the pathogenesis of osteosarcoma. Here, we review these cancer signaling pathways, including Notch, Wnt, Hedgehog, phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/AKT, and JAK/STAT, and their specific role in osteosarcoma. In addition, we highlight numerous natural compounds that have been documented to target these pathways effectively, including curcumin, diallyl trisulfide, resveratrol, apigenin, cyclopamine, and sulforaphane. We elucidate through references that these natural compounds can induce cancer signaling pathway manipulation and possibly facilitate new treatment modalities for osteosarcoma.

  2. Natural compounds targeting major cell signaling pathways: a novel paradigm for osteosarcoma therapy

    Directory of Open Access Journals (Sweden)

    Pablo Angulo

    2017-01-01

    Full Text Available Abstract Osteosarcoma is the most common primary bone cancer affecting children and adolescents worldwide. Despite an incidence of three cases per million annually, it accounts for an inordinate amount of morbidity and mortality. While the use of chemotherapy (cisplatin, doxorubicin, and methotrexate in the last century initially resulted in marginal improvement in survival over surgery alone, survival has not improved further in the past four decades. Patients with metastatic osteosarcoma have an especially poor prognosis, with only 30% overall survival. Hence, there is a substantial need for new therapies. The inability to control the metastatic progression of this localized cancer stems from a lack of complete knowledge of the biology of osteosarcoma. Consequently, there has been an aggressive undertaking of scientific investigation of various signaling pathways that could be instrumental in understanding the pathogenesis of osteosarcoma. Here, we review these cancer signaling pathways, including Notch, Wnt, Hedgehog, phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K/AKT, and JAK/STAT, and their specific role in osteosarcoma. In addition, we highlight numerous natural compounds that have been documented to target these pathways effectively, including curcumin, diallyl trisulfide, resveratrol, apigenin, cyclopamine, and sulforaphane. We elucidate through references that these natural compounds can induce cancer signaling pathway manipulation and possibly facilitate new treatment modalities for osteosarcoma.

  3. Mammalian target of rapamycin complex 1 signalling is essential for germinal centre reaction.

    Science.gov (United States)

    Li, Bingshou; Li, Zhirong; Wang, Pengcheng; Huang, Qizhao; Xu, Lifan; He, Ran; Ye, Lilin; Bai, Qiang

    2017-10-01

    The mammalian target of rapamycin (mTOR) is a serine-threonine kinase that has been shown to be essential for the differentiation and function of various immune cells. Earlier in vitro studies showed that mTOR signalling regulates B-cell biology by supporting their activation and proliferation. However, how mTOR signalling temporally regulates in vivo germinal centre B (GCB) cell development and differentiation into short-lived plasma cells, long-lived plasma cells and memory cells is still not well understood. In this study, we used a combined conditional/inducible knock-out system to investigate the temporal regulation of mTOR complex 1 (mTORC1) in the GCB cell response to acute lymphocytic choriomeningitis virus infection by deleting Raptor, a main component of mTORC1, specifically in B cells in pre- and late GC phase. Early Raptor deficiency strongly inhibited GCB cell proliferation and differentiation and plasma cell differentiation. Nevertheless, late GC Raptor deficiency caused only decreases in the size of memory B cells and long-lived plasma cells through poor maintenance of GCB cells, but it did not change their differentiation. Collectively, our data revealed that mTORC1 signalling supports GCB cell responses at both early and late GC phases during viral infection but does not regulate GCB cell differentiation into memory B cells and plasma cells at the late GC stage. © 2017 John Wiley & Sons Ltd.

  4. Dietary phytochemicals for possible preventive and therapeutic option of uterine fibroids: Signaling pathways as target.

    Science.gov (United States)

    Islam, Md Soriful; Segars, James H; Castellucci, Mario; Ciarmela, Pasquapina

    2017-02-01

    A growing interest has emerged on dietary phytochemicals to control diverse pathological conditions. Unfortunately, dietary phytochemical research in uterine fibroids is still under construction. Uterine fibroids/leiomyomas are benign tumors developing from the myometrium of the uterus in premenopausal women. They may occur in more than 70% of women, and approximately 25% of women show clinically significant symptoms. These include heavy and prolonged menstrual bleeding, pelvic pressure (urinary frequency, incontinence, and difficulty with urination), pelvic pain, pelvic mass, infertility, and reproductive dysfunction. Due to lack of medical treatments surgery has been definitive choice for fibroid management. Moreover, surgery negatively affects women's quality of life, and its associated cost appears to be expensive. The molecular mechanism of fibroids development and growth is not fully elucidated. However, accumulated evidence shows that several signaling pathways, including Smad 2/3, PI3K/AKT/mTOR, ERK 1/2 and β-catenin are involved in the leiomyoma pathogenesis, indicating that they could serve as targets for prevention and/or treatment of this tumor. Therefore, in this review, we discuss the involvement of signaling pathways in leiomyoma development and growth, and introduce some potential dietary phytochemicals that could modulate those signaling pathways. Copyright © 2016 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  5. Neuronal target identification requires AHA-1-mediated fine-tuning of Wnt signaling in C. elegans.

    Directory of Open Access Journals (Sweden)

    Jingyan Zhang

    2013-06-01

    Full Text Available Electrical synaptic transmission through gap junctions is a vital mode of intercellular communication in the nervous system. The mechanism by which reciprocal target cells find each other during the formation of gap junctions, however, is poorly understood. Here we show that gap junctions are formed between BDU interneurons and PLM mechanoreceptors in C. elegans and the connectivity of BDU with PLM is influenced by Wnt signaling. We further identified two PAS-bHLH family transcription factors, AHA-1 and AHR-1, which function cell-autonomously within BDU and PLM to facilitate the target identification process. aha-1 and ahr-1 act genetically upstream of cam-1. CAM-1, a membrane-bound receptor tyrosine kinase, is present on both BDU and PLM cells and likely serves as a Wnt antagonist. By binding to a cis-regulatory element in the cam-1 promoter, AHA-1 enhances cam-1 transcription. Our study reveals a Wnt-dependent fine-tuning mechanism that is crucial for mutual target cell identification during the formation of gap junction connections.

  6. Targeting protein kinase-b3 (akt3) signaling in melanoma.

    Science.gov (United States)

    Madhunapantula, SubbaRao V; Robertson, Gavin P

    2017-03-01

    Deregulated Akt activity leading to apoptosis inhibition, enhanced proliferation and drug resistance has been shown to be responsible for 35-70% of advanced metastatic melanomas. Of the three isoforms, the majority of melanomas have elevated Akt3 expression and activity. Hence, potent inhibitors targeting Akt are urgently required, which is possible only if (a) the factors responsible for the failure of Akt inhibitors in clinical trials is known; and (b) the information pertaining to synergistically acting targeted therapeutics is available. Areas covered: This review provides a brief introduction of the PI3K-Akt signaling pathway and its role in melanoma development. In addition, the functional role of key Akt pathway members such as PRAS40, GSK3 kinases, WEE1 kinase in melanoma development are discussed together with strategies to modulate these targets. Efficacy and safety of Akt inhibitors is also discussed. Finally, the mechanism(s) through which Akt leads to drug resistance is discussed in this expert opinion review. Expert opinion: Even though Akt play key roles in melanoma tumor progression, cell survival and drug resistance, many gaps still exist that require further understanding of Akt functions, especially in the (a) metastatic spread; (b) circulating melanoma cells survival; and (c) melanoma stem cells growth.

  7. Evidence for ubiquitin-regulated nuclear and subnuclear trafficking among Paramyxovirinae matrix proteins.

    Directory of Open Access Journals (Sweden)

    Mickey Pentecost

    2015-03-01

    Full Text Available The paramyxovirus matrix (M protein is a molecular scaffold required for viral morphogenesis and budding at the plasma membrane. Transient nuclear residence of some M proteins hints at non-structural roles. However, little is known regarding the mechanisms that regulate the nuclear sojourn. Previously, we found that the nuclear-cytoplasmic trafficking of Nipah virus M (NiV-M is a prerequisite for budding, and is regulated by a bipartite nuclear localization signal (NLSbp, a leucine-rich nuclear export signal (NES, and monoubiquitination of the K258 residue within the NLSbp itself (NLSbp-lysine. To define whether the sequence determinants of nuclear trafficking identified in NiV-M are common among other Paramyxovirinae M proteins, we generated the homologous NES and NLSbp-lysine mutations in M proteins from the five major Paramyxovirinae genera. Using quantitative 3D confocal microscopy, we determined that the NES and NLSbp-lysine are required for the efficient nuclear export of the M proteins of Nipah virus, Hendra virus, Sendai virus, and Mumps virus. Pharmacological depletion of free ubiquitin or mutation of the conserved NLSbp-lysine to an arginine, which inhibits M ubiquitination, also results in nuclear and nucleolar retention of these M proteins. Recombinant Sendai virus (rSeV-eGFP bearing the NES or NLSbp-lysine M mutants rescued at similar efficiencies to wild type. However, foci of cells expressing the M mutants displayed marked fusogenicity in contrast to wild type, and infection did not spread. Recombinant Mumps virus (rMuV-eGFP bearing the homologous mutations showed similar defects in viral morphogenesis. Finally, shotgun proteomics experiments indicated that the interactomes of Paramyxovirinae M proteins are significantly enriched for components of the nuclear pore complex, nuclear transport receptors, and nucleolar proteins. We then synthesize our functional and proteomics data to propose a working model for the ubiquitin

  8. Targeting Protein Kinase C Downstream of Growth Factor and Adhesion Signalling

    Directory of Open Access Journals (Sweden)

    Catríona M. Dowling

    2015-07-01

    Full Text Available The signaling outputs of Receptor Tyrosine Kinases, G-protein coupled receptors and integrins converge to mediate key cell process such as cell adhesion, cell migration, cell invasion and cell proliferation. Once activated by their ligands, these cell surface proteins recruit and direct a diverse range of proteins to disseminate the appropriate response downstream of the specific environmental cues. One of the key groups of proteins required to regulate these activities is the family of serine/threonine intracellular kinases called Protein Kinase Cs. The activity and subcellular location of PKCs are mediated by a series of tightly regulated events and is dependent on several posttranslational modifications and the availability of second messengers. Protein Kinase Cs exhibit both pro- and anti-tumorigenic effects making them an interesting target for anti-cancer treatment.

  9. Targeted inhibition of hedgehog signaling by cyclopamine prodrugs for advanced prostate cancer

    Science.gov (United States)

    Kumar, Srinivas K.; Roy, Indrajit; Anchoori, Ravi K.; Fazli, Sarah; Maitra, Anirban; Beachy, Philip A.; Khan, Saeed R.

    2009-01-01

    A promising agent for use in prostate cancer therapy is the Hedgehog (Hh) signaling pathway inhibitor, cyclopamine. This compound, however, has the potential for causing serious side effects in non-tumor tissues. To minimize these bystander toxicities, we have designed and synthesized two novel peptide-cyclopamine conjugates as prostate-specific antigen (PSA)-activated prodrugs for use against prostate cancer. These prodrugs were composed of cyclopamine coupled to one of two peptides (either HSSKLQ or SSKYQ) that can be selectively cleaved by PSA, converting the mature prodrug into an active Hedgehog inhibitor within the malignant cells. Of the two prodrugs, Mu-SSKYQ-Cyclopamine was rapidly hydrolyzed, with a half-life of 3.2 h, upon incubation with the PSA enzyme. Thus, modulating cyclopamine at the secondary amine with PSA-cleavable peptides is a promising strategy for developing prodrugs to target prostate cancer. PMID:18249125

  10. In Vivo Phosphoproteomics Analysis Reveals the Cardiac Targets of β-Adrenergic Receptor Signaling

    DEFF Research Database (Denmark)

    Lundby, Alicia; Andersen, Martin N; Steffensen, Annette B

    2013-01-01

    used quantitative in vivo phosphoproteomics to identify 670 site-specific phosphorylation changes in murine hearts in response to acute treatment with specific βAR agonists. The residues adjacent to the regulated phosphorylation sites exhibited a sequence-specific preference (R......) of the potassium channel KV7.1, increased current amplitude. Our data set represents a quantitative analysis of phosphorylated proteins regulated in vivo upon stimulation of seven-transmembrane receptors, and our findings reveal previously unknown phosphorylation sites that regulate myocardial contractility......-X-X-pS/T), and integrative analysis of sequence motifs and interaction networks suggested that the kinases AMPK (adenosine 5'-monophosphate-activated protein kinase), Akt, and mTOR (mammalian target of rapamycin) mediate βAR signaling, in addition to the well-established pathways mediated by PKA (cyclic adenosine...

  11. Molecular Pathways: Cachexia Signaling-A Targeted Approach to Cancer Treatment.

    Science.gov (United States)

    Miyamoto, Yuji; Hanna, Diana L; Zhang, Wu; Baba, Hideo; Lenz, Heinz-Josef

    2016-08-15

    Cancer cachexia is a multifactorial syndrome characterized by an ongoing loss of skeletal muscle mass, which negatively affects quality of life and portends a poor prognosis. Numerous molecular substrates and mechanisms underlie the dysregulation of skeletal muscle synthesis and degradation observed in cancer cachexia, including proinflammatory cytokines (TNFα, IL1, and IL6), and the NF-κB, IGF1/AKT/mTOR, and myostatin/activin-SMAD pathways. Recent preclinical and clinical studies have demonstrated that anti-cachexia drugs (such as MABp1 and soluble receptor antagonist of myostatin/activin) not only prevent muscle wasting but also may prolong overall survival. In this review, we focus on the significance of cachexia signaling in patients with cancer and highlight promising drugs targeting tumor cachexia in clinical development. Clin Cancer Res; 22(16); 3999-4004. ©2016 AACR. ©2016 American Association for Cancer Research.

  12. Targeting Protein Kinase C Downstream of Growth Factor and Adhesion Signalling

    Energy Technology Data Exchange (ETDEWEB)

    Dowling, Catríona M., E-mail: Catriona.Dowling@ul.ie; Kiely, Patrick A., E-mail: Catriona.Dowling@ul.ie [Department of Life Sciences, Materials and Surface Science Institute and Stokes Institute, University of Limerick, Limerick 78666 (Ireland); Health Research Institute (HRI), University of Limerick, Limerick 78666 (Ireland)

    2015-07-15

    The signaling outputs of Receptor Tyrosine Kinases, G-protein coupled receptors and integrins converge to mediate key cell process such as cell adhesion, cell migration, cell invasion and cell proliferation. Once activated by their ligands, these cell surface proteins recruit and direct a diverse range of proteins to disseminate the appropriate response downstream of the specific environmental cues. One of the key groups of proteins required to regulate these activities is the family of serine/threonine intracellular kinases called Protein Kinase Cs. The activity and subcellular location of PKCs are mediated by a series of tightly regulated events and is dependent on several posttranslational modifications and the availability of second messengers. Protein Kinase Cs exhibit both pro- and anti-tumorigenic effects making them an interesting target for anti-cancer treatment.

  13. The quassinoid derivative NBT-272 targets both the AKT and ERK signaling pathways in embryonal tumors.

    Science.gov (United States)

    Castelletti, Deborah; Fiaschetti, Giulio; Di Dato, Valeria; Ziegler, Urs; Kumps, Candy; De Preter, Katleen; Zollo, Massimo; Speleman, Frank; Shalaby, Tarek; De Martino, Daniela; Berg, Thorsten; Eggert, Angelika; Arcaro, Alexandre; Grotzer, Michael A

    2010-12-01

    The quassinoid analogue NBT-272 has been reported to inhibit MYC, thus warranting a further effort 7to better understand its preclinical properties in models of embryonal tumors (ET), a family of childhood malignancies sharing relevant biological and genetic features such as deregulated expression of MYC oncogenes. In our study, NBT-272 displayed a strong antiproliferative activity in vitro that resulted from the combination of diverse biological effects, ranging from G(1)/S arrest of the cell cycle to apoptosis and autophagy. The compound prevented the full activation of both eukaryotic translation initiation factor 4E (eIF4E) and its binding protein 4EBP-1, regulating cap-dependent protein translation. Interestingly, all responses induced by NBT-272 in ET could be attributed to interference with 2 main proproliferative signaling pathways, that is, the AKT and the MEK/extracellular signal-regulated kinase pathways. These findings also suggested that the depleting effect of NBT-272 on MYC protein expression occurred via indirect mechanisms, rather than selective inhibition. Finally, the ability of NBT-272 to arrest tumor growth in a xenograft model of neuroblastoma plays a role in the strong antitumor activity of this compound, both in vitro and in vivo, with its potential to target cell-survival pathways that are relevant for the development and progression of ET.

  14. Tryptanthrin inhibits angiogenesis by targeting the VEGFR2-mediated ERK1/2 signalling pathway.

    Directory of Open Access Journals (Sweden)

    Xuemei Liao

    Full Text Available Angiogenesis is a key step for tumour growth and metastasis, and anti-angiogenesis has been proposed as an important strategy for cancer therapy. Tryptanthrin is a weakly basic alkaloid isolated from the dried roots of medicinal indigo plants and has been shown to possess anti-tumour activities on various cancer cell types. This study aims to investigate the in vitro and in vivo anti-angiogenic activities of tryptanthrin and to unravel its underlying molecular action mechanisms. Our results show that tryptanthrin inhibited the in vitro proliferation, migration, and tube formation of the human microvascular endothelial cells (HMEC-1 in a concentration-dependent manner and significantly suppressed angiogenesis in Matrigel plugs in mice. Mechanistic studies indicated that tryptanthrin reduced the expression of several pro-angiogenic factors (Ang-1, PDGFB and MMP2. Tryptanthrin was also found to suppress the VEGFR2-mediated ERK1/2 signalling pathway in HMEC-1 cells and molecular docking simulation indicated that tryptanthrin could bound to the ATP-binding site of VEGFR2. Collectively, the present study demonstrated that tryptanthrin exhibited both in vitro and in vivo anti-angiogenic activities by targeting the VEGFR2-mediated ERK1/2 signalling pathway and might have therapeutic potential for the treatment of angiogenesis-related diseases.

  15. The Role of Mammalian Target of Rapamycin (mTOR in Insulin Signaling

    Directory of Open Access Journals (Sweden)

    Mee-Sup Yoon

    2017-10-01

    Full Text Available The mammalian target of rapamycin (mTOR is a serine/threonine kinase that controls a wide spectrum of cellular processes, including cell growth, differentiation, and metabolism. mTOR forms two distinct multiprotein complexes known as mTOR complex 1 (mTORC1 and mTOR complex 2 (mTORC2, which are characterized by the presence of raptor and rictor, respectively. mTOR controls insulin signaling by regulating several downstream components such as growth factor receptor-bound protein 10 (Grb10, insulin receptor substrate (IRS-1, F-box/WD repeat-containing protein 8 (Fbw8, and insulin like growth factor 1 receptor/insulin receptor (IGF-IR/IR. In addition, mTORC1 and mTORC2 regulate each other through a feedback loop to control cell growth. This review outlines the current understanding of mTOR regulation in insulin signaling in the context of whole body metabolism.

  16. Accuracy Refinement Algorithm for Mobile Target Location Tracking by Radio Signal Strength Indication Approach

    Science.gov (United States)

    Lau, Erin-Ee-Lin; Chung, Wan-Young

    A novel RSSI (Received Signal Strength Indication) refinement algorithm is proposed to enhance the resolution for indoor and outdoor real-time location tracking system. The proposed refinement algorithm is implemented in two separate phases. During the first phase, called the pre-processing step, RSSI values at different static locations are collected and processed to build a calibrated model for each reference node. Different measurement campaigns pertinent to each parameter in the model are implemented to analyze the sensitivity of RSSI. The propagation models constructed for each reference nodes are needed by the second phase. During the next phase, called the runtime process, real-time tracking is performed. Smoothing algorithm is proposed to minimize the dynamic fluctuation of radio signal received from each reference node when the mobile target is moving. Filtered RSSI values are converted to distances using formula calibrated in the first phase. Finally, an iterative trilateration algorithm is used for position estimation. Experiments relevant to the optimization algorithm are carried out in both indoor and outdoor environments and the results validated the feasibility of proposed algorithm in reducing the dynamic fluctuation for more accurate position estimation.

  17. Quorum sensing communication between bacteria and human cells: signals, targets and functions

    Directory of Open Access Journals (Sweden)

    Angelika eHolm

    2014-06-01

    Full Text Available Both direct and long-range interactions between pathogenic Pseudomonas aeruginosa bacteria and their eukaryotic hosts are important in the outcome of infections. For cell-to-cell communication, these bacteria employ the quorum sensing (QS system to pass on information of the density of the bacterial population and collectively switch on virulence factor production, biofilm formation and resistance development. Thus, QS allows bacteria to behave as a community to perform tasks which would be impossible for individual cells, e.g. to overcome defense and immune systems and establish infections in higher organisms. This review highlights these aspects of QS and our own recent research on how P.aeruginosa communicates with human cells using the small QS signal molecules N-acyl homoserine lactones (AHL. We focus on how this conversation changes the behavior and function of neutrophils, macrophages and epithelial cells and on how the signaling machinery in human cells responsible for the recognition of AHL. Understanding the bacteria-host relationships at both cellular and molecular levels is essential for the identification of new targets and for the development of novel strategies to fight bacterial infections in the future.

  18. A sequential multi-target Mps1 phosphorylation cascade promotes spindle checkpoint signaling.

    Science.gov (United States)

    Ji, Zhejian; Gao, Haishan; Jia, Luying; Li, Bing; Yu, Hongtao

    2017-01-10

    The master spindle checkpoint kinase Mps1 senses kinetochore-microtubule attachment and promotes checkpoint signaling to ensure accurate chromosome segregation. The kinetochore scaffold Knl1, when phosphorylated by Mps1, recruits checkpoint complexes Bub1-Bub3 and BubR1-Bub3 to unattached kinetochores. Active checkpoint signaling ultimately enhances the assembly of the mitotic checkpoint complex (MCC) consisting of BubR1-Bub3, Mad2, and Cdc20, which inhibits the anaphase-promoting complex or cyclosome bound to Cdc20 (APC/C(Cdc20)) to delay anaphase onset. Using in vitro reconstitution, we show that Mps1 promotes APC/C inhibition by MCC components through phosphorylating Bub1 and Mad1. Phosphorylated Bub1 binds to Mad1-Mad2. Phosphorylated Mad1 directly interacts with Cdc20. Mutations of Mps1 phosphorylation sites in Bub1 or Mad1 abrogate the spindle checkpoint in human cells. Therefore, Mps1 promotes checkpoint activation through sequentially phosphorylating Knl1, Bub1, and Mad1. This sequential multi-target phosphorylation cascade makes the checkpoint highly responsive to Mps1 and to kinetochore-microtubule attachment.

  19. Oncogenic role and therapeutic target of leptin signaling in breast cancer and cancer stem cells

    Science.gov (United States)

    Guo, Shanchun; Liu, Mingli; Wang, Guangdi; Torroella-Kouri, Marta; Gonzalez-Perez, Ruben R.

    2012-01-01

    Significant correlations between obesity and incidence of various cancers have been reported. Obesity, considered a mild inflammatory process, is characterized by a high level of secretion of several cytokines from adipose tissue. These molecules have disparate effects, which could be relevant to cancer development. Among the inflammatory molecules, leptin, mainly produced by adipose tissue and overexpressed with its receptor (Ob-R) in cancer cells is the most studied adipokine. Mutations of leptin or Ob-R genes associated with obesity or cancer are rarely found. However, leptin is an anti-apoptotic molecule in many cell types, and its central roles in obesity-related cancers are based on its pro-angiogenic, pro-inflammatory and mitogenic actions. Notably, these leptin actions are commonly reinforced through entangled crosstalk with multiple oncogenes, cytokines and growth factors. Leptin-induced signals comprise several pathways commonly triggered by many cytokines (i.e, canonical: JAK2/STAT; MAPK/ERK1/2 and PI-3K/AKT1 and, non-canonical signaling pathways: PKC, JNK and p38 MAP kinase). Each of these leptin-induced signals is essential to its biological effects on food intake, energy balance, adiposity, immune and endocrine systems, as well as oncogenesis. This review is mainly focused on the current knowledge of the oncogenic role of leptin in breast cancer. Additionally, leptin pro-angiogenic molecular mechanisms and its potential role in breast cancer stem cells will be reviewed. Strict biunivocal binding-affinity and activation of leptin/Ob-R complex makes it a unique molecular target for prevention and treatment of breast cancer, particularly in obesity contexts. PMID:22289780

  20. Interleukin-6/STAT3 signaling as a promising target to improve the efficacy of cancer immunotherapy.

    Science.gov (United States)

    Kitamura, Hidemitsu; Ohno, Yosuke; Toyoshima, Yujiro; Ohtake, Junya; Homma, Shigenori; Kawamura, Hideki; Takahashi, Norihiko; Taketomi, Akinobu

    2017-10-01

    Overcoming the immunosuppressive state in tumor microenvironments is a critical issue for improving the efficacy of cancer immunotherapy. Interleukin (IL)-6, a pleiotropic cytokine, is highly produced in the tumor-bearing host. Previous studies have indicated that IL-6 suppresses the antigen presentation ability of dendritic cells (DC) through activation of signal transducer and activator of transcription 3 (STAT3). Thus, we focused on the precise effect of the IL-6/STAT3 signaling cascade on human DC and the subsequent induction of antitumor T cell immune responses. Tumor-infiltrating CD11b + CD11c + cells isolated from colorectal cancer tissues showed strong induction of the IL-6 gene, downregulated surface expression of human leukocyte antigen (HLA)-DR, and an attenuated T cell-stimulating ability compared with those from peripheral blood mononuclear cells, suggesting that the tumor microenvironment suppresses antitumor effector cells. In vitro experiments revealed that IL-6-mediated STAT3 activation reduced surface expression of HLA-DR on CD14 + monocyte-derived DC. Moreover, we confirmed that cyclooxygenase 2, lysosome protease and arginase activities were involved in the IL-6-mediated downregulation of the surface expression levels of HLA class II on human DC. These findings suggest that IL-6-mediated STAT3 activation in the tumor microenvironment inhibits functional maturation of DC to activate effector T cells, blocking introduction of antitumor immunity in cancers. Therefore, we propose in this review that blockade of the IL-6/STAT3 signaling pathway and target molecules in DC may be a promising strategy to improve the efficacy of immunotherapies for cancer patients. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  1. PACAP neurons in the hypothalamic ventromedial nucleus are targets of central leptin signaling.

    Science.gov (United States)

    Hawke, Zoe; Ivanov, Tina R; Bechtold, David A; Dhillon, Harveen; Lowell, Brad B; Luckman, Simon M

    2009-11-25

    The adipose-derived hormone, leptin, was discovered over 10 years ago, but only now are we unmasking its downstream pathways which lead to reduced energy intake (feeding) and increased energy expenditure (thermogenesis). Recent transgenic models have challenged the long-standing supposition that the hypothalamic arcuate nucleus (Arc) is omnipotent in the central response to leptin, and research focus is beginning to shift to examine roles of extra-arcuate sites. Dhillon et al. (2006) demonstrated that targeted knock out of the signaling form of the leptin receptor (lepr-B) in steroidogenic factor 1 (SF-1) cells of the hypothalamic ventromedial nucleus (VMN) produces obesity of a similar magnitude to the pro-opiomelanocortin (POMC)-driven lepr-B deleted mouse, via a functionally distinct mechanism. These findings reveal that SF-1 cells of the VMN could be equally as important as POMC cells in mediating leptin's anti-obesity effects. However, the identification of molecular and cellular correlates of this relationship remains tantalizingly unknown. Here, we have shown that mRNA expression of the VMN-expressed neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) is regulated according to energy status and that it exerts catabolic effects when administered centrally to mice. Furthermore, we have shown that SF-1 and PACAP mRNAs are colocalized in the VMN, and that leptin signaling via lepr-B is required for normal PACAP expression in these cells. Finally, blocking endogenous central PACAP signaling with the antagonist PACAP(6-38) markedly attenuates leptin-induced hypophagia and hyperthermia in vivo. Thus, it appears that PACAP is an important mediator of central leptin effects on energy balance.

  2. Targeting Glutamatergic Signaling and the PI3 Kinase Pathway to Halt Melanoma Progression

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    Stephen A. Rosenberg

    2015-02-01

    Full Text Available Our group has previously reported that the majority of human melanomas (>60% express the metabotropic glutamate receptor 1 (GRM1 and that the glutamate release inhibitor riluzole, a drug currently used to treat amyotrophic lateral sclerosis, can induce apoptosis in GRM1-expressing melanoma cells. Our group previously reported that in vitro riluzole treatment reduces cell growth in three-dimensional (3D soft agar colony assays by 80% in cells with wildtype phosphoinositide 3-kinase (PI3K pathway activation. However, melanoma cell lines harboring constitutive activating mutations of the PI3K pathway (PTEN and NRAS mutations showed only a 35% to 40% decrease in colony formation in soft agar in the presence of riluzole. In this study, we have continued our preclinical studies of riluzole and its effect on melanoma cells alone and in combination with inhibitors of the PI3 kinase pathway: the AKT inhibitor, API-2, and the mammalian target of rapamycin (mTOR inhibitor, rapamycin. We modeled these combinatorial therapies on various melanoma cell lines in 3D and 2D systems and in vivo. Riluzole combined with mTOR inhibition is more effective at halting melanoma anchorage-independent growth and xenograft tumor progression than either agent alone. PI3K signaling changes associated with this combinatorial treatment shows that 3D (nanoculture modeling of cell signaling more closely resembles in vivo signaling than monolayer models. Riluzole combined with mTOR inhibition is effective at halting tumor cell progression independent of BRAF mutational status. This makes this combinatorial therapy a potentially viable alternative for metastatic melanoma patients who are BRAF WT and are therefore ineligible for vemurafenib therapy.

  3. Targeting the cis-dimerization of LINGO-1 with low MW compounds affects its downstream signalling.

    Science.gov (United States)

    Cobret, L; De Tauzia, M L; Ferent, J; Traiffort, E; Hénaoui, I; Godin, F; Kellenberger, E; Rognan, D; Pantel, J; Bénédetti, H; Morisset-Lopez, S

    2015-02-01

    The transmembrane protein LINGO-1 is a negative regulator in the nervous system mainly affecting axonal regeneration, neuronal survival, oligodendrocyte differentiation and myelination. However, the molecular mechanisms regulating its functions are poorly understood. In the present study, we investigated the formation and the role of LINGO-1 cis-dimers in the regulation of its biological activity. LINGO-1 homodimers were identified in both HEK293 and SH-SY5Y cells using co-immunoprecipitation experiments and BRET saturation analysis. We performed a hypothesis-driven screen for identification of small-molecule protein-protein interaction modulators of LINGO-1 using a BRET-based assay, adapted for screening. The compound identified was further assessed for effects on LINGO-1 downstream signalling pathways using Western blotting analysis and AlphaScreen technology. LINGO-1 was present as homodimers in primary neuronal cultures. LINGO-1 interacted homotypically in cis-orientation and LINGO-1 cis-dimers were formed early during LINGO-1 biosynthesis. A BRET-based assay allowed us to identify phenoxybenzamine as the first conformational modulator of LINGO-1 dimers. In HEK-293 cells, phenoxybenzamine was a positive modulator of LINGO-1 function, increasing the LINGO-1-mediated inhibition of EGF receptor signalling and Erk phosphorylation. Our data suggest that LINGO-1 forms constitutive cis-dimers at the plasma membrane and that low MW compounds affecting the conformational state of these dimers can regulate LINGO-1 downstream signalling pathways. We propose that targeting the LINGO-1 dimerization interface opens a new pharmacological approach to the modulation of its function and provides a new strategy for drug discovery. © 2014 The British Pharmacological Society.

  4. Xenon Protects Against Septic Acute Kidney Injury via miR-21 Target Signaling Pathway*

    Science.gov (United States)

    Jia, Ping; Teng, Jie; Zou, Jianzhou; Fang, Yi; Wu, Xie; Liang, Mingyu

    2015-01-01

    Objectives: Septic acute kidney injury is one of the most common and life-threatening complications in critically ill patients, and there is no approved effective treatment. We have shown xenon provides renoprotection against ischemia-reperfusion injury and nephrotoxicity in rodents via inhibiting apoptosis. Here, we studied the effects of xenon preconditioning on septic acute kidney injury and its mechanism. Design: Experimental animal investigation. Setting: University research laboratory. Subjects: Experiments were performed with male C57BL/6 mice, 10 weeks of age, weighing 20–25 g. Interventions: We induced septic acute kidney injury by a single intraperitoneal injection of Escherichia coli lipopolysaccharide at a dose of 20 mg/kg. Mice were exposed for 2 hours to either 70% xenon or 70% nitrogen, 24 hours before the onset of septic acute kidney injury. In vivo knockdown of miR-21 was performed using locked nucleic acid-modified anti-miR, the role of miR-21 in renal protection conferred by the xenon preconditioning was examined, and miR-21 signaling pathways were analyzed. Measurements and Main Results: Xenon preconditioning provided morphologic and functional renoprotection, characterized by attenuation of renal tubular damage, apoptosis, and a reduction in inflammation. Furthermore, xenon treatment significantly upregulated the expression of miR-21 in kidney, suppressed proinflammatory factor programmed cell death protein 4 expression and nuclear factor-κB activity, and increased interleukin-10 production. Meanwhile, xenon preconditioning also suppressed the expression of proapoptotic protein phosphatase and tensin homolog deleted on chromosome 10, activating protein kinase B signaling pathway, subsequently increasing the expression of antiapoptotic B-cell lymphoma-2, and inhibiting caspase-3 activity. Knockdown of miR-21 upregulated its target effectors programmed cell death protein 4 and phosphatase and tensin homolog deleted on chromosome 10

  5. Xenon Protects Against Septic Acute Kidney Injury via miR-21 Target Signaling Pathway.

    Science.gov (United States)

    Jia, Ping; Teng, Jie; Zou, Jianzhou; Fang, Yi; Wu, Xie; Liang, Mingyu; Ding, Xiaoqiang

    2015-07-01

    Septic acute kidney injury is one of the most common and life-threatening complications in critically ill patients, and there is no approved effective treatment. We have shown xenon provides renoprotection against ischemia-reperfusion injury and nephrotoxicity in rodents via inhibiting apoptosis. Here, we studied the effects of xenon preconditioning on septic acute kidney injury and its mechanism. Experimental animal investigation. University research laboratory. Experiments were performed with male C57BL/6 mice, 10 weeks of age, weighing 20-25 g. We induced septic acute kidney injury by a single intraperitoneal injection of Escherichia coli lipopolysaccharide at a dose of 20 mg/kg. Mice were exposed for 2 hours to either 70% xenon or 70% nitrogen, 24 hours before the onset of septic acute kidney injury. In vivo knockdown of miR-21 was performed using locked nucleic acid-modified anti-miR, the role of miR-21 in renal protection conferred by the xenon preconditioning was examined, and miR-21 signaling pathways were analyzed. Xenon preconditioning provided morphologic and functional renoprotection, characterized by attenuation of renal tubular damage, apoptosis, and a reduction in inflammation. Furthermore, xenon treatment significantly upregulated the expression of miR-21 in kidney, suppressed proinflammatory factor programmed cell death protein 4 expression and nuclear factor-κB activity, and increased interleukin-10 production. Meanwhile, xenon preconditioning also suppressed the expression of proapoptotic protein phosphatase and tensin homolog deleted on chromosome 10, activating protein kinase B signaling pathway, subsequently increasing the expression of antiapoptotic B-cell lymphoma-2, and inhibiting caspase-3 activity. Knockdown of miR-21 upregulated its target effectors programmed cell death protein 4 and phosphatase and tensin homolog deleted on chromosome 10 expression, resulted in an increase in apoptosis, and exacerbated lipopolysaccharide

  6. Targeting the p53 signaling pathway in cancer therapy - The promises, challenges, and perils

    Science.gov (United States)

    Stegh, Alexander H.

    2012-01-01

    Introduction Research over the past three decades has identified p53 as a multifunctional transcription factor, which regulates the expression of >2,500 target genes. p53 impacts myriad, highly diverse cellular processes, including the maintenance of genomic stability and fidelity, metabolism, longevity, and represents one of the most important and extensively studied tumor suppressors. Activated by various stresses, foremost genotoxic damage, hypoxia, heat shock and oncogenic assault, p53 blocks cancer progression by provoking transient or permanent growth arrest, by enabling DNA repair or by advancing cellular death programs. This potent and versatile anti-cancer activity profile, together with genomic and mutational analyses documenting inactivation of p53 in more than 50% of human cancers, motivated drug development efforts to (re-) activate p53 in established tumors. Areas covered In this review the complexities of p53 signaling in cancer are summarized. Current strategies and challenges to restore p53’s tumor suppressive function in established tumors, i.e. adenoviral gene transfer and small molecules to activate p53, to inactivate p53 inhibitors and to restore wild type function of p53 mutant proteins are discussed. Expert opinion It is indubitable that p53 represents an attractive target for the development of anti-cancer therapies. Whether p53 is ‘druggable’, however, remains an area of active research and discussion, as p53 has pro-survival functions and chronic p53 activation accelerates aging, which may compromise the long-term homeostasis of an organism. Thus, the complex biology and dual functions of p53 in cancer prevention and age-related cellular responses pose significant challenges on the development of p53-targeting cancer therapies. PMID:22239435

  7. Notch signaling pathway targeted therapy suppresses tumor progression and metastatic spread in pancreatic cancer.

    Science.gov (United States)

    Yabuuchi, Shinichi; Pai, Shweta G; Campbell, Nathaniel R; de Wilde, Roeland F; De Oliveira, Elizabeth; Korangath, Preethi; Streppel, Mirte M; Rasheed, Zeshaan A; Hidalgo, Manuel; Maitra, Anirban; Rajeshkumar, N V

    2013-07-10

    Pancreatic ductal adenocarcinoma (PDA) remains a lethal human malignancy with historically limited success in treatment. The role of aberrant Notch signaling, which requires the constitutive activation of γ-secretase, in the initiation and progression of PDA is well defined and inhibitors of this pathway are currently in clinical trials. Here we investigated the in vivo therapeutic effect of PF-03084014, a selective γ-secretase inhibitor, alone and in combination with gemcitabine in pancreatic cancer xenografts. PF-03084014 treatment inhibited the cleavage of nuclear Notch 1 intracellular domain and Notch targets Hes-1 and Hey-1. Gemcitabine treatment showed good response but not capable of inducing tumor regressions and targeting the tumor-resident cancer stem cells (CD24(+)CD44(+) and ALDH(+) tumor cells). A combination of PF-03084014 and gemcitabine treatment resulted tumor regression in 3 of 4 subcutaneously implanted xenograft models. PF-03084014, and in combination with gemcitabine reduced putative cancer stem cells, indicating that PF-03084014 target the especially dangerous and resilient cancer stem cells within pancreatic tumors. Tumor re-growth curves plotted after drug treatments demonstrated that the effect of the combination therapy was sustainable than that of gemcitabine. Notably, in a highly aggressive orthotopic model, PF-03084014 and gemcitabine combination was effective in inducing apoptosis, inhibition of tumor cell proliferation and angiogenesis, resulting in the attenuation of primary tumor growth as well as controlling metastatic dissemination, compared to gemcitabine treatment. In summary, our preclinical data suggest that PF-03084014 has greater anti-tumor activity in combination with gemcitabine in PDA and provides rationale for further investigation of this combination in PDA. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  8. Effect of structure on sensing performance of a target induced signaling probe shifting DNA-based (TISPS-DNA) sensor.

    Science.gov (United States)

    Yu, Xiang; Yu, Zhigang; Li, Fengqin; Xu, Yanmei; He, Xunjun; Xu, Lan; Shi, Wenbing; Zhang, Guiling; Yan, Hong

    2017-05-15

    A type of "signal on" displacement-based sensors named target induced signaling probe shifting DNA-based (TISPS-DNA) sensor were developed for a designated DNA detection. The signaling mechanism of the signaling probe (SP) shifting different from the classical conformation/flexibility change mode endows the sensor with high sensitivity. Through using thiolated or no thiolated capturing probe (CP), two 3-probe sensing structures, sensor-1 and sensor-2, were designed and constructed. The systematical comparing research results show that both sensors exhibit some similarities or big differences in sensing performance. On the one hand, the similarity in structures determines the similarity in some aspects of signaling mechanism, background signal, signal changing form, anti-fouling ability and versatility; on the other hand, the slight difference in structures also results in two opposite hybridization modes of gradual increasing resistance and gradual decreasing resistance which can affect the hybridization efficiency between the assistant probe (AP) and the SP, further producing some big differences in sensing performance, for example, apparently different signal enhancement (SE) change, point mutation discrimination ability and response speed. Under the optimized fabrication and detection conditions, both sensors feature high sensitivity for target DNAs with the detection limits of ∼10 fM for sensor-1 and ∼7 fM for sensor-2, respectively. Among many acquired sensing virtues, the sensor-1 shows a peculiar specificity adjustability which is also a highlight in this work. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Targeting Cannabinoid Signaling in the Immune System: “High”-ly Exciting Questions, Possibilities, and Challenges

    Directory of Open Access Journals (Sweden)

    Attila Oláh

    2017-11-01

    Full Text Available It is well known that certain active ingredients of the plants of Cannabis genus, i.e., the “phytocannabinoids” [pCBs; e.g., (−-trans-Δ9-tetrahydrocannabinol (THC, (−-cannabidiol, etc.] can influence a wide array of biological processes, and the human body is able to produce endogenous analogs of these substances [“endocannabinoids” (eCB, e.g., arachidonoylethanolamine (anandamide, AEA, 2-arachidonoylglycerol (2-AG, etc.]. These ligands, together with multiple receptors (e.g., CB1 and CB2 cannabinoid receptors, etc., and a complex enzyme and transporter apparatus involved in the synthesis and degradation of the ligands constitute the endocannabinoid system (ECS, a recently emerging regulator of several physiological processes. The ECS is widely expressed in the human body, including several members of the innate and adaptive immune system, where eCBs, as well as several pCBs were shown to deeply influence immune functions thereby regulating inflammation, autoimmunity, antitumor, as well as antipathogen immune responses, etc. Based on this knowledge, many in vitro and in vivo studies aimed at exploiting the putative therapeutic potential of cannabinoid signaling in inflammation-accompanied diseases (e.g., multiple sclerosis or in organ transplantation, and to dissect the complex immunological effects of medical and “recreational” marijuana consumption. Thus, the objective of the current article is (i to summarize the most recent findings of the field; (ii to highlight the putative therapeutic potential of targeting cannabinoid signaling; (iii to identify open questions and key challenges; and (iv to suggest promising future directions for cannabinoid-based drug development.

  10. Targeting Slit-Roundabout signaling inhibits tumor angiogenesis in chemical-induced squamous cell carcinogenesis.

    Science.gov (United States)

    Wang, Li-Jing; Zhao, Yuan; Han, Bing; Ma, Yu-Guang; Zhang, Jie; Yang, Ding-Ming; Mao, Jian-Wen; Tang, Fu-Tian; Li, Wei-Dong; Yang, Yang; Wang, Rui; Geng, Jian-Guo

    2008-03-01

    Slit is a secreted protein known to function through the Roundabout (Robo) receptor as a repellent for axon guidance and neuronal migration, and as an inhibitor in leukocyte chemotaxis. We have previously shown that Slit2 is also secreted by a variety of human cancer cells whereby it acts as a chemoattractant to vascular endothelial cells for tumor angiogenesis. We used a blocking antibody to investigate the role of Slit-Robo signaling in tumor angiogenesis during oral carcinogenesis. In this report we undertook a multistage model of 7,12-dimethyl-1,2-benzanthracene-induced squamous cell carcinoma in the hamster buccal pouch. R5, a monoclonal antibody against the first immunoglobulin domain of Robo1, was used to study whether R5 blocks the Slit-Robo interaction and furthermore inhibits tumor angiogenesis and growth in our model. In addition, the expression of Slit2, von Willebrand factor, and vascular endothelial growth factor were examined using human tissue of oral cheek mucosa with oral squamous cell carcinoma. Our data showed that Slit2 was expressed minimally in normal and hyperplastic mucosa, moderately in dysplastic mucosa, and highly in neoplastic mucosa obtained from hamster buccal pouch. We also found that increased Slit2 expression was associated with higher tumor angiogenesis, as reflected by increased vascular endothelial growth factor expression and microvessel density. A similar Slit2 expression profile was found in human tissue. Importantly, interruption of the Slit2-Robo interaction using R5 inhibited tumor angiogenesis and growth in our in vivo model, which indicates that Slit2-mediated tumor angiogenesis is a critical process underlying the carcinogenesis of chemical-induced squamous cell carcinoma. Therefore, targeting Slit-Robo signaling may offer a novel antiangiogenesis approach for oral cancer therapy.

  11. Targeting GRB7/ERK/FOXM1 signaling pathway impairs aggressiveness of ovarian cancer cells.

    Directory of Open Access Journals (Sweden)

    David W Chan

    Full Text Available Ovarian cancer is a highly lethal disease with poor prognosis and especially in high-grade tumor. Emerging evidence has reported that aberrant upregulation and activation of GRB7, ERK as well as FOXM1 are closely associated with aggresivenesss of human cancers. However, the interplay between these factors in the pathogenesis of human cancers still remains unclear. In this study, we found that GRB7 (P<0.0001, ERK phosphorylation (P<0.0001 and FOXM1 (P = 0.001 were frequently increased and associated with high-grade tumors, as well as a high tendency in association with advanced stage ovarian cancer by immunohistochemical analysis. Intriguingly, the expressions of GRB7 (P<0.0001, ERK phosphorylation (P<0.001 and FOXM1 (P<0.001 showed a significant stepwise increase pattern along Grade 1 to Grade 3 ovarian cancers. Biochemical studies using western blot analysis demonstrated that enforced expression or knockdown of GRB7 showed GRB7 could elevate the levels of ERK phosphorylation and FOXM1, whereas enforced expression of FOXM1 could not alter levels of GRB7 and ERK phosphorylation. But inhibition of ERK signaling by U0126 or PD98059 could reduce the level of FOXM1 in GRB7-overexpressing ovarian cancer cells, suggesting that GRB7, ERK and FOXM1 are regulated orderly. Moreover, inhibition of ERK activity by U0126 or PD98059, or decreased FOXM1 expression by Thiostrepton significantly inhibited cell migration/invasion, tumor growth in vitro and in vivo. Collectively, our findings confer that targeting GRB7/ERK/FOXM1 signaling cascade may be a promising molecular therapeutic choice in combating ovarian cancer.

  12. Dovitinib Acts As a Novel Radiosensitizer in Hepatocellular Carcinoma by Targeting SHP-1/STAT3 Signaling

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Chao-Yuan [Department of Oncology, National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei, Taiwan (China); Department of Radiological Technology, Yuanpei University, Hsinchu, Taiwan (China); Tai, Wei-Tien [Department of Medical Research, National Taiwan University Hospital, Taipei, Taiwan (China); National Center of Excellence for Clinical Trial and Research, National Taiwan University Hospital, Taipei, Taiwan (China); Wu, Szu-Yuan [Institute of Toxicology, College of Medicine, National Taiwan University, Taipei, Taiwan (China); Department of Radiation Oncology, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan (China); Department of Internal Medicine, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Department of Biotechnology, Hungkuang University, Taichung, Taiwan (China); Shih, Chih-Ting; Chen, Min-Hsuan; Tsai, Ming-Hsien [Department of Medical Research, National Taiwan University Hospital, Taipei, Taiwan (China); Kuo, Chiung-Wen [Department of Medical Imaging and Radiological Technology, Yuanpei University of Medical Technology, Hsinchu, Taiwan (China); Shiau, Chung-Wai [Institute of Biopharmaceutical Sciences, National Yang-Ming University, Taipei, Taiwan (China); Hung, Man-Hsin, E-mail: cindybeaty@gmail.com [Division of Medical Oncology, Department of Oncology, Taipei Veterans General Hospital, Taipei, Taiwan (China); Division of Hematology and Oncology, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan (China); Program in Molecular Medicine, School of Life Science, National Yang-Ming University, Taipei, Taiwan (China); School of Medicine, National Yang-Ming University, Taipei, Taiwan (China); Chen, Kuen-Feng, E-mail: kfchen1970@ntu.edu.tw [Department of Medical Research, National Taiwan University Hospital, Taipei, Taiwan (China); National Center of Excellence for Clinical Trial and Research, National Taiwan University Hospital, Taipei, Taiwan (China)

    2016-06-01

    Purpose: Hepatocellular carcinoma (HCC) is among the most lethal human malignancies, and curative therapy is not an option for most patients. There is growing interest in the potential benefit of combining targeted therapies with radiation therapy (RT). This study aimed to characterize the efficacy and mechanism of an investigational drug, dovitinib, used in combination with RT. Methods and Materials: HCC cell lines (PLC5, Hep3B, SK-Hep1, HA59T, and Huh-7) were treated with dovitinib, RT, or both, and apoptosis and signal transduction were analyzed. Results: Dovitinib treatment resulted in Src homology region 2 (SH2) domain-containing phosphatase 1 (SHP-1)-mediated downregulation of p-STAT3 and promoted potent apoptosis of HCC cells. Ectopic expression of STAT3, or inhibition of SHP-1, diminished the effects of dovitinib on HCC cells. By ectopic expression and purified recombinant proteins of various mutant forms of SHP-1, the N-SH2 domain of SHP-1 was found to be required for dovitinib treatment. Overexpression of STAT3 or catalytic-dead mutant SHP-1 restored RT-induced reduction of HCC cell survival. Conversely, ectopic expression of SHP-1 or activation of SHP-1 by dovitinib enhanced the effects of RT against HCC in vitro and in vivo. Conclusions: SHP-1/STAT3 signaling is critically associated with the radiosensitivity of HCC cells. Combination therapy with RT and the SHP-1 agonist, such as dovitinib, resulted in enhanced in vitro and in vivo anti-HCC effects.

  13. Genetic polymorphisms regulating dopamine signaling in the frontal cortex interact to affect target detection under high working memory load.

    Science.gov (United States)

    Smith, Christopher T; Swift-Scanlan, Theresa; Boettiger, Charlotte A

    2014-02-01

    Frontal-dependent task performance is typically modulated by dopamine (DA) according to an inverted-U pattern, whereby intermediate levels of DA signaling optimizes performance. Numerous studies implicate trait differences in DA signaling based on differences in the catechol-O-methyltransferase (COMT) gene in executive function task performance. However, little work has investigated genetic variations in DA signaling downstream from COMT. One candidate is the DA- and cAMP-regulated phosphoprotein of molecular weight 32 kDa (DARPP-32), which mediates signaling through the D1-type DA receptor, the dominant DA receptor in the frontal cortex. Using an n-back task, we used signal detection theory to measure performance in a healthy adult population (n = 97) genotyped for single nucleotide polymorphisms in the COMT (rs4680) and DARPP-32 (rs907094) genes. Correct target detection (hits) and false alarms were used to calculate d' measures for each working memory load (0-, 2-, and 3-back). At the highest load (3-back) only, we observed a significant COMT × DARPP-32 interaction, such that the DARPP-32 T/T genotype enhanced target detection in COMT(ValVal) individuals, but impaired target detection in COMT(Met) carriers. These findings suggest that enhanced dopaminergic signaling via the DARPP-32 T allele aids target detection in individuals with presumed low frontal DA (COMT(ValVal)) but impairs target detection in those with putatively higher frontal DA levels (COMT(Met) carriers). Moreover, these data support an inverted-U model with intermediate levels of DA signaling optimizing performance on tasks requiring maintenance of mental representations in working memory.

  14. Targeting of RAGE-ligand signaling impairs breast cancer cell invasion and metastasis.

    Science.gov (United States)

    Kwak, T; Drews-Elger, K; Ergonul, A; Miller, P C; Braley, A; Hwang, G H; Zhao, D; Besser, A; Yamamoto, Y; Yamamoto, H; El-Ashry, D; Slingerland, J M; Lippman, M E; Hudson, B I

    2017-03-01

    The receptor for advanced glycation end products (RAGE) is highly expressed in various cancers and is correlated with poorer outcome in breast and other cancers. Here we tested the role of targeting RAGE by multiple approaches in the tumor and tumor microenvironment, to inhibit the metastatic process. We first tested how RAGE impacts tumor cell-intrinsic mechanisms using either RAGE overexpression or knockdown with short hairpin RNAs (shRNAs). RAGE ectopic overexpression in breast cancer cells increased MEK-EMT (MEK-epithelial-to-mesenchymal transition) signaling, transwell invasion and soft agar colony formation, and in vivo promoted lung metastasis independent of tumor growth. RAGE knockdown with multiple independent shRNAs in breast cancer cells led to decreased transwell invasion and soft agar colony formation, without affecting proliferation. In vivo, targeting RAGE shRNA knockdown in human and mouse breast cancer cells, decreased orthotopic tumor growth, reduced tumor angiogenesis and recruitment of inflammatory cells, and markedly decreased metastasis to the lung and liver in multiple xenograft and syngeneic mouse models. To test the non-tumor cell microenvironment role of RAGE, we performed syngeneic studies with orthotopically injected breast cancer cells in wild-type and RAGE-knockout C57BL6 mice. RAGE-knockout mice displayed striking impairment of tumor cell growth compared with wild-type mice, along with decreased mitogen-activated protein kinase signaling, tumor angiogenesis and inflammatory cell recruitment. To test the combined inhibition of RAGE in both tumor cell-intrinsic and non-tumor cells of the microenvironment, we performed in vivo treatment of xenografted tumors with FPS-ZM1 (1 mg/kg, two times per week). Compared with vehicle, FPS-ZM1 inhibited primary tumor growth, inhibited tumor angiogenesis and inflammatory cell recruitment and, most importantly, prevented metastasis to the lung and liver. These data demonstrate that RAGE drives tumor

  15. Targeting G-Protein Signaling for the Therapeutics of Prostate Tumor Bone Metastases and the Associated Chronic Bone Pain

    Science.gov (United States)

    2015-09-01

    Cancer Bone Metastasis, heterotrimeric G protein  subunits, G protein -coupled receptors, signal transduction 16. SECURITY CLASSIFICATION OF: 17...comparable to that of Gαt-expressing cells (Fig.2A), indicating the involvement of G subunits released from Gi/o proteins . Interestingly, the growth...1 AWARD NUMBER: W81XWH-12-1-0213 TITLE: Targeting G- Protein Signaling for the Therapeutics of Prostate Tumor Bone Metastases and the

  16. HES1, a target of Notch signaling, is elevated in canine osteosarcoma, but reduced in the most aggressive tumors

    OpenAIRE

    Dailey, Deanna D; Anfinsen, Kristin P.; Pfaff, Liza E; Ehrhart, EJ; Charles, J Brad; B?nsdorff, Tina B.; Thamm, Douglas H; Powers, Barbara E; Thora J Jonasdottir; Duval, Dawn L.

    2013-01-01

    Background Hairy and enhancer of split 1 (HES1), a basic helix-loop-helix transcriptional repressor, is a downstream target of Notch signaling. Notch signaling and HES1 expression have been linked to growth and survival in a variety of human cancer types and have been associated with increased metastasis and invasiveness in human osteosarcoma cell lines. Osteosarcoma (OSA) is an aggressive cancer demonstrating both high metastatic rate and chemotherapeutic resistance. The current study examin...

  17. Elimination of head and neck cancer initiating cells through targeting glucose regulated protein78 signaling

    Directory of Open Access Journals (Sweden)

    Huang Chih-Yang

    2010-10-01

    Full Text Available Abstract Background Head and neck squamous cell carcinoma (HNSCC is a highly lethal cancer that contains cellular and functional heterogeneity. Previously, we enriched a subpopulation of highly tumorigenic head and neck cancer initiating cells (HN-CICs from HNSCC. However, the molecular mechanisms by which to govern the characteristics of HN-CICs remain unclear. GRP78, a stress-inducible endoplasmic reticulum chaperone, has been reported to play a crucial role in the maintenance of embryonic stem cells, but the role of GRP78 in CICs has not been elucidated. Results Initially, we recognized GRP78 as a putative candidate on mediating the stemness and tumorigenic properties of HN-CICs by differential systemic analyses. Subsequently, cells with GRP78 anchored at the plasma membrane (memGRP78+ exerted cancer stemness properties of self-renewal, differentiation and radioresistance. Of note, xenotransplantation assay indicated merely 100 memGRP78+ HNSCCs resulted in tumor growth. Moreover, knockdown of GRP78 significantly reduced the self-renewal ability, side population cells and expression of stemness genes, but inversely promoted cell differentiation and apoptosis in HN-CICs. Targeting GRP78 also lessened tumorigenicity of HN-CICs both in vitro and in vivo. Clinically, co-expression of GRP78 and Nanog predicted the worse survival prognosis of HNSCC patients by immunohistochemical analyses. Finally, depletion of GRP78 in HN-CICs induced the expression of Bax, Caspase 3, and PTEN. Conclusions In summary, memGRP78 should be a novel surface marker for isolation of HN-CICs, and targeting GRP78 signaling might be a potential therapeutic strategy for HNSCC through eliminating HN-CICs.

  18. Keap1-Nrf2 Signaling: A Target for Cancer Prevention by Sulforaphane

    Science.gov (United States)

    Kensler, Thomas W; Egner, Patricia A; Agyeman, Abena S.; Visvanathan, Kala; Groopman, John D; Chen, Jian-Guo; Chen, Tao-Yang; Fahey, Jed W; Talalay, Paul

    2013-01-01

    Sulforaphane is a promising agent under preclinical evaluation in many models of disease prevention. This bioactive phytochemical affects many molecular targets in cellular and animal models; however, amongst the most sensitive is Keap1, a key sensor for the adaptive stress response system regulated through the transcription factor Nrf2. Keap1 is a sulfhydryl-rich protein that represses Nrf2 signaling by facilitating the poly ubiquitination of Nrf2 thereby enabling its subsequent proteasomal degradation. Interaction of sulforaphane with Keap1 disrupts this function and allows for nuclear accumulation of Nrf2 and activation of its transcriptional program. Enhanced transcription of Nrf2 target genes provokes a strong cytoprotective response that enhances resistance to carcinogenesis and other diseases mediated by exposures to electrophiles and oxidants. Clinical evaluation of sulforaphane has been largely conducted by utilizing preparations of broccoli or broccoli sprouts rich in either sulforaphane or its precursor form in plants, a stable β-thioglucose conjugate termed glucoraphanin. We have conducted a series of clinical trials in Qidong, China, a region where exposures to food- and air-borne carcinogens has been considerable, to evaluate the suitability of broccoli sprout beverages, rich in either glucoraphanin (GRR) or sulforaphane SFR or both for their bioavailability, tolerability and pharmacodynamic action in population-based interventions. Results from these clinical trials indicate that interventions with well characterized preparations of broccoli sprouts may enhance the detoxication of aflatoxins and air-borne toxins, which may in turn attenuate their associated health risks, including cancer, in exposed individuals. PMID:22752583

  19. Targeting Pulmonary Endothelial Hemoglobin α Improves Nitric Oxide Signaling and Reverses Pulmonary Artery Endothelial Dysfunction.

    Science.gov (United States)

    Alvarez, Roger A; Miller, Megan P; Hahn, Scott A; Galley, Joseph C; Bauer, Eileen; Bachman, Timothy; Hu, Jian; Sembrat, John; Goncharov, Dmitry; Mora, Ana L; Rojas, Mauricio; Goncharova, Elena; Straub, Adam C

    2017-12-01

    Pulmonary hypertension is characterized by pulmonary endothelial dysfunction. Previous work showed that systemic artery endothelial cells (ECs) express hemoglobin (Hb) α to control nitric oxide (NO) diffusion, but the role of this system in pulmonary circulation has not been evaluated. We hypothesized that up-regulation of Hb α in pulmonary ECs contributes to NO depletion and pulmonary vascular dysfunction in pulmonary hypertension. Primary distal pulmonary arterial vascular smooth muscle cells, lung tissue sections from unused donor (control) and idiopathic pulmonary artery (PA) hypertension lungs, and rat and mouse models of SU5416/hypoxia-induced pulmonary hypertension (PH) were used. Immunohistochemical, immunocytochemical, and immunoblot analyses and transfection, infection, DNA synthesis, apoptosis, migration, cell count, and protein activity assays were performed in this study. Cocultures of human pulmonary microvascular ECs and distal pulmonary arterial vascular smooth muscle cells, lung tissue from control and pulmonary hypertensive lungs, and a mouse model of chronic hypoxia-induced PH were used. Immunohistochemical, immunoblot analyses, spectrophotometry, and blood vessel myography experiments were performed in this study. We find increased expression of Hb α in pulmonary endothelium from humans and mice with PH compared with controls. In addition, we show up-regulation of Hb α in human pulmonary ECs cocultured with PA smooth muscle cells in hypoxia. We treated pulmonary ECs with a Hb α mimetic peptide that disrupts the association of Hb α with endothelial NO synthase, and found that cells treated with the peptide exhibited increased NO signaling compared with a scrambled peptide. Myography experiments using pulmonary arteries from hypoxic mice show that the Hb α mimetic peptide enhanced vasodilation in response to acetylcholine. Our findings reveal that endothelial Hb α functions as an endogenous scavenger of NO in the pulmonary endothelium

  20. BRAF gene alterations and enhanced mammalian target of rapamycin signaling in gangliogliomas.

    Science.gov (United States)

    Kakkar, Aanchal; Majumdar, Atreye; Pathak, Pankaj; Kumar, Anupam; Kumari, Kalpana; Tripathi, Manjari; Sharma, Mehar C; Suri, Vaishali; Tandon, Vivek; Chandra, Sarat P; Sarkar, Chitra

    2017-01-01

    Gangliogliomas (GGs) are slow-growing glioneuronal tumors seen in children and young adults. They are associated with intractable epilepsy, and have recently been found to harbor BRAF (B- rapidly accelerated fibrosarcoma) gene mutations. However, the mammalian target of rapamycin (mTOR) signaling pathway, downstream of BRAF, has not been evaluated extensively in GGs. GG cases were retrieved, clinical data obtained, and histopathological features reviewed. Sequencing for BRAF V600E mutation, analysis of BRAF copy number by quantitative real-time polymerase chain reaction, and immunohistochemistry for mTOR pathway markers p-S6 and p-4EBP1 were performed. Sixty-four cases of GG were identified (0.9% of central nervous system tumors). Of these, 28 had sufficient tumor tissue for further evaluation. Mixed glial and neuronal morphology was the commonest (64%) type. Focal cortical dysplasia was identified in the adjacent cortex (6 cases). BRAF V600E mutation was identified in 30% of GGs; BRAF copy number gain was observed in 50% of them. p-S6 and p-4EBP1 immunopositivity was seen in 57% cases each. Thus, mTOR pathway activation was seen in 81% cases, and was independent of BRAF alterations. 87% patients had Engel grade I outcome, while 13% had Engel grade II outcome. Both the Engel grade II cases analyzed showed BRAF V600E mutation. BRAF V600E mutation is frequent in GGs, as is BRAF gain; the former may serve as a target for personalized therapy in patients with residual tumors, necessitating its assessment in routine pathology reporting of these tumors. Evidence of mTOR pathway activation highlights similarities in the pathogenetic mechanisms underlying GG and focal cortical dysplasia, and suggests that mTOR inhibitors may be of utility in GG patients with persistent seizures after surgery.

  1. Targeted gene-silencing reveals the functional significance of myocardin signaling in the failing heart.

    Directory of Open Access Journals (Sweden)

    Mario Torrado

    Full Text Available BACKGROUND: Myocardin (MYOCD, a potent transcriptional coactivator of smooth muscle (SM and cardiac genes, is upregulated in failing myocardium in animal models and human end-stage heart failure (HF. However, the molecular and functional consequences of myocd upregulation in HF are still unclear. METHODOLOGY/PRINCIPAL FINDINGS: The goal of the present study was to investigate if targeted inhibition of upregulated expression of myocd could influence failing heart gene expression and function. To this end, we used the doxorubicin (Dox-induced diastolic HF (DHF model in neonatal piglets, in which, as we show, not only myocd but also myocd-dependent SM-marker genes are highly activated in failing left ventricular (LV myocardium. In this model, intra-myocardial delivery of short-hairpin RNAs, designed to target myocd variants expressed in porcine heart, leads on day 2 post-delivery to: (1 a decrease in the activated expression of myocd and myocd-dependent SM-marker genes in failing myocardium to levels seen in healthy control animals, (2 amelioration of impaired diastolic dysfunction, and (3 higher survival rates of DHF piglets. The posterior restoration of elevated myocd expression (on day 7 post-delivery led to overexpression of myocd-dependent SM-marker genes in failing LV-myocardium that was associated with a return to altered diastolic function. CONCLUSIONS/SIGNIFICANCE: These data provide the first evidence that a moderate inhibition (e.g., normalization of the activated MYOCD signaling in the diseased heart may be promising from a therapeutic point of view.

  2. Inhibition of Mammalian Target of Rapamycin Signaling with Rapamycin Prevents Trauma-Induced Heterotopic Ossification.

    Science.gov (United States)

    Qureshi, Ammar T; Dey, Devaveena; Sanders, Erin M; Seavey, Jonathan G; Tomasino, Allison M; Moss, Kaitlyn; Wheatley, Benjamin; Cholok, David; Loder, Shawn; Li, John; Levi, Benjamin; Davis, Thomas A

    2017-11-01

    A pressing clinical need exists for 63% to 65% of combat-wounded service members and 11% to 20% of civilians who develop heterotopic ossification (HO) after blast-related extremity injury and traumatic injuries, respectively. The mammalian target of rapamycin pathway is a central cellular sensor of injury. We evaluated the prophylactic effects of rapamycin, a selective inhibitor of mammalian target of rapamycin signaling, on HO formation in a rat model of blast-related, polytraumatic extremity injury. Rapamycin was administered intraperitoneally daily for 14 days at 0.5 mg/kg or 2.5 mg/kg. Ectopic bone formation was monitored by micro-computed tomography and confirmed by histologic examination. Connective tissue progenitor cells, platelet-derived growth factor receptor-α-positive cells, and α-smooth muscle actin-positive blood vessels were assayed at postoperative day 7 by colony formation and immunofluorescence. Early gene expression changes were determined by low-density microarray. There was significant attenuation of 1) total new bone and soft tissue ectopic bone with 0.5 mg/kg (38.5% and 14.7%) and 2.5 mg/kg rapamycin (90.3% and 82.9%), respectively, 2) connective tissue progenitor cells, 3) platelet-derived growth factor receptor-α-positive cells, 4) α-smooth muscle actin-positive blood vessels, and 5) of key extracellular matrix remodeling (CD44, Col1a1, integrins), osteogenesis (Sp7, Runx2, Bmp2), inflammation (Cxcl5, 10, IL6, Ccl2), and angiogenesis (Angpt2) genes. No wound healing complications were noted. Our data demonstrate the efficacy of rapamycin in inhibiting blast trauma-induced HO by a multipronged mechanism. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  3. Alcohol consumption mitigates apoptosis and mammalian target of rapamycin signaling in myocardium.

    Science.gov (United States)

    Elmadhun, Nassrene Y; Sabe, Ashraf A; Lassaletta, Antonio D; Sellke, Frank W

    2014-06-01

    Epidemiologic studies have shown that individuals who consume low to moderate alcohol have a lower risk of cardiovascular disease developing compared with abstainers. Although experimental studies confirmed this observation, the effect of alcohol on ischemic myocardium is still unclear. We developed a clinically relevant animal model of chronic myocardial ischemia to investigate the effects of moderate alcohol consumption on the myocardium. Fourteen Yorkshire swine underwent placement of an ameroid constrictor to induce chronic myocardial ischemia. Postoperatively, one group was supplemented with 90 mL 50% EtOH daily (n = 7) and one group was supplemented with 80 g sucrose daily to normalize caloric intake between groups (n = 7). After 7 weeks, all animals underwent sternotomy, and harvest of the chronically ischemic myocardium and nonischemic myocardium. Tissues were analyzed for protein expression and stained for apoptosis quantification. In the ischemic myocardium, alcohol down-regulated the following proapoptotic proteins: tumor necrosis factor-α, forkhead box protein 03, BCL2-associated death promoter, and cysteine aspartic acid-specific protease 9; up-regulated the following prosurvival proteins: 5'adenosine monophosphate-activated protein kinase, phosphorylated 5'adenosine monophosphate-activated protein kinase, and phosphorylated forkhead box protein 03; and down-regulated mammalian target of rapamycin (MTOR) signaling by down-regulating MTOR, phosphorylated MTOR, and up-regulating Deptor. In the nonischemic myocardium, alcohol up-regulated prosurvival proteins: protein kinase B, phosphorylated protein kinase B, phosphorylated B-cell CLL/lymphoma 2, 5'adenosine monophosphate-activated protein kinase, phosphorylated BCL2-associated death promoter, phosphorylated forkhead box protein 03, and down-regulated MTOR signaling by down-regulating phosphorylated MTOR and up-regulating Deptor. Alcohol also decreased cell death as measured by terminal

  4. The microRNA-132/212 family fine-tunes multiple targets in Angiotensin II signalling in cardiac fibroblasts

    DEFF Research Database (Denmark)

    Eskildsen, Tilde V; Schneider, Mikael; Sandberg, Maria B

    2015-01-01

    INTRODUCTION: MicroRNAs (miRNAs) are emerging as key regulators of cardiovascular development and disease; however, the cardiac miRNA target molecules are not well understood. We and others have described the Angiotensin II (AngII)-induced miR-132/212 family as novel regulators of cardiovascular......, signalling molecules and transcription factors. Subsequent comprehensive in silico analysis identified 24 target genes, of which 22 genes were qPCR validated. We identified seven genes involved in AngII signalling pathways. CONCLUSION: We here report novel insight of an extensive network of molecular...

  5. Signal and data processing of small targets 1989; Proceedings of the Meeting, Orlando, FL, Mar. 27-29, 1989

    Science.gov (United States)

    Drummond, Oliver E. (Editor)

    1989-01-01

    The present conference on digital signal processing, association and filtering techniques, and multiple-sensor/multiple-tracking techniques, discusses single-frame velocity estimation, efficient target extraction for laser radar imagery, precision target tracking for small extended objects, IR clutter partitioning for matched filter design, the maximum-likelihood approach to gamma circumvention, position estimation for optical point targets using staring detector arrays, and a multiple-scan signal processing technique for area-moving target indication. Also discussed are a proportional integral estimator, the prediction of track purity in tracking performance evaluations, synchronization and fault-tolerance in a distributed tracker, the benefits of soft sensors and probabilistic fusion, and testing track initiation algorithms fusing two-dimensional tracks.

  6. Distinct requirements for TrkB and TrkC signaling in target innervation by sensory neurons

    Science.gov (United States)

    Postigo, Antonio; Calella, Anna Maria; Fritzsch, Bernd; Knipper, Marlies; Katz, David; Eilers, Andreas; Schimmang, Thomas; Lewin, Gary R.; Klein, Rudiger; Minichiello, Liliana

    2002-01-01

    Signaling by brain-derived neurotrophic factor (BDNF) via the TrkB receptor, or by neurotrophin-3 (NT3) through the TrkC receptor support distinct populations of sensory neurons. The intracellular signaling pathways activated by Trk (tyrosine kinase) receptors, which in vivo promote neuronal survival and target innervation, are not well understood. Using mice with TrkB or TrkC receptors lacking the docking site for Shc adaptors (trkB(shc/shc) and trkC(shc/shc) mice), we show that TrkB and TrkC promote survival of sensory neurons mainly through Shc site-independent pathways, suggesting that these receptors use similar pathways to prevent apoptosis. In contrast, the regulation of target innervation appears different: in trkB(shc/shc) mice neurons lose target innervation, whereas in trkC(shc/shc) mice the surviving TrkC-dependent neurons maintain target innervation and function. Biochemical analysis indicates that phosphorylation at the Shc site positively regulates autophosphorylation of TrkB, but not of TrkC. Our findings show that although TrkB and TrkC signals mediating survival are largely similar, TrkB and TrkC signals required for maintenance of target innervation in vivo are regulated by distinct mechanisms.

  7. The malarial host-targeting signal is conserved in the Irish potato famine pathogen.

    Directory of Open Access Journals (Sweden)

    2006-05-01

    consistent with the idea that the context of their presentation is constrained. These studies provide the first evidence to our knowledge that eukaryotic microbes share equivalent pathogenic HT signals and thus conserved mechanisms to access host cells across plant and animal kingdoms that may present unique targets for prophylaxis across divergent pathogens.

  8. The malarial host-targeting signal is conserved in the Irish potato famine pathogen.

    Science.gov (United States)

    Bhattacharjee, Souvik; Hiller, N Luisa; Liolios, Konstantinos; Win, Joe; Kanneganti, Thirumala-Devi; Young, Carolyn; Kamoun, Sophien; Haldar, Kasturi

    2006-05-01

    that the context of their presentation is constrained. These studies provide the first evidence to our knowledge that eukaryotic microbes share equivalent pathogenic HT signals and thus conserved mechanisms to access host cells across plant and animal kingdoms that may present unique targets for prophylaxis across divergent pathogens.

  9. Hypothalamic Leptin and Ghrelin Signaling as Targets for Improvement in Metabolic Control.

    Science.gov (United States)

    Frago, Laura M; Chowen, Julie A

    2015-01-01

    Metabolic homeostasis requires a tight balance between energy intake and energy expenditure; hence, the physiological circuits implicated in the regulation of energy metabolism must be able to quickly adjust to changes in either side of the equation. Circulating orexigenic and anorexigenic factors, including ghrelin and leptin, are produced in the gastrointestinal tract and adipose tissue, respectively, in relation to an individual's nutritional status. These signals interact with central metabolic circuits to regulate the production and secretion of neuropeptides implicated in the control of appetite and energy expenditure. However, this physiological equilibrium can be perturbed by diverse processes, with weight gain occurring due to a positive energy balance and weight loss taking place if there is a negative energy balance. If a situation of positive energy balance continues for an extended period of time, excess weight is accumulated and this can eventually result in obesity. Obesity has become one of the most important health problems facing the industrialized world, indicating that metabolic equilibrium is frequently disrupted. Understanding how and why this occurs will allow new therapeutical targets to be identified.

  10. Surface modified fluorescent quantum dots with neurotransmitter ligands for potential targeting of cell signaling applications.

    Science.gov (United States)

    Mansur, Alexandra A P; Saliba, Juliana B; Mansur, Herman S

    2013-11-01

    The possibility of combining nanotechnology with nanomedicine opens a broad field of research which may truly revolutionize our society. The neural system plays a crucial role in the human body, and most related diseases can dramatically change the quality of life. Thus, the present study reports a novel approach for using neurotransmitters as ligands in the synthesis of surface-modified fluorescent nanocrystals for potential use in cell labeling applications. Briefly, CdS quantum dots (QDs) were prepared using L-glutamic and L-aspartic as surface capping agents via a one-step chemical processing method, which resulted in stable aqueous colloidal systems at room temperature and ambient pressure. UV-visible spectroscopy, photoluminescence spectroscopy (PL), Fourier transform infrared (FTIR) spectroscopy, and transmission electron microscopy (TEM) were used to characterize the synthesis and relative stability of peptide-capped CdS nanocrystals. The results demonstrate that both ligands were effective in nucleating and stabilizing CdS quantum dots in colloidal aqueous suspensions, with an estimated dimension below 3.3 nm and with fluorescence activity. Thus, novel nanohybrids were developed based on QDs bioconjugated to surface-active neurotransmitter moieties suitable for investigation as potential biomarkers in cell targeting and signaling applications. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. Target of rapamycin signaling orchestrates growth-defense trade-offs in plants.

    Science.gov (United States)

    De Vleesschauwer, David; Filipe, Osvaldo; Hoffman, Gena; Seifi, Hamed Soren; Haeck, Ashley; Canlas, Patrick; Van Bockhaven, Jonas; De Waele, Evelien; Demeestere, Kristof; Ronald, Pamela; Hofte, Monica

    2018-01-01

    Plant defense to microbial pathogens is often accompanied by significant growth inhibition. How plants merge immune system function with normal growth and development is still poorly understood. Here, we investigated the role of target of rapamycin (TOR), an evolutionary conserved serine/threonine kinase, in the plant defense response. We used rice as a model system and applied a combination of chemical, genetic, genomic and cell-based analyses. We demonstrate that ectopic expression of TOR and Raptor (regulatory-associated protein of mTOR), a protein previously demonstrated to interact with TOR in Arabidopsis, positively regulates growth and development in rice. Transcriptome analysis of rice cells treated with the TOR-specific inhibitor rapamycin revealed that TOR not only dictates transcriptional reprogramming of extensive gene sets involved in central and secondary metabolism, cell cycle and transcription, but also suppresses many defense-related genes. TOR overexpression lines displayed increased susceptibility to both bacterial and fungal pathogens, whereas plants with reduced TOR signaling displayed enhanced resistance. Finally, we found that TOR antagonizes the action of the classic defense hormones salicylic acid and jasmonic acid. Together, these results indicate that TOR acts as a molecular switch for the activation of cell proliferation and plant growth at the expense of cellular immunity. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  12. Dysregulation of Mammalian Target of Rapamycin Signaling in Mouse Models of Autism.

    Science.gov (United States)

    Huber, Kimberly M; Klann, Eric; Costa-Mattioli, Mauro; Zukin, R Suzanne

    2015-10-14

    The mammalian target of rapamycin (mTOR) is a central regulator of a diverse array of cellular processes, including cell growth, proliferation, autophagy, translation, and actin polymerization. Components of the mTOR cascade are present at synapses and influence synaptic plasticity and spine morphogenesis. A prevailing view is that the study of mTOR and its role in autism spectrum disorders (ASDs) will elucidate the molecular mechanisms by which mTOR regulates neuronal function under physiological and pathological conditions. Although many ASDs arise as a result of mutations in genes with multiple molecular functions, they appear to converge on common biological pathways that give rise to autism-relevant behaviors. Dysregulation of mTOR signaling has been identified as a phenotypic feature common to fragile X syndrome, tuberous sclerosis complex 1 and 2, neurofibromatosis 1, phosphatase and tensin homolog, and potentially Rett syndrome. Below are a summary of topics covered in a symposium that presents dysregulation of mTOR as a unifying theme in a subset of ASDs. Copyright © 2015 the authors 0270-6474/15/3513836-07$15.00/0.

  13. BRAF inhibitors suppress apoptosis through off-target inhibition of JNK signaling

    Science.gov (United States)

    Vin, Harina; Ojeda, Sandra S; Ching, Grace; Leung, Marco L; Chitsazzadeh, Vida; Dwyer, David W; Adelmann, Charles H; Restrepo, Monica; Richards, Kristen N; Stewart, Larissa R; Du, Lili; Ferguson, Scarlett B; Chakravarti, Deepavali; Ehrenreiter, Karin; Baccarini, Manuela; Ruggieri, Rosamaria; Curry, Jonathan L; Kim, Kevin B; Ciurea, Ana M; Duvic, Madeleine; Prieto, Victor G; Ullrich, Stephen E; Dalby, Kevin N; Flores, Elsa R; Tsai, Kenneth Y

    2013-01-01

    Vemurafenib and dabrafenib selectively inhibit the v-Raf murine sarcoma viral oncogene homolog B1 (BRAF) kinase, resulting in high response rates and increased survival in melanoma. Approximately 22% of individuals treated with vemurafenib develop cutaneous squamous cell carcinoma (cSCC) during therapy. The prevailing explanation for this is drug-induced paradoxical ERK activation, resulting in hyperproliferation. Here we show an unexpected and novel effect of vemurafenib/PLX4720 in suppressing apoptosis through the inhibition of multiple off-target kinases upstream of c-Jun N-terminal kinase (JNK), principally ZAK. JNK signaling is suppressed in multiple contexts, including in cSCC of vemurafenib-treated patients, as well as in mice. Expression of a mutant ZAK that cannot be inhibited reverses the suppression of JNK activation and apoptosis. Our results implicate suppression of JNK-dependent apoptosis as a significant, independent mechanism that cooperates with paradoxical ERK activation to induce cSCC, suggesting broad implications for understanding toxicities associated with BRAF inhibitors and for their use in combination therapies. DOI: http://dx.doi.org/10.7554/eLife.00969.001 PMID:24192036

  14. Mitogen activated protein kinase signaling in the kidney: Target for intervention?

    NARCIS (Netherlands)

    de Borst, M.H.; Wassef, L.; Kelly, D.J.; van Goor, H.; Navis, Ger Jan

    2006-01-01

    Mitogen activated protein kinases (MAPKs) are intracellular signal transduction molecules, which connect cell-surface receptor signals to intracellular processes. MAPKs regulate a range of cellular activities including cell proliferation, gene expression, apoptosis, cell differentiation and cytokine

  15. Phospholipid-mediated signaling systems as novel targets for treatment of heart disease

    National Research Council Canada - National Science Library

    Tappia, Paramjit S

    2007-01-01

    .... The identification of the changes in such signaling systems as well as understanding the contribution of phospholipid-signaling pathways to the pathophysiology of heart disease are rapidly emerging...

  16. Discovering Small Molecule Inhibitors Targeted to Ligand-Stimulated RAGE-DIAPH1 Signaling Transduction

    Science.gov (United States)

    Pan, Jinhong

    The receptor of advanced glycation end product (RAGE) is a multiligand receptor of the immunoglobulin superfamily of cell surface molecules, which plays an important role in immune responses. Full-length RAGE includes three extracellular immunoglobulin domains, a transmembrane domain and an intracellular domain. It is a pattern recognition receptor that can bind diverse ligands. NMR spectroscopy and x-ray crystallization studies of the extracellular domains of RAGE indicate that RAGE ligands bind by distinct charge- and hydrophobicity-dependent mechanisms. It is found that calgranulin binding to the C1C2 domain or AGEs binding to the V domain activates extracellular signaling, which triggers interactions of the RAGE cytoplasmic tail (ctRAGE) with intracellular effector, such as diaphanous 1 (DIAPH1), to initiate signal transduction cascades. ctRAGE is essential for RAGE-ligand-mediated signal transduction and consequent modulation of gene expression and cellular properties. RAGE is over-expressed in diseased tissues of most RAGE-associated pathogenic conditions, such as complications of Alzheimer's diseases, diabetes, vascular diseases, inflammation, cancers and neurodegeneration. They are the major diseases affecting a large population worldwide. RAGE can function as a biomarker or drug target for these diseases. The cytoplasmic tail of RAGE can be used as a drug target to inhibit RAGE-induced intracellular signaling by small molecule inhibitors to treat RAGE-associated diseases. We developed a high throughput screening assay with which we probed a small molecule library of 58,000 compounds to find that 777 small molecules displayed 50% inhibition and 97 compounds demonstrated dose-dependent inhibition of the binding of ctRAGE-DIAPH1. Eventually, there were 13 compounds which displayed dose-dependent inhibition of ctRAGE binding to DIAPH1 and direct binding to ctRAGE analyzed by 15N HSQC-NMR and native tryptophan fluorescence titration experiments; thus, they were

  17. Isoimperatorin ameliorates osteoarthritis by downregulating the mammalian target of rapamycin C1 signaling pathway.

    Science.gov (United States)

    Ouyang, Jiayao; Jiang, Huaji; Fang, Hang; Cui, Wenbo; Cai, Daozhang

    2017-12-01

    Osteoarthritis (OA) is the most common disease of the joints, and is characterized by the breakdown of cartilage and degradation of the extracellular matrix. OA causes a high level of patient suffering and incurs large societal costs; however, the current strategies for treating OA are restricted due to limited understanding of the underlying molecular and cellular mechanisms. In the present study, the beneficial effects of isoimperatorin (Iso) were investigated using an experimental mouse model of OA, and its mechanism of action on primary chondrocytes was elucidated. Destabilization of the medial meniscus was performed on 8‑week‑old male mice to induce OA in the knees. Iso (500 mg/g/day) was intragastrically administered for 4 weeks. Degeneration of articular cartilage was assessed by histology using the Osteoarthritis Research Society International scoring system. The expression of matrix metalloproteinase (MMP)13, Runt‑related transcription factor (Runx)2, type X collagen (Col X) and vascular endothelial growth factor (VEGF) in the knee joints was examined by immunohistochemistry. In vitro, murine primary chondrocytes were treated with various concentrations of Iso, followed by 10 ng/ml interleukin‑1. The mRNA expression levels of MMP13, Runx2, Col X and VEGF were determined by reverse transcription‑quantitative polymerase chain reaction. The levels of autophagy and mammalian target of rapamycin (mTOR) signaling were determined by western blotting. Iso significantly ameliorated the severity of articular cartilage degradation in mice with experimental OA. The expression levels of MMP13, Runx2, Col X and VEGF were reduced in Iso‑treated mice. In murine primary chondrocytes, Iso also reduced MMP13, Runx2, Col X and VEGF expression, and activated autophagy by downregulating the mTOR complex 1 (mTORC1) signaling pathway. Therefore, the results of the present study demonstrated that Iso ameliorates OA‑induced pathological alterations by delaying

  18. WNT1 inducible signaling pathway protein 3, WISP-3, a novel target gene in colorectal carcinomas with microsatellite instability.

    Science.gov (United States)

    Thorstensen, L; Diep, C B; Meling, G I; Aagesen, T H; Ahrens, C H; Rognum, T O; Lothe, R A

    2001-12-01

    Microsatellite instability (MSI) is the phenotype of colorectal carcinomas with defect mismatch repair. Genes with repetitive sequences within their coding regions are targets for mutations in these tumors. We have evaluated 2 novel candidate genes for potential involvement in development of MSI colorectal carcinomas and compared them with alterations in known target genes. The MSI status was determined by multiplex polymerase chain reactions (PCRs) of 5-17 markers in a Norwegian series of 275 colorectal carcinomas. All MSI tumors were analyzed for gene mutations using fluorescence PCR followed by capillary electrophoresis. Two novel candidate genes, WNT1-inducible signaling pathway protein 3 (WISP-3) and caspase-1, and 9 known target genes were analyzed. Thirteen percent of the tumors were MSI-high (H) and 12% were MSI-low (L). Thirty-three of 37 MSI-H vs. 1 of 34 MSI-L tumors showed mutations in the target genes (P WISP-3 was mutated in 31% of the MSI-H tumors. The frequencies of frameshift mutations in the known target genes were comparable with other studies. The relative high frequency of mutation, higher than those seen for other known target genes, the predicted truncation of the protein product, and the homology with WISP-1 and WISP-2, 2 proteins induced downstream of WNT1 signaling, strongly suggest WISP-3 as a novel target in development of MSI-H colorectal carcinomas.

  19. Genomic Targets and Features of BarA-UvrY (-SirA Signal Transduction Systems.

    Directory of Open Access Journals (Sweden)

    Tesfalem R Zere

    Full Text Available The two-component signal transduction system BarA-UvrY of Escherichia coli and its orthologs globally regulate metabolism, motility, biofilm formation, stress resistance, virulence of pathogens and quorum sensing by activating the transcription of genes for regulatory sRNAs, e.g. CsrB and CsrC in E. coli. These sRNAs act by sequestering the RNA binding protein CsrA (RsmA away from lower affinity mRNA targets. In this study, we used ChIP-exo to identify, at single nucleotide resolution, genomic sites for UvrY (SirA binding in E. coli and Salmonella enterica. The csrB and csrC genes were the strongest targets of crosslinking, which required UvrY phosphorylation by the BarA sensor kinase. Crosslinking occurred at two sites, an inverted repeat sequence far upstream of the promoter and a site near the -35 sequence. DNAse I footprinting revealed specific binding of UvrY in vitro only to the upstream site, indicative of additional binding requirements and/or indirect binding to the downstream site. Additional genes, including cspA, encoding the cold-shock RNA-binding protein CspA, showed weaker crosslinking and modest or negligible regulation by UvrY. We conclude that the global effects of UvrY/SirA on gene expression are primarily mediated by activating csrB and csrC transcription. We also used in vivo crosslinking and other experimental approaches to reveal new features of csrB/csrC regulation by the DeaD and SrmB RNA helicases, IHF, ppGpp and DksA. Finally, the phylogenetic distribution of BarA-UvrY was analyzed and found to be uniquely characteristic of γ-Proteobacteria and strongly anti-correlated with fliW, which encodes a protein that binds to CsrA and antagonizes its activity in Bacillus subtilis. We propose that BarA-UvrY and orthologous TCS transcribe sRNA antagonists of CsrA throughout the γ-Proteobacteria, but rarely or never perform this function in other species.

  20. The Role of Wnt Signaling in the Development of Alzheimer’s Disease: A Potential Therapeutic Target?

    Directory of Open Access Journals (Sweden)

    Wenbin Wan

    2014-01-01

    Full Text Available Accumulating evidence supports a key role for Wnt signaling in the development of the central nervous system (CNS during embryonic development and in the regulation of the structure and function of the adult brain. Alzheimer’s disease (AD is the most common form of senile dementia, which is characterized by β-amyloid (Aβ deposition in specific brain regions. However, the molecular mechanism underlying AD pathology remains elusive. Dysfunctional Wnt signaling is associated with several diseases such as epilepsy, cancer, metabolic disease, and AD. Increasing evidence suggests that downregulation of Wnt signaling, induced by Aβ, is associated with disease progression of AD. More importantly, persistent activation of Wnt signaling through Wnt ligands, or inhibition of negative regulators of Wnt signaling, such as Dickkopf-1 (DKK-1 and glycogen synthase kinase-3β (GSK-3β that are hyperactive in the disease state, is able to protect against Aβ toxicity and ameliorate cognitive performance in AD. Together, these data suggest that Wnt signaling might be a potential therapeutic target of AD. Here, we review recent studies related to the progression of AD where Wnt signaling might be relevant and participate in the development of the disease. Then, we focus on the potential relevance of manipulating the Wnt signaling pathway for the treatment of AD.

  1. Lubiprostone targets prostanoid signaling and promotes ion transporter trafficking, mucus exocytosis, and contractility.

    Science.gov (United States)

    Jakab, Robert L; Collaco, Anne M; Ameen, Nadia A

    2012-11-01

    Lubiprostone is a chloride channel activator in clinical use for the treatment of chronic constipation, but the mechanisms of action of the drug are poorly understood. The aim of this study was to determine whether lubiprostone exerts secretory effects in the intestine by membrane trafficking of ion transporters and associated machinery. Immunolabeling and quantitative fluorescence intensity were used to examine lubiprostone-induced trafficking of the cystic fibrosis transmembrane conductance regulator (CFTR), sodium/potassium-coupled chloride co-transporter 1 (NKCC1), electrogenic sodium/bicarbonate co-transporter 1 (NBCe1), down-regulated in adenoma (DRA), putative anion transporter 1 (PAT1), sodium/proton exchanger 3 (NHE3), Ca(2+) activated chloride channel 2 (ClC-2) serotonin and its transporter SERT, E prostanoid receptors EP4 and EP1, sodium/potassium ATPase (Na-K-ATPase) and protein kinase A (PKA). The effects of lubiprostone on mucus exocytosis in rat intestine and human rectosigmoid explants were also examined. Lubiprostone induced contraction of villi and proximal colonic plicae and membrane trafficking of transporters that was more pronounced in villus/surface cells compared to the crypt. Membrane trafficking was determined by: (1) increased membrane labeling for CFTR, PAT1, NKCC1, and NBCe1 and decreased membrane labeling for NHE3, DRA and ClC-2; (2) increased serotonin, SERT, EP4, EP1 and PKA labeling in enterochromaffin cells; (3) increased SERT, EP4, EP1, PKA and Na-K-ATPase in enterocytes; and (4) increased mucus exocytosis in goblet cells. These data suggest that lubiprostone can target serotonergic, EP4/PKA and EP1 signaling in surface/villus regions; stimulate membrane trafficking of CFTR/NBCe1/NKCC1 in villus epithelia and PAT1/NBCe1/NKCC1 in colonic surface epithelia; suppress NHE3/DRA trafficking and fluid absorption; and enhance mucus-mobilization and mucosal contractility.

  2. Targeting renal purinergic signalling for the treatment of lithium-induced nephrogenic diabetes insipidus.

    Science.gov (United States)

    Kishore, B K; Carlson, N G; Ecelbarger, C M; Kohan, D E; Müller, C E; Nelson, R D; Peti-Peterdi, J; Zhang, Y

    2015-06-01

    Lithium still retains its critical position in the treatment of bipolar disorder by virtue of its ability to prevent suicidal tendencies. However, chronic use of lithium is often limited by the development of nephrogenic diabetes insipidus (NDI), a debilitating condition. Lithium-induced NDI is due to resistance of the kidney to arginine vasopressin (AVP), leading to polyuria, natriuresis and kaliuresis. Purinergic signalling mediated by extracellular nucleotides (ATP/UTP), acting via P2Y receptors, opposes the action of AVP on renal collecting duct (CD) by decreasing the cellular cAMP and thus AQP2 protein levels. Taking a cue from this phenomenon, we discovered the potential involvement of ATP/UTP-activated P2Y2 receptor in lithium-induced NDI in rats and showed that P2Y2 receptor knockout mice are significantly resistant to Li-induced polyuria, natriuresis and kaliuresis. Extension of these studies revealed that ADP-activated P2Y12 receptor is expressed in the kidney, and its irreversible blockade by the administration of clopidogrel bisulphate (Plavix(®)) ameliorates Li-induced NDI in rodents. Parallel in vitro studies showed that P2Y12 receptor blockade by the reversible antagonist PSB-0739 sensitizes CD to the action of AVP. Thus, our studies unravelled the potential beneficial effects of targeting P2Y2 or P2Y12 receptors to counter AVP resistance in lithium-induced NDI. If established in further studies, our findings may pave the way for the development of better and safer methods for the treatment of NDI by bringing a paradigm shift in the approach from the current therapies that predominantly counter the anti-AVP effects to those that enhance the sensitivity of the kidney to AVP action. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.

  3. DOA Estimation of Low Altitude Target Based on Adaptive Step Glowworm Swarm Optimization-multiple Signal Classification Algorithm

    Directory of Open Access Journals (Sweden)

    Zhou Hao

    2015-06-01

    Full Text Available The traditional MUltiple SIgnal Classification (MUSIC algorithm requires significant computational effort and can not be employed for the Direction Of Arrival (DOA estimation of targets in a low-altitude multipath environment. As such, a novel MUSIC approach is proposed on the basis of the algorithm of Adaptive Step Glowworm Swarm Optimization (ASGSO. The virtual spatial smoothing of the matrix formed by each snapshot is used to realize the decorrelation of the multipath signal and the establishment of a fullorder correlation matrix. ASGSO optimizes the function and estimates the elevation of the target. The simulation results suggest that the proposed method can overcome the low altitude multipath effect and estimate the DOA of target readily and precisely without radar effective aperture loss.

  4. The microRNA-132/212 family fine-tunes multiple targets in Angiotensin II signalling in cardiac fibroblasts

    DEFF Research Database (Denmark)

    Eskildsen, Tilde V; Schneider, Mikael; Sandberg, Maria B

    2015-01-01

    function including regulation of cardiac hypertrophy, heart failure and blood pressure possibly through AT1R signalling. However, the miR-132/212 targets in the heart remain unknown. MATERIALS AND METHODS: To understand the role of these miRNAs in cardiac signalling networks, we undertook comprehensive......INTRODUCTION: MicroRNAs (miRNAs) are emerging as key regulators of cardiovascular development and disease; however, the cardiac miRNA target molecules are not well understood. We and others have described the Angiotensin II (AngII)-induced miR-132/212 family as novel regulators of cardiovascular...... in silico and in vitro experiments to identify miR-132/212 molecular targets in primary rat cardiac fibroblasts. RESULTS: MiR-132/212 overexpression increased fibroblast cell size and mRNA arrays detected several hundred genes that were differentially expressed, including a wide panel of receptors...

  5. Estimating the delay-Doppler of target echo in a high clutter underwater environment using wideband linear chirp signals: Evaluation of performance with experimental data.

    Science.gov (United States)

    Yu, Ge; Yang, T C; Piao, Shengchun

    2017-10-01

    A chirp signal is a signal with linearly varying instantaneous frequency over the signal bandwidth, also known as a linear frequency modulated (LFM) signal. It is widely used in communication, radar, active sonar, and other applications due to its Doppler tolerance property in signal detection using the matched filter (MF) processing. Modern sonar uses high-gain, wideband signals to improve the signal to reverberation ratio. High gain implies a high product of the signal bandwidth and duration. However, wideband and/or long duration LFM signals are no longer Doppler tolerant. The shortcoming of the standard MF processing is loss of performance, and bias in range estimation. This paper uses the wideband ambiguity function and the fractional Fourier transform method to estimate the target velocity and restore the performance. Target velocity or Doppler provides a clue for differentiating the target from the background reverberation and clutter. The methods are applied to simulated and experimental data.

  6. A model system for targeted drug release triggered by biomolecular signals logically processed through enzyme logic networks.

    Science.gov (United States)

    Mailloux, Shay; Halámek, Jan; Katz, Evgeny

    2014-03-07

    A new Sense-and-Act system was realized by the integration of a biocomputing system, performing analytical processes, with a signal-responsive electrode. A drug-mimicking release process was triggered by biomolecular signals processed by different logic networks, including three concatenated AND logic gates or a 3-input OR logic gate. Biocatalytically produced NADH, controlled by various combinations of input signals, was used to activate the electrochemical system. A biocatalytic electrode associated with signal-processing "biocomputing" systems was electrically connected to another electrode coated with a polymer film, which was dissolved upon the formation of negative potential releasing entrapped drug-mimicking species, an enzyme-antibody conjugate, operating as a model for targeted immune-delivery and consequent "prodrug" activation. The system offers great versatility for future applications in controlled drug release and personalized medicine.

  7. Signaling by 4-hydroxy-2-nonenal: Exposure protocols, target selectivity and degradation.

    Science.gov (United States)

    Zhang, Hongqiao; Forman, Henry Jay

    2017-03-01

    4-hydroxy-2-nonenal (HNE), a major non-saturated aldehyde product of lipid peroxidation, has been extensively studied as a signaling messenger. In these studies a wide range of HNE concentrations have been used, ranging from the unstressed plasma concentration to far beyond what would be found in actual pathophysiological condition. In addition, accumulating evidence suggest that signaling protein modification by HNE is specific with only those proteins with cysteine, histidine, and lysine residues located in certain sequence or environments adducted by HNE. HNE-signaling is further regulated through the turnover of HNE-signaling protein adducts through proteolytic process that involve proteasomes, lysosomes and autophagy. This review discusses the HNE concentrations and exposure modes used in signaling studies, the selectivity of the HNE-adduction site, and the turnover of signaling protein adducts. Copyright © 2016 Elsevier Inc. All rights reserved.

  8. Future Perspectives: Therapeutic Targeting of Notch Signalling May Become a Strategy in Patients Receiving Stem Cell Transplantation for Hematologic Malignancies

    Directory of Open Access Journals (Sweden)

    Elisabeth Ersvaer

    2011-01-01

    Full Text Available The human Notch system consists of 5 ligands and 4 membrane receptors with promiscuous ligand binding, and Notch-initiated signalling interacts with a wide range of other intracellular pathways. The receptor signalling seems important for regulation of normal and malignant hematopoiesis, development of the cellular immune system, and regulation of immune responses. Several Notch-targeting agents are now being developed, including natural receptor ligands, agonistic and antagonistic antibodies, and inhibitors of intracellular Notch-initiated signalling. Some of these agents are in clinical trials, and several therapeutic strategies seem possible in stem cell recipients: (i agonists may be used for stem cell expansion and possibly to enhance posttransplant lymphoid reconstitution; (ii receptor-specific agonists or antagonists can be used for immunomodulation; (iii Notch targeting may have direct anticancer effects. Although the effects of therapeutic targeting are difficult to predict due to promiscuous ligand binding, targeting of this system may represent an opportunity to achieve combined effects with earlier posttransplant reconstitution, immunomodulation, or direct anticancer effects.

  9. Green tea polyphenol, (-)-epigallocatechin-3-gallate, induces toxicity in human skin cancer cells by targeting β-catenin signaling.

    Science.gov (United States)

    Singh, Tripti; Katiyar, Santosh K

    2013-12-01

    The green tea polyphenol, (-)-epigallocatechin-3-gallate (EGCG), has been shown to have anti-carcinogenic effects in several skin tumor models, and efforts are continued to investigate the molecular targets responsible for its cytotoxic effects to cancer cells. Our recent observation that β-catenin is upregulated in skin tumors suggested the possibility that the anti-skin carcinogenic effects of EGCG are mediated, at least in part, through its effects on β-catenin signaling. We have found that treatment of the A431 and SCC13 human skin cancer cell lines with EGCG resulted in reduced cell viability and increased cell death and that these cytotoxic effects were associated with inactivation of β-catenin signaling. Evidence of EGCG-induced inactivation of β-catenin included: (i) reduced accumulation of nuclear β-catenin; (ii) enhanced levels of casein kinase1α, reduced phosphorylation of glycogen synthase kinase-3β, and increased phosphorylation of β-catenin on critical serine(45,33/37) residues; and (iii) reduced levels of matrix metalloproteinase (MMP)-2 and MMP-9, which are down-stream targets of β-catenin. Treatment of cells with prostaglandin E2 (PGE2) enhanced the accumulation of β-catenin and enhanced β-catenin signaling. Treatment with either EGCG or an EP2 antagonist (AH6809) reduced the PGE2-enhanced levels of cAMP, an upstream regulator of β-catenin. Inactivation of β-catenin by EGCG resulted in suppression of cell survival signaling proteins. siRNA knockdown of β-catenin in A431 and SCC13 cells reduced cell viability. Collectively, these data suggest that induction of cytotoxicity in skin cancer cells by EGCG is mediated by targeting of β-catenin signaling and that the β-catenin signaling is upregulated by inflammatory mediators. © 2013.

  10. Subnuclear localization, rates and effectiveness of UVC-induced unscheduled DNA synthesis visualized by fluorescence widefield, confocal and super-resolution microscopy.

    Science.gov (United States)

    Pierzyńska-Mach, Agnieszka; Szczurek, Aleksander; Cella Zanacchi, Francesca; Pennacchietti, Francesca; Drukała, Justyna; Diaspro, Alberto; Cremer, Christoph; Darzynkiewicz, Zbigniew; Dobrucki, Jurek W

    2016-01-01

    Unscheduled DNA synthesis (UDS) is the final stage of the process of repair of DNA lesions induced by UVC. We detected UDS using a DNA precursor, 5-ethynyl-2'-deoxyuridine (EdU). Using wide-field, confocal and super-resolution fluorescence microscopy and normal human fibroblasts, derived from healthy subjects, we demonstrate that the sub-nuclear pattern of UDS detected via incorporation of EdU is different from that when BrdU is used as DNA precursor. EdU incorporation occurs evenly throughout chromatin, as opposed to just a few small and large repair foci detected by BrdU. We attribute this difference to the fact that BrdU antibody is of much larger size than EdU, and its accessibility to the incorporated precursor requires the presence of denatured sections of DNA. It appears that under the standard conditions of immunocytochemical detection of BrdU only fragments of DNA of various length are being denatured. We argue that, compared with BrdU, the UDS pattern visualized by EdU constitutes a more faithful representation of sub-nuclear distribution of the final stage of nucleotide excision repair induced by UVC. Using the optimized integrated EdU detection procedure we also measured the relative amount of the DNA precursor incorporated by cells during UDS following exposure to various doses of UVC. Also described is the high degree of heterogeneity in terms of the UVC-induced EdU incorporation per cell, presumably reflecting various DNA repair efficiencies or differences in the level of endogenous dT competing with EdU within a population of normal human fibroblasts.

  11. WNT signalling events near the cell membrane and their pharmacological targeting for the treatment of cancer

    NARCIS (Netherlands)

    Driehuis, Else; Clevers, Hans

    2017-01-01

    WNT signalling is an essential signalling pathway for all multicellular animals. Although first described more than 30 years ago, new components and regulators of the pathway are still being discovered. Considering its importance in both embryonic development and adult homeostasis, it is not

  12. The core spliceosome as target and effector of non-canonical ATM signalling

    NARCIS (Netherlands)

    Tresini, Maria; Warmerdam, Daniël O; Kolovos, Petros; Snijder, Loes; Vrouwe, Mischa G; Demmers, Jeroen A A; van IJcken, Wilfred F J; Grosveld, Frank G; Medema, René H; Hoeijmakers, Jan H J; Mullenders, Leon H F; Vermeulen, Wim; Marteijn, Jurgen A

    2015-01-01

    In response to DNA damage, tissue homoeostasis is ensured by protein networks promoting DNA repair, cell cycle arrest or apoptosis. DNA damage response signalling pathways coordinate these processes, partly by propagating gene-expression-modulating signals. DNA damage influences not only the

  13. Enzalutamide: targeting the androgen signalling pathway in metastatic castration-resistant prostate cancer

    NARCIS (Netherlands)

    Schalken, J.A.; Fitzpatrick, J.M.

    2016-01-01

    Significant progress has been made in the understanding of the underlying cancer biology of castration-resistant prostate cancer (CRPC) with the androgen receptor (AR) signalling pathway remaining implicated throughout the prostate cancer disease continuum. Reactivation of the AR signalling pathway

  14. Genome-wide identification of Bcl11b gene targets reveals role in brain-derived neurotrophic factor signaling.

    Directory of Open Access Journals (Sweden)

    Bin Tang

    Full Text Available B-cell leukemia/lymphoma 11B (Bcl11b is a transcription factor showing predominant expression in the striatum. To date, there are no known gene targets of Bcl11b in the nervous system. Here, we define targets for Bcl11b in striatal cells by performing chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq in combination with genome-wide expression profiling. Transcriptome-wide analysis revealed that 694 genes were significantly altered in striatal cells over-expressing Bcl11b, including genes showing striatal-enriched expression similar to Bcl11b. ChIP-seq analysis demonstrated that Bcl11b bound a mixture of coding and non-coding sequences that were within 10 kb of the transcription start site of an annotated gene. Integrating all ChIP-seq hits with the microarray expression data, 248 direct targets of Bcl11b were identified. Functional analysis on the integrated gene target list identified several zinc-finger encoding genes as Bcl11b targets, and further revealed a significant association of Bcl11b to brain-derived neurotrophic factor/neurotrophin signaling. Analysis of ChIP-seq binding regions revealed significant consensus DNA binding motifs for Bcl11b. These data implicate Bcl11b as a novel regulator of the BDNF signaling pathway, which is disrupted in many neurological disorders. Specific targeting of the Bcl11b-DNA interaction could represent a novel therapeutic approach to lowering BDNF signaling specifically in striatal cells.

  15. Dendritic targeting in the leg neuropil of Drosophila: the role of midline signalling molecules in generating a myotopic map.

    Directory of Open Access Journals (Sweden)

    David J Brierley

    2009-09-01

    Full Text Available Neural maps are emergent, highly ordered structures that are essential for organizing and presenting synaptic information. Within the embryonic nervous system of Drosophila motoneuron dendrites are organized topographically as a myotopic map that reflects their pattern of innervation in the muscle field. Here we reveal that this fundamental organizational principle exists in adult Drosophila, where the dendrites of leg motoneurons also generate a myotopic map. A single postembryonic neuroblast sequentially generates different leg motoneuron subtypes, starting with those innervating proximal targets and medial neuropil regions and producing progeny that innervate distal muscle targets and lateral neuropil later in the lineage. Thus the cellular distinctions in peripheral targets and central dendritic domains, which make up the myotopic map, are linked to the birth-order of these motoneurons. Our developmental analysis of dendrite growth reveals that this myotopic map is generated by targeting. We demonstrate that the medio-lateral positioning of motoneuron dendrites in the leg neuropil is controlled by the midline signalling systems Slit-Robo and Netrin-Fra. These results reveal that dendritic targeting plays a major role in the formation of myotopic maps and suggests that the coordinate spatial control of both pre- and postsynaptic elements by global neuropilar signals may be an important mechanism for establishing the specificity of synaptic connections.

  16. [Piperine regulates glucose metabolism disorder in HepG2 cells of insulin resistance models via targeting upstream target of AMPK signaling pathway].

    Science.gov (United States)

    Wan, Chun-Ping; Wei, Ya-Gai; Li, Xiao-Xue; Zhang, Li-Jun; Yang, Rui; Bao, Zhao-Ri-Ge-Tu

    2017-02-01

    To investigate the effect of piperine on the disorder of glucose metabolism in the cell model with insulin resistance (IR) and explore the molecules mechanism on intervening the upstream target of AMPK signaling pathway. The insulin resistance models in HepG2 cells were established by fat emulsion stimulation. Then glucose consumption in culture supernatant was detected by GOD-POD method. Enzyme-linked immunosorbent assay(ELISA) was used to measure the levels of leptin(LEP) and adiponectin(APN) in culture supernatant; Real-time quantitative PCR was used to assess the mRNA expression of APN and LEP; and the protein expression levels of LepR, AdipoR1, AdipoR2 and the activation of AMPK signaling pathway were detected by Western blot analysis. The results showed that piperine, rosiglitazone and AMPK agonist AICAR could significantly elevate the glucose consumption in insulin resistance cell models, enhance the level of APN, promote APN mRNA transcripts and increase the protein expression of Adipo receptor. Meanwhile,AMPKα mRNA and р-AMPKα protein expressions were also increased in piperine treated cells, but both LEP mRNA expression and LepR protein expressions were decreased in piperine treated group. The results indicated that piperine could significantly ameliorate the glucose metabolism disorder in insulin resistance cell models through regulating upstream molecules (APN and LEP) of AMPK signaling pathway, and thus activate the AMPK signaling pathway. Copyright© by the Chinese Pharmaceutical Association.

  17. Activin signaling targeted by insulin/dFOXO regulates aging and muscle proteostasis in Drosophila.

    Directory of Open Access Journals (Sweden)

    Hua Bai

    2013-11-01

    Full Text Available Reduced insulin/IGF signaling increases lifespan in many animals. To understand how insulin/IGF mediates lifespan in Drosophila, we performed chromatin immunoprecipitation-sequencing analysis with the insulin/IGF regulated transcription factor dFOXO in long-lived insulin/IGF signaling genotypes. Dawdle, an Activin ligand, is bound and repressed by dFOXO when reduced insulin/IGF extends lifespan. Reduced Activin signaling improves performance and protein homeostasis in muscles of aged flies. Activin signaling through the Smad binding element inhibits the transcription of Autophagy-specific gene 8a (Atg8a within muscle, a factor controlling the rate of autophagy. Expression of Atg8a within muscle is sufficient to increase lifespan. These data reveal how insulin signaling can regulate aging through control of Activin signaling that in turn controls autophagy, representing a potentially conserved molecular basis for longevity assurance. While reduced Activin within muscle autonomously retards functional aging of this tissue, these effects in muscle also reduce secretion of insulin-like peptides at a distance from the brain. Reduced insulin secretion from the brain may subsequently reinforce longevity assurance through decreased systemic insulin/IGF signaling.

  18. Human nonsense-mediated RNA decay regulates EMT by targeting the TGF-ß signaling pathway in lung adenocarcinoma.

    Science.gov (United States)

    Cao, Lu; Qi, Lisha; Zhang, Lin; Song, Wangzhao; Yu, Yue; Xu, Cong; Li, Lingmei; Guo, Yuhong; Yang, Lingyi; Liu, Changxu; Huang, Qiujuan; Wang, Yalei; Sun, Baocun; Meng, Bin; Zhang, Bin; Cao, Wenfeng

    2017-09-10

    Nonsense-mediated mRNA decay (NMD) is a highly conserved pathway that selectively degrades aberrant RNA transcripts. In this study, we proved that NMD regulates the epithelial-mesenchymal transition (EMT) of lung adenocarcinoma (ADC). Moreover, we found that NMD core factor UP-frameshift 1 tends to be expressed at lower levels in human ADC tissues than in normal lung tissues, thereby raising the possibility that NMD may be downregulated to permit ADC oncogenesis. Our experiments in human ADC cell lines showed that downregulating NMD can promote EMT. Moreover, EMT can be inhibited by upregulating NMD. We tested the role of TGF-ß signaling and found that NMD influences EMT by targeting the TGF-ß signaling pathway. Our findings reveal that NMD is a potential tumor regulatory mechanism and may be a potential therapeutic target for ADC. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. miR-958 inhibits Toll signaling and Drosomycin expression via direct targeting of Toll and Dif in Drosophila melanogaster.

    Science.gov (United States)

    Li, Shengjie; Li, Yao; Shen, Li; Jin, Ping; Chen, Liming; Ma, Fei

    2017-02-01

    Drosophila melanogaster is widely used as a model system to study innate immunity and signaling pathways related to innate immunity, including the Toll signaling pathway. Although this pathway is well studied, the precise mechanisms of posttranscriptional regulation of key components of the Toll signaling pathway by microRNAs (miRNAs) remain obscure. In this study, we used an in silico strategy in combination with the Gal80ts-Gal4 driver system to identify microRNA-958 (miR-958) as a candidate Toll pathway regulating miRNA in Drosophila We report that overexpression of miR-958 significantly reduces the expression of Drosomycin, a key antimicrobial peptide involved in Toll signaling and the innate immune response. We further demonstrate in vitro and in vivo that miR-958 targets the Toll and Dif genes, key components of the Toll signaling pathway, to negatively regulate Drosomycin expression. In addition, a miR-958 sponge rescued the expression of Toll and Dif, resulting in increased expression of Drosomycin. These results, not only revealed a novel function and modulation pattern of miR-958, but also provided a new insight into the underlying molecular mechanisms of Toll signaling in regulation of innate immunity. Copyright © 2017 the American Physiological Society.

  20. Indirect targeting of IGF receptor signaling in vivo by substrate-selective inhibition of PAPP-A proteolytic activity

    Science.gov (United States)

    Kalra, Bhanu; Savjani, Gopal; Kumar, Ajay; Conover, Cheryl A.; Oxvig, Claus

    2014-01-01

    The insulin-like growth factor (IGF) signaling pathway is involved in certain human cancers, and the feasibility of directly targeting the IGF receptor has been actively investigated. However, recent evidence from clinical trials suggests that this approach can be problematic. We have developed an alternative strategy to indirectly inhibit the IGF signaling by targeting the metalloproteinase, pregnancy-associated plasma protein-A (PAPP-A). PAPP-A associated with the cell surface cleaves IGF binding protein-4 (IGFBP-4), when IGF is bound to IGFBP-4, and thereby increases IGF bioavailability for receptor activation in an autocrine/paracrine manner. We hypothesized that inhibition of PAPP-A would suppress excessive local IGF signaling in tissues where this is caused by increased PAPP-A proteolytic activity. To test this hypothesis, we developed an inhibitory monoclonal antibody, mAb 1/41, which targets a unique substrate-binding exosite of PAPP-A. This inhibitor selectively and specifically inhibits proteolytic cleavage of IGFBP-4 with an inhibitory constant (Ki) of 135 pM. In addition, it inhibited intracellular signaling of the IGF receptor (AKT phosphorylation) in monolayers of A549 cells, an IGF-responsive lung cancer-derived cell line found to express high levels of PAPP-A. We further showed that mAb 1/41 is effective towards PAPP-A bound to cell surfaces, and that it is capable of inhibiting PAPP-A activity in vivo. Using a murine xenograft model of A549 cells, we demonstrated that mAb 1/41 administered intraperitoneally significantly inhibited tumor growth. Analysis of xenograft tumor tissue recovered from treated mice showed penetration of mAb 1/41, reduced IGFBP-4 proteolysis, and reduced AKT phosphorylation. Our study provides proof of concept that IGF signaling can be selectively reduced by targeting a regulatory proteinase that functions extracellularly, upstream of the IGF receptor. PAPP-A targeting thus represents an alternative therapeutic strategy for

  1. Attentional Changes during Implicit Learning: Signal Validity Protects a Target Stimulus from the Attentional Blink

    Science.gov (United States)

    Livesey, Evan J.; Harris, Irina M.; Harris, Justin A.

    2009-01-01

    Participants in 2 experiments performed 2 simultaneous tasks: one, a dual-target detection task within a rapid sequence of target and distractor letters; the other, a cued reaction time task requiring participants to make a cued left-right response immediately after each letter sequence. Under these rapid visual presentation conditions, it is…

  2. A NUFFT Based Step-frequency Chirp Signal High Resolution Imaging Algorithm and Target Recognition Algorithm

    Directory of Open Access Journals (Sweden)

    Xiang Yin

    2015-12-01

    Full Text Available Radar Automatic Target Recognition (RATR is the key technique to be breaked through in the fuure development of intelligent weapon system. Compared to the 2-D SAR image target recognition, High Resolution Range Profile (HRRP target recognition has the advantage of low data dimension, low requirement of radar system's calculation and storage ability, and the imaging algorithm is also not complicated. HRRP imaging is the first and the key process in target recognition, its speed and imaging quality can directly influence the real-time capability and accuracy of target recognition. In this paper a new HRRP imaging algorithm — NUFFT algorithm is proposed, the derivation of mathematical expression is given, both for the echo simulation process and the imaging process. In the meantime, by analyzing each step's calculation complexity, we compared the calculation complexity of four different imaging algorithms, we also simulate two target's imaging and target recognition processing. Theoretical analysis and simulation both prove that the proposed algorithm's calculation complexity is improved in various degree compared with the others, thus can be effectively used in target recognition.

  3. Targeting FAK Radiosensitizes 3-Dimensional Grown Human HNSCC Cells Through Reduced Akt1 and MEK1/2 Signaling

    Energy Technology Data Exchange (ETDEWEB)

    Hehlgans, Stephanie [OncoRay-National Center for Radiation Research in Oncology, Medical Faculty Carl Gustav Carus, Dresden University of Technology, Dresden (Germany); Department of Radiotherapy and Oncology, University of Frankfurt, Frankfurt am Main (Germany); Institute of Radiopharmacy, Helmholtz Center Dresden-Rossendorf, Dresden (Germany); Eke, Iris [OncoRay-National Center for Radiation Research in Oncology, Medical Faculty Carl Gustav Carus, Dresden University of Technology, Dresden (Germany); Cordes, Nils, E-mail: Nils.Cordes@OncoRay.de [OncoRay-National Center for Radiation Research in Oncology, Medical Faculty Carl Gustav Carus, Dresden University of Technology, Dresden (Germany); Institute of Radiopharmacy, Helmholtz Center Dresden-Rossendorf, Dresden (Germany); Department of Radiation Oncology, University Hospital and Medical Faculty Carl Gustav Carus, Dresden University of Technology, Dresden (Germany)

    2012-08-01

    Purpose: Focal adhesion kinase (FAK), a main regulator of integrin signaling and cell migration, is frequently overexpressed and hyperphosphorylated in human head-and-neck squamous cell carcinoma (HNSCC). We have previously shown that pharmacologic FAK inhibition leads to radiosensitization of 3-dimensionally grown HNSCC cell lines. To further evaluate the role of FAK in radioresistance and as a potential cancer target, we examined FAK and FAK downstream signaling in HNSCC cell lines grown in more physiologic extracellular matrix-based 3-dimensional cell cultures. Methods and Materials: Seven HNSCC cell lines were grown in 3-dimensional extracellular matrix and the clonogenic radiation survival, expression, and phosphorylation of FAK, paxillin, Akt1, extracellular signal-regulated kinase (ERK)1/2, and MEK1/2 were analyzed after siRNA-mediated knockdown of FAK, Akt1, MEK1, FAK+Akt1, or FAK+MEK1 compared with controls or stable overexpression of FAK. The role of MEK1/2 for clonogenic survival and signaling was investigated using the MEK inhibitor U0126 with or without irradiation. Results: FAK knockdown moderately or significantly enhanced the cellular radiosensitivity of 3-dimensionally grown HNSCC cells. The FAK downstream targets paxillin, Akt1, and ERK1/2 were substantially dephosphorylated under FAK depletion. FAK overexpression, in contrast, increased radiation survival and paxillin, Akt1, and ERK1/2 phosphorylation. The degree of radiosensitization upon Akt1, ERK1/2, or MEK1 depletion or U0126 was superimposable to FAK knockdown. Combination knockdown conditions (ie, Akt1/FAK, MEK1/FAK, or U0126/FAK) failed to provide additional radiosensitization. Conclusions: Our data provide further evidence for FAK as important determinant of radiation survival, which acts in the same signaling axis as Akt1 and ERK1/2. These data strongly support our hypothesis that FAK is a relevant molecular target for HNSCC radiotherapy.

  4. How to target apoptosis signaling pathways for the treatment of pediatric cancers

    Directory of Open Access Journals (Sweden)

    Simone eFulda

    2013-02-01

    Full Text Available Apoptosis represents one of the most important forms of cell death in higher organisms and is typically dysregulated in human cancers, including pediatric tumors. This implies that ineffective engagement of cell death programs can contribute to tumor formation as well as tumor progression. In addition, the majority of cytotoxic therapeutic principles rely on the activation of cell death signaling pathways in cancer cells. Blockade of signaling networks that lead to cell death can therefore confer treatment resistance. A variety of genetic and epigenetic events as well as dysfunctional regulation of signaling networks have been identified as underlying causes of cell death resistance in childhood malignancies. Apoptosis pathways can be therapeutically exploited by enhancing proapoptotic signals or by neutralizing antiapoptotic programs. The challenge in the coming years will be to successfully transfer this knowledge into the development of innovative treatment approaches for children with cancer.

  5. Signal Transducer and Activator of Transcription 3 in Liver Diseases: A Novel Therapeutic Target

    OpenAIRE

    Wang, Hua; Lafdil, Fouad; Kong, Xiaoni; Gao, Bin

    2011-01-01

    Signal transducer and activator of transcription 3 (STAT3) is a transcription factor that is activated by many cytokines and growth factors and plays a key role in cell survival, proliferation, and differentiation. STAT3 activation is detected virtually in all rodent models of liver injury and in human liver diseases. In this review, we highlight recent advances of STAT3 signaling in liver injury, steatosis, inflammation, regeneration, fibrosis, and hepatocarcinogenesis. The cytokines and sma...

  6. A sequential multi-target Mps1 phosphorylation cascade promotes spindle checkpoint signaling

    OpenAIRE

    Ji, Zhejian; Gao, Haishan; Jia, Luying; Li, Bing; Yu, Hongtao

    2017-01-01

    The master spindle checkpoint kinase Mps1 senses kinetochore-microtubule attachment and promotes checkpoint signaling to ensure accurate chromosome segregation. The kinetochore scaffold Knl1, when phosphorylated by Mps1, recruits checkpoint complexes Bub1?Bub3 and BubR1?Bub3 to unattached kinetochores. Active checkpoint signaling ultimately enhances the assembly of the mitotic checkpoint complex (MCC) consisting of BubR1?Bub3, Mad2, and Cdc20, which inhibits the anaphase-promoting complex or ...

  7. Identification of Novel Plant Peroxisomal Targeting Signals by a Combination of Machine Learning Methods and in Vivo Subcellular Targeting Analyses[W

    Science.gov (United States)

    Lingner, Thomas; Kataya, Amr R.; Antonicelli, Gerardo E.; Benichou, Aline; Nilssen, Kjersti; Chen, Xiong-Yan; Siemsen, Tanja; Morgenstern, Burkhard; Meinicke, Peter; Reumann, Sigrun

    2011-01-01

    In the postgenomic era, accurate prediction tools are essential for identification of the proteomes of cell organelles. Prediction methods have been developed for peroxisome-targeted proteins in animals and fungi but are missing specifically for plants. For development of a predictor for plant proteins carrying peroxisome targeting signals type 1 (PTS1), we assembled more than 2500 homologous plant sequences, mainly from EST databases. We applied a discriminative machine learning approach to derive two different prediction methods, both of which showed high prediction accuracy and recognized specific targeting-enhancing patterns in the regions upstream of the PTS1 tripeptides. Upon application of these methods to the Arabidopsis thaliana genome, 392 gene models were predicted to be peroxisome targeted. These predictions were extensively tested in vivo, resulting in a high experimental verification rate of Arabidopsis proteins previously not known to be peroxisomal. The prediction methods were able to correctly infer novel PTS1 tripeptides, which even included novel residues. Twenty-three newly predicted PTS1 tripeptides were experimentally confirmed, and a high variability of the plant PTS1 motif was discovered. These prediction methods will be instrumental in identifying low-abundance and stress-inducible peroxisomal proteins and defining the entire peroxisomal proteome of Arabidopsis and agronomically important crop plants. PMID:21487095

  8. Phosphoproteomic profiling of in vivo signaling in liver by the mammalian target of rapamycin complex 1 (mTORC1.

    Directory of Open Access Journals (Sweden)

    Gokhan Demirkan

    Full Text Available Our understanding of signal transduction networks in the physiological context of an organism remains limited, partly due to the technical challenge of identifying serine/threonine phosphorylated peptides from complex tissue samples. In the present study, we focused on signaling through the mammalian target of rapamycin (mTOR complex 1 (mTORC1, which is at the center of a nutrient- and growth factor-responsive cell signaling network. Though studied extensively, the mechanisms involved in many mTORC1 biological functions remain poorly understood.We developed a phosphoproteomic strategy to purify, enrich and identify phosphopeptides from rat liver homogenates. Using the anticancer drug rapamycin, the only known target of which is mTORC1, we characterized signaling in liver from rats in which the complex was maximally activated by refeeding following 48 hr of starvation. Using protein and peptide fractionation methods, TiO(2 affinity purification of phosphopeptides and mass spectrometry, we reproducibly identified and quantified over four thousand phosphopeptides. Along with 5 known rapamycin-sensitive phosphorylation events, we identified 62 new rapamycin-responsive candidate phosphorylation sites. Among these were PRAS40, gephyrin, and AMP kinase 2. We observed similar proportions of increased and reduced phosphorylation in response to rapamycin. Gene ontology analysis revealed over-representation of mTOR pathway components among rapamycin-sensitive phosphopeptide candidates.In addition to identifying potential new mTORC1-mediated phosphorylation events, and providing information relevant to the biology of this signaling network, our experimental and analytical approaches indicate the feasibility of large-scale phosphoproteomic profiling of tissue samples to study physiological signaling events in vivo.

  9. Altered Toll-Like Receptor 9 Signaling in Mycobacterium avium subsp. paratuberculosis-Infected Bovine Monocytes Reveals Potential Therapeutic Targets

    Science.gov (United States)

    Arsenault, Ryan J.; Li, Yue; Maattanen, Pekka; Scruten, Erin; Doig, Kimberley; Potter, Andrew; Griebel, Philip; Kusalik, Anthony

    2013-01-01

    Mycobacterium avium subsp. paratuberculosis is the causative agent of Johne's disease in cattle. The complex, multifaceted interaction of M. avium subsp. paratuberculosis with its host includes dampening the ability of infected cells to respond to stimuli that promote M. avium subsp. paratuberculosis clearance. By disrupting host defenses, M. avium subsp. paratuberculosis creates an intracellular environment that favors the establishment and maintenance of infection. Toll-like receptors (TLRs) are important sensors that initiate innate immune responses to microbial challenge and are also immunotherapeutic targets. For example, TLR9 contributes to host defense against M. avium subsp. paratuberculosis, and its agonists (CpG oligodeoxynucleotides [ODNs]) are under investigation for treatment of Johne's disease and other infections. Here we demonstrate that M. avium subsp. paratuberculosis infection changes the responsiveness of bovine monocytes to TLR9 stimulation. M. avium subsp. paratuberculosis inhibits classical TLR9-mediated responses despite a 10-fold increase in TLR9 expression and maintained uptake of CpG ODNs. Other TLR9-mediated responses, such as oxidative burst, which occur through noncanonical signaling, remain functional. Kinome analysis verifies that classic TLR9 signaling is blocked by M. avium subsp. paratuberculosis infection and that signaling instead proceeds through a Pyk2-mediated mechanism. Pyk2-mediated signaling does not hinder infection, as CpG ODNs fail to promote M. avium subsp. paratuberculosis clearance. Indeed, Pyk2 signaling appears to be an important aspect of M. avium subsp. paratuberculosis infection, as Pyk2 inhibitors significantly reduce the number of intracellular M. avium subsp. paratuberculosis bacteria. The actions of M. avium subsp. paratuberculosis on TLR9 signaling may represent a strategy to generate a host environment which is better suited for infection, revealing potential new targets for therapeutic intervention. PMID

  10. Na,K-ATPase β1-subunit is a target of sonic hedgehog signaling and enhances medulloblastoma tumorigenicity.

    Science.gov (United States)

    Lee, Seung Joon; Litan, Alisa; Li, Zhiqin; Graves, Bruce; Lindsey, Stephan; Barwe, Sonali P; Langhans, Sigrid A

    2015-08-19

    The Sonic hedgehog (Shh) signaling pathway plays an important role in cerebellar development, and mutations leading to hyperactive Shh signaling have been associated with certain forms of medulloblastoma, a common form of pediatric brain cancer. While the fundamentals of this pathway are known, the molecular targets contributing to Shh-mediated proliferation and transformation are still poorly understood. Na,K-ATPase is a ubiquitous enzyme that maintains intracellular ion homeostasis and functions as a signaling scaffold and a cell adhesion molecule. Changes in Na,K-ATPase function and subunit expression have been reported in several cancers and loss of the β1-subunit has been associated with a poorly differentiated phenotype in carcinoma but its role in medulloblastoma progression is not known. Human medulloblastoma cell lines and primary cultures of cerebellar granule cell precursors (CGP) were used to determine whether Shh regulates Na,K-ATPase expression. Smo/Smo medulloblastoma were used to assess the Na,K-ATPase levels in vivo. Na,K-ATPase β1-subunit was knocked down in DAOY cells to test its role in medulloblastoma cell proliferation and tumorigenicity. Na,K-ATPase β1-subunit levels increased with differentiation in normal CGP cells. Activation of Shh signaling resulted in reduced β1-subunit mRNA and protein levels and was mimicked by overexpression of Gli1and Bmi1, both members of the Shh signaling cascade; overexpression of Bmi1 reduced β1-subunit promoter activity. In human medulloblastoma cells, low β1-subunit levels were associated with increased cell proliferation and in vivo tumorigenesis. Na,K-ATPase β1-subunit is a target of the Shh signaling pathway and loss of β1-subunit expression may contribute to tumor development and progression not only in carcinoma but also in medulloblastoma, a tumor of neuronal origin.

  11. Targeting cFMS signaling to restore immune function and eradicate HIV reservoirs

    Science.gov (United States)

    Gerngross, Lindsey

    While combination anti-retroviral therapy (cART) has improved the length and quality of life of individuals living with HIV-1 infection, the prevalence of HIV-associated neurocognitive disorders (HAND) has increased and remains a significant clinical concern. The neuropathogenesis of HAND is not completely understood, however, latent HIV infection in the central nervous system (CNS) and chronic neuroinflammation are believed to play a prominent role. CNS-associated macrophages and resident microglia are significant contributors to CNS inflammation and constitute the chief reservoir of HIV-1 infection in the CNS. Previous studies from our lab suggest monocyte/macrophage invasion of the CNS in HIV may be driven by altered monocyte/macrophage homeostasis. We have reported expansion of a monocyte subset (CD14+CD16 +CD163+) in peripheral blood of HIV+ patients that is phenotypically similar to macrophages/microglia that accumulate in the CNS as seen in post-mortem tissue. The factors driving the expansion of this monocyte subset are unknown, however, signaling through cFMS, a type III receptor tyrosine kinase (RTK), may play a role. Macrophage-colony stimulating factor (M-CSF), a ligand of cFMS, has been shown to be elevated in the cerebral spinal fluid (CSF) of individuals with the most severe form of HAND, HIV-associated dementia (HAD). M-CSF promotes a Macrophage-2-like phenotype and increases CD16 and CD163 expression in cultured monocytes. M-CSF has also been shown to increase the susceptibility of macrophages to HIV infection and enhance virus production. These findings, in addition to the known function of M-CSF in promoting macrophage survival, supports a role for M-CSF in the development and maintenance of macrophage viral reservoirs in tissues where these cells accumulate, including the CNS. Interestingly, a second ligand for cFMS, IL-34, was recently identified and reported to share some functions with M-CSF, suggesting that both ligands may contribute to HIV

  12. Selective attention modulates the effect of target location probability on redundant signal processing.

    Science.gov (United States)

    Chang, Ting-Yun; Little, Daniel R; Yang, Cheng-Ta

    2016-08-01

    We investigated the decision process underlying the detection of targets at multiple locations. In three experiments using the same observers, target location probability and attentional instructions were manipulated. A redundant-target detection task was conducted in which participants were required to detect a dot presented at one of two locations. When the dot appeared at the two locations with equal frequency (Experiment 1), those participants who were found to have limited to unlimited capacity were shown to adopt a parallel, self-terminating strategy. By contrast, those participants who had supercapacity were shown to process redundant targets in a coactive manner. When targets were presented with unequal probability, two participants adopted a parallel, self-terminating strategy regardless of whether they were informed the target location probability (Experiment 3) or not (Experiment 2). For the remaining two participants, the strategy changed from parallel, self-terminating to serial, self-terminating as a result of the probability instructions. In Experiments 2 and 3, all the participants were of unlimited to limited capacity. Taken together, these results suggest that target location probability differently affects the selection of a decision strategy and highlight the role of controlled attention in selecting a decision strategy.

  13. Detection and localization of multiple short range targets using FMCW radar signal

    KAUST Repository

    Jardak, Seifallah

    2016-07-26

    In this paper, a 24 GHz frequency-modulated continuous wave radar is used to detect and localize both stationary and moving targets. Depending on the application, the implemented software offers different modes of operation. For example, it can simply output raw data samples for advanced offline processing or directly carry out a two dimensional fast Fourier transform to estimate the location and velocity of multiple targets. To suppress clutter and detect only moving targets, two methods based on the background reduction and the slow time processing techniques are implemented. A trade-off between the two methods is presented based on their performance and the required processing time. © 2016 IEEE.

  14. Obstacle Avoidance and Target Acquisition for Robot Navigation Using a Mixed Signal Analog/Digital Neuromorphic Processing System

    Science.gov (United States)

    Milde, Moritz B.; Blum, Hermann; Dietmüller, Alexander; Sumislawska, Dora; Conradt, Jörg; Indiveri, Giacomo; Sandamirskaya, Yulia

    2017-01-01

    Neuromorphic hardware emulates dynamics of biological neural networks in electronic circuits offering an alternative to the von Neumann computing architecture that is low-power, inherently parallel, and event-driven. This hardware allows to implement neural-network based robotic controllers in an energy-efficient way with low latency, but requires solving the problem of device variability, characteristic for analog electronic circuits. In this work, we interfaced a mixed-signal analog-digital neuromorphic processor ROLLS to a neuromorphic dynamic vision sensor (DVS) mounted on a robotic vehicle and developed an autonomous neuromorphic agent that is able to perform neurally inspired obstacle-avoidance and target acquisition. We developed a neural network architecture that can cope with device variability and verified its robustness in different environmental situations, e.g., moving obstacles, moving target, clutter, and poor light conditions. We demonstrate how this network, combined with the properties of the DVS, allows the robot to avoid obstacles using a simple biologically-inspired dynamics. We also show how a Dynamic Neural Field for target acquisition can be implemented in spiking neuromorphic hardware. This work demonstrates an implementation of working obstacle avoidance and target acquisition using mixed signal analog/digital neuromorphic hardware. PMID:28747883

  15. Obstacle Avoidance and Target Acquisition for Robot Navigation Using a Mixed Signal Analog/Digital Neuromorphic Processing System

    Directory of Open Access Journals (Sweden)

    Moritz B. Milde

    2017-07-01

    Full Text Available Neuromorphic hardware emulates dynamics of biological neural networks in electronic circuits offering an alternative to the von Neumann computing architecture that is low-power, inherently parallel, and event-driven. This hardware allows to implement neural-network based robotic controllers in an energy-efficient way with low latency, but requires solving the problem of device variability, characteristic for analog electronic circuits. In this work, we interfaced a mixed-signal analog-digital neuromorphic processor ROLLS to a neuromorphic dynamic vision sensor (DVS mounted on a robotic vehicle and developed an autonomous neuromorphic agent that is able to perform neurally inspired obstacle-avoidance and target acquisition. We developed a neural network architecture that can cope with device variability and verified its robustness in different environmental situations, e.g., moving obstacles, moving target, clutter, and poor light conditions. We demonstrate how this network, combined with the properties of the DVS, allows the robot to avoid obstacles using a simple biologically-inspired dynamics. We also show how a Dynamic Neural Field for target acquisition can be implemented in spiking neuromorphic hardware. This work demonstrates an implementation of working obstacle avoidance and target acquisition using mixed signal analog/digital neuromorphic hardware.

  16. Obstacle Avoidance and Target Acquisition for Robot Navigation Using a Mixed Signal Analog/Digital Neuromorphic Processing System.

    Science.gov (United States)

    Milde, Moritz B; Blum, Hermann; Dietmüller, Alexander; Sumislawska, Dora; Conradt, Jörg; Indiveri, Giacomo; Sandamirskaya, Yulia

    2017-01-01

    Neuromorphic hardware emulates dynamics of biological neural networks in electronic circuits offering an alternative to the von Neumann computing architecture that is low-power, inherently parallel, and event-driven. This hardware allows to implement neural-network based robotic controllers in an energy-efficient way with low latency, but requires solving the problem of device variability, characteristic for analog electronic circuits. In this work, we interfaced a mixed-signal analog-digital neuromorphic processor ROLLS to a neuromorphic dynamic vision sensor (DVS) mounted on a robotic vehicle and developed an autonomous neuromorphic agent that is able to perform neurally inspired obstacle-avoidance and target acquisition. We developed a neural network architecture that can cope with device variability and verified its robustness in different environmental situations, e.g., moving obstacles, moving target, clutter, and poor light conditions. We demonstrate how this network, combined with the properties of the DVS, allows the robot to avoid obstacles using a simple biologically-inspired dynamics. We also show how a Dynamic Neural Field for target acquisition can be implemented in spiking neuromorphic hardware. This work demonstrates an implementation of working obstacle avoidance and target acquisition using mixed signal analog/digital neuromorphic hardware.

  17. WNT antagonist, DKK2, is a Notch signaling target in intestinal stem cells: augmentation of a negative regulation system for canonical WNT signaling pathway by the Notch-DKK2 signaling loop in primates.

    Science.gov (United States)

    Katoh, Masuko; Katoh, Masaru

    2007-01-01

    Notch and WNT signaling pathways are key components of the stem cell signaling network. Canonical WNT signaling to intestinal progenitor cells leads to transcriptional activation of the JAG1 gene, encoding Serrate-type Notch ligand. JAG1 then binds to the Notch receptor on adjacent stem cells to induce Notch receptor proteolyses for the release of Notch intracellular domain (NICD). NICD is associated with CSL/RBPSUH and Mastermind (MAML1, MAML2, or MAML3) to activate Notch target genes, such as HES1 and HES5. Although WNT-dependent Notch signaling activation in intestinal stem cells is clarified, the effects of Notch signaling activation on WNT signaling in progenitor cells remain unclear. We searched for Notch-response element (NRE) in the promoter region of genes encoding secreted WNT signaling inhibitors, including DKK1, DKK2, DKK3, DKK4, SFRP1, SFRP2, SFRP3, SFRP4, SFRP5 and WIF1. Double NREs were identified within human DKK2 promoter by bioinformatics and human intelligence (Humint). The human DKK2 gene was characterized as Notch signaling target in intestinal stem cells. Because DKK2 is a key player in the stem cell signaling network, the DKK2 gene at human chromosome 4q25 is a candidate tumor suppressor gene inactivated due to epigenetic silencing and/or deletion. The chimpanzee DKK2 gene was identified within the NW_105990.1 genome sequence, while the cow Dkk2 gene was identified within the AC156664.2 and AC158038.2 genome sequences. Chimpanzee DKK2 and cow Dkk2 showed 98.5% and 95.8% total-amino-acid identity with human DKK2, respectively. Double NREs in human DKK2 promoter were conserved in chimpanzee DKK2 promoter, partially in rat Dkk2 promoter, but not in cow and mouse Dkk2 promoters. The Notch-DKK2 signaling loop, created or potentiated in primates, was complementary to WNT-DKK1 and BMP-IHH-SFRP1 signaling loops for negative regulation of canonical WNT signaling pathway. Together, these facts indicate that DKK2 promoter evolution resulted in the

  18. An In Vivo Chemical Genetic Screen Identifies Phosphodiesterase 4 as a Pharmacological Target for Hedgehog Signaling Inhibition

    Directory of Open Access Journals (Sweden)

    Charles H. Williams

    2015-04-01

    Full Text Available Hedgehog (Hh signaling plays an integral role in vertebrate development, and its dysregulation has been accepted widely as a driver of numerous malignancies. While a variety of small molecules target Smoothened (Smo as a strategy for Hh inhibition, Smo gain-of-function mutations have limited their clinical implementation. Modulation of targets downstream of Smo could define a paradigm for treatment of Hh-dependent cancers. Here, we describe eggmanone, a small molecule identified from a chemical genetic zebrafish screen, which induced an Hh-null phenotype. Eggmanone exerts its Hh-inhibitory effects through selective antagonism of phosphodiesterase 4 (PDE4, leading to protein kinase A activation and subsequent Hh blockade. Our study implicates PDE4 as a target for Hh inhibition, suggests an improved strategy for Hh-dependent cancer therapy, and identifies a unique probe of downstream-of-Smo Hh modulation.

  19. Targeting cancer stem cell-specific markers and/or associated signaling pathways for overcoming cancer drug resistance.

    Science.gov (United States)

    Ranji, Peyman; Salmani Kesejini, Tayyebali; Saeedikhoo, Sara; Alizadeh, Ali Mohammad

    2016-10-01

    Cancer stem cells (CSCs) are a small subpopulation of tumor cells with capabilities of self-renewal, dedifferentiation, tumorigenicity, and inherent chemo-and-radio therapy resistance. Tumor resistance is believed to be caused by CSCs that are intrinsically challenging to common treatments. A number of CSC markers including CD44, CD133, receptor tyrosine kinase, aldehyde dehydrogenases, epithelial cell adhesion molecule/epithelial specific antigen, and ATP-binding cassette subfamily G member 2 have been proved as the useful targets for defining CSC population in solid tumors. Furthermore, targeting CSC markers through new therapeutic strategies will ultimately improve treatments and overcome cancer drug resistance. Therefore, the identification of novel strategies to increase sensitivity of CSC markers has major clinical implications. This review will focus on the innovative treatment methods such as nano-, immuno-, gene-, and chemotherapy approaches for targeting CSC-specific markers and/or their associated signaling pathways.

  20. Hedgehog signaling pathway regulated the target genes for adipogenesis in silkworm Bombyx mori.

    Science.gov (United States)

    Liang, Shuang; Chen, Rui-Ting; Zhang, Deng-Pan; Xin, Hu-Hu; Lu, Yan; Wang, Mei-Xian; Miao, Yun-Gen

    2015-10-01

    Hedgehog (Hh) signals regulate invertebrate and vertebrate development, yet the role of the pathway in adipose development remains poorly understood. In this report, we found that Hh pathway components are expressed in the fat body of silkworm larvae. Functional analysis of these components in a BmN cell line model revealed that activation of the Hh gene stimulated transcription of Hh pathway components, but inhibited the expression of the adipose marker gene AP2. Conversely, specific RNA interference-mediated knockdown of Hh resulted in increased AP2 expression. This further showed the regulation of Hh signal on the adipose marker gene. In silkworm larval models, enhanced adipocyte differentiation and an increase in adipocyte cell size were observed in silkworms that had been treated with a specific Hh signaling pathway antagonist, cyclopamine. The fat-body-specific Hh blockade tests were consistent with Hh signaling inhibiting silkworm adipogenesis. Our results indicate that the role of Hh signaling in inhibiting fat formation is conserved in vertebrates and invertebrates. © 2014 Institute of Zoology, Chinese Academy of Sciences.

  1. Research and Analysis Laser Target Optics Characteristics and Signal Recognition Processing in Detection Screen System

    National Research Council Canada - National Science Library

    Hanshan Li; Yanran Li

    2014-01-01

      In order to improve the measurement accuracy of the laser measurement distance system, this paper studies the laser target optics characteristics based on the laser detection principle in the laser...

  2. Are suppressors of cytokine signaling proteins recently identified in atherosclerosis possible therapeutic targets?

    Science.gov (United States)

    Tang, Jingjing; Raines, Elaine W

    2005-10-01

    Atherosclerosis is a slowly progressing chronic inflammatory disease characterized by focal arterial lesions that can ultimately occlude the entire blood vessel and lead to sudden death. Lesions associated with cardiovascular events are those enriched in macrophages and other inflammatory cells. Activation of inflammatory cells within lesions induces the release of cytokines which promotes more inflammation and associated tissue damage if cytokine signaling pathways remain unregulated. Thus, pathways capable of suppressing proinflammatory cytokine signaling hold the potential to limit life-threatening cardiovascular events caused by atherogenesis. This review focuses on suppressors of cytokine signaling proteins recently identified in the atherosclerosis-prone ApoE(-/-) mouse and provides perspectives of their potential for intervention in atherosclerotic lesion progression.

  3. A targeted glycan-related gene screen reveals heparan sulfate proteoglycan sulfation regulates WNT and BMP trans-synaptic signaling.

    Directory of Open Access Journals (Sweden)

    Neil Dani

    Full Text Available A Drosophila transgenic RNAi screen targeting the glycan genome, including all N/O/GAG-glycan biosynthesis/modification enzymes and glycan-binding lectins, was conducted to discover novel glycan functions in synaptogenesis. As proof-of-product, we characterized functionally paired heparan sulfate (HS 6-O-sulfotransferase (hs6st and sulfatase (sulf1, which bidirectionally control HS proteoglycan (HSPG sulfation. RNAi knockdown of hs6st and sulf1 causes opposite effects on functional synapse development, with decreased (hs6st and increased (sulf1 neurotransmission strength confirmed in null mutants. HSPG co-receptors for WNT and BMP intercellular signaling, Dally-like Protein and Syndecan, are differentially misregulated in the synaptomatrix of these mutants. Consistently, hs6st and sulf1 nulls differentially elevate both WNT (Wingless; Wg and BMP (Glass Bottom Boat; Gbb ligand abundance in the synaptomatrix. Anterograde Wg signaling via Wg receptor dFrizzled2 C-terminus nuclear import and retrograde Gbb signaling via synaptic MAD phosphorylation and nuclear import are differentially activated in hs6st and sulf1 mutants. Consequently, transcriptional control of presynaptic glutamate release machinery and postsynaptic glutamate receptors is bidirectionally altered in hs6st and sulf1 mutants, explaining the bidirectional change in synaptic functional strength. Genetic correction of the altered WNT/BMP signaling restores normal synaptic development in both mutant conditions, proving that altered trans-synaptic signaling causes functional differentiation defects.

  4. Mammalian Target of Rapamycin Signaling Pathway Changes with Intestinal Epithelial Cells Renewal Along Crypt-Villus Axis

    Directory of Open Access Journals (Sweden)

    Huansheng Yang

    2016-07-01

    Full Text Available Background/Aims: Understanding the mechanism that involves in regulating epithelial cells renewal is the fundamental of regulating intestinal mucosa development and functions and related diseases. The mechanistic target of rapamycin (mTOR signaling pathway involves in controlling various major processes by integrating intracellular and extracellular cues. The present experiment was conducted to test the correlation between the mTOR signaling pathway and intestinal epithelial cells renewal along crypt-villus axis (CVA. Methods: Intestinal epithelial cells were sequentially isolated from the jejunum of piglets along CVA, and the amount or phosphorylation level of proteins involved in cell cycle, mTOR signaling pathway, gene expression, and the antioxidant capacity in the isolated cells were measured. Results: The results showed that the amount of proteins involved in cell cycle decreased from crypt to villus tip. The amount or phosphorylation level of proteins related to mTOR signaling pathway in intestinal epithelial cells mainly decreased during maturation along CVA. The amount of proteins involved in gene expression and the antioxidant capacity also decreased from crypt to the top of villi. Conclusions: These results indicate that the mTOR signaling pathway may be involved in regulating the intestinal epithelial cells renewal along CVA and it may partly through affecting the antioxidant capacity and gene expression of intestinal epithelial cells. Further histological verification is needed to confirm the results of the present experiments.

  5. Dual PDF signaling pathways reset clocks via TIMELESS and acutely excite target neurons to control circadian behavior.

    Science.gov (United States)

    Seluzicki, Adam; Flourakis, Matthieu; Kula-Eversole, Elzbieta; Zhang, Luoying; Kilman, Valerie; Allada, Ravi

    2014-03-01

    Molecular circadian clocks are interconnected via neural networks. In Drosophila, PIGMENT-DISPERSING FACTOR (PDF) acts as a master network regulator with dual functions in synchronizing molecular oscillations between disparate PDF(+) and PDF(-) circadian pacemaker neurons and controlling pacemaker neuron output. Yet the mechanisms by which PDF functions are not clear. We demonstrate that genetic inhibition of protein kinase A (PKA) in PDF(-) clock neurons can phenocopy PDF mutants while activated PKA can partially rescue PDF receptor mutants. PKA subunit transcripts are also under clock control in non-PDF DN1p neurons. To address the core clock target of PDF, we rescued per in PDF neurons of arrhythmic per⁰¹ mutants. PDF neuron rescue induced high amplitude rhythms in the clock component TIMELESS (TIM) in per-less DN1p neurons. Complete loss of PDF or PKA inhibition also results in reduced TIM levels in non-PDF neurons of per⁰¹ flies. To address how PDF impacts pacemaker neuron output, we focally applied PDF to DN1p neurons and found that it acutely depolarizes and increases firing rates of DN1p neurons. Surprisingly, these effects are reduced in the presence of an adenylate cyclase inhibitor, yet persist in the presence of PKA inhibition. We have provided evidence for a signaling mechanism (PKA) and a molecular target (TIM) by which PDF resets and synchronizes clocks and demonstrates an acute direct excitatory effect of PDF on target neurons to control neuronal output. The identification of TIM as a target of PDF signaling suggests it is a multimodal integrator of cell autonomous clock, environmental light, and neural network signaling. Moreover, these data reveal a bifurcation of PKA-dependent clock effects and PKA-independent output effects. Taken together, our results provide a molecular and cellular basis for the dual functions of PDF in clock resetting and pacemaker output.

  6. Toxcast Chemical and Bioactivity Profiles for In Vitro Targets in the Retinoid Signaling System (SOT)

    Science.gov (United States)

    A predictive model for prenatal developmental toxicity using ToxCast Phase I showed the RAR assay set to be the strongest weighting factor (Sipes et al. 2011). Retinoid signaling mediates growth and differentiation of the embryo. ToxCast has 6 reporter assays for trans-activation...

  7. Functional Proteomic Analysis of Signaling Networks and Response to Targeted Therapy

    Science.gov (United States)

    2009-03-01

    Jordan JD, Landau EM, Iyengar R (2000) Signaling networks: the origins of cellular multitasking. Cell 103: 193–200. 53. Eungdamrong NJ, Iyengar R...I, Shamir R (2009) Identifying functional modules using expression profiles and confidence-scored protein interactions. Bioinformatics 25: 1158–1164

  8. Locating proteins in the cell using TargetP, SignalP and related tools

    DEFF Research Database (Denmark)

    Emanuelsson, O.; Brunak, Søren; von Heijne, G.

    2007-01-01

    of methods to predict subcellular localization based on these sorting signals and other sequence properties. We then outline how to use a number of internet-accessible tools to arrive at a reliable subcellular localization prediction for eukaryotic and prokaryotic proteins. In particular, we provide detailed...

  9. Nuclear hormone receptor signals as new therapeutic targets for urothelial carcinoma.

    Science.gov (United States)

    Miyamoto, H; Zheng, Y; Izumi, K

    2012-01-01

    Unlike prostate and breast cancers, urothelial carcinoma of the urinary bladder is not yet considered as an endocrine-related neoplasm, and hormonal therapy for bladder cancer remains experimental. Nonetheless, there is increasing evidence indicating that nuclear hormone receptor signals are implicated in the development and progression of bladder cancer. Androgen-mediated androgen receptor (AR) signals have been convincingly shown to induce bladder tumorigenesis. Androgens also promote the growth of AR-positive bladder cancer cells, although it is controversial whether AR plays a dominant role in bladder cancer progression. Both stimulatory and inhibitory functions of estrogen receptor signals in bladder cancer have been reported. Various studies have also demonstrated the involvement of other nuclear receptors, including progesterone receptor, glucocorticoid receptor, vitamin D receptor, and retinoid receptors, as well as some orphan receptors, in bladder cancer. This review summarizes and discusses available data suggesting the modulation of bladder carcinogenesis and cancer progression via nuclear hormone receptor signaling pathways. These pathways have the potential to be an extremely important area of bladder cancer research, leading to the development of effective chemopreventive/therapeutic approaches, using hormonal manipulation. Considerable uncertainty remains regarding the selection of patients who are likely to benefit from hormonal therapy and optimal options for the treatment.

  10. Targeting the epithelial to mesenchymal transition in glioblastoma: the emerging role of MET signaling

    Directory of Open Access Journals (Sweden)

    Lee JK

    2014-10-01

    Full Text Available Jin-Ku Lee,1,2,* Kyeung Min Joo,3 Jeongwu Lee,4 Yeup Yoon,5,* Do-Hyun Nam2,5 1Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Seoul, Korea; 2Department of Neurosurgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea; 3Department of Anatomy and Cell Biology, Sungkyunkwan University School of Medicine, Seoul, Korea; 4Department of Stem Cell Biology and Regenerative Medicine, Lerner Research Institute, Cleveland Clinic, Cleveland, OH, USA; 5Samsung Advanced Institute for Health Sciences and Technology, Sungkyunkwan University School of Medicine, Seoul, Korea  *These authors contributed equally to this work Abstract: Glioblastoma multiforme (GBM is the most common human primary brain malignancy and has a dismal prognosis. Aggressive treatments using maximal surgical resection, radiotherapy, and temozolomide result in median survival of only 14.6 months in patients with GBM. Numerous clinical approaches using small molecule inhibitors have shown disappointing results because of the genetic heterogeneity of GBM. The epithelial to mesenchymal transition (EMT is a crucial biological process occurring in the early development stages of many species. However, cancer cells often obtain the ability to invade and metastasize through the EMT, which triggers the scattering of cells. The hepatocyte growth factor (HGF/MET signaling pathway is indicative of the EMT during both embryogenesis and the invasive growth of tumors, because HGF potently induces mesenchymal transition in epithelial-driven cells. Activation of MET signaling or co-overexpression of HGF and MET frequently represents aggressive growth and poor prognosis of various cancers, including GBM. Thus, efforts to treat cancers by inhibiting MET signaling using neutralizing antibodies or small molecule inhibitors have progressed during the last decade. In this review, we discuss HGF/MET signaling in the development of diseases

  11. A Novel Chemical Inhibitor of ABA Signaling Targets All ABA Receptors.

    Science.gov (United States)

    Ye, Yajin; Zhou, Lijuan; Liu, Xue; Liu, Hao; Li, Deqiang; Cao, Minjie; Chen, Haifeng; Xu, Lin; Zhu, Jian-Kang; Zhao, Yang

    2017-04-01

    Abscisic acid (ABA), the most important stress-induced phytohormone, regulates seed dormancy, germination, plant senescence, and the abiotic stress response. ABA signaling is repressed by group A type 2C protein phosphatases (PP2Cs), and then ABA binds to its receptor of the ACTIN RESISTANCE1 (PYR1), PYR1-LIKE (PYL), and REGULATORY COMPONENTS OF ABA RECEPTORS (RCAR) family, which, in turn, inhibits PP2Cs and activates downstream ABA signaling. The agonist/antagonist of ABA receptors have the potential to reveal the ABA signaling machinery and to become lead compounds for agrochemicals; however, until now, no broad-spectrum antagonists of ABA receptors blocking all PYR/PYL-PP2C interactions have been identified. Here, using chemical genetics screenings, we identified ABA ANTAGONIST1 (AA1), the first broad-spectrum antagonist of ABA receptors in Arabidopsis (Arabidopsis thaliana). Physiological analyses revealed that AA1 is sufficiently active to block ABA signaling. AA1 interfered with all the PYR/PYL-HAB1 interactions, and the diminished PYR/PYL-HAB1 interactions, in turn, restored the activity of HAB1. AA1 binds to all 13 members. Molecular dockings, the non-AA1-bound PYL2 variant, and competitive binding assays demonstrated that AA1 enters into the ligand-binding pocket of PYL2. Using AA1, we tested the genetic relationships of ABA receptors with other core components of ABA signaling, demonstrating that AA1 is a powerful tool with which to sidestep this genetic redundancy of PYR/PYLs. In addition, the application of AA1 delays leaf senescence. Thus, our study developed an efficient broad-spectrum antagonist of ABA receptors and demonstrated that plant senescence can be chemically controlled through AA1, with a simple and easy-to-synthesize structure, allowing its availability and utility as a chemical probe synthesized in large quantities, indicating its potential application in agriculture. © 2017 American Society of Plant Biologists. All Rights Reserved.

  12. Targeting of macrophage galactose-type C-type lectin (MGL) induces DC signaling and activation

    DEFF Research Database (Denmark)

    Napoletano, Chiara; Zizzari, Ilaria G; Rughetti, Aurelia

    2012-01-01

    Dendritic cells (DCs) sense the microenvironment through several types of receptors recognizing pathogen-associated molecular patterns. In particular, C-type lectins, expressed by distinct subsets of DCs, recognize and internalize specific carbohydrate antigen in a Ca(2+) -dependent manner....... Targeting of these receptors is becoming an efficient strategy of delivering antigens in DC-based anticancer immunotherapy. Here we investigated the role of the macrophage galactose type C-lectin receptor (MGL), expressed by immature DCs (iDCs), as a molecular target for a-N-acetylgalactosamine (Gal...

  13. The CD47-signal regulatory protein alpha (SIRPa) interaction is a therapeutic target for human solid tumors.

    Science.gov (United States)

    Willingham, Stephen B; Volkmer, Jens-Peter; Gentles, Andrew J; Sahoo, Debashis; Dalerba, Piero; Mitra, Siddhartha S; Wang, Jian; Contreras-Trujillo, Humberto; Martin, Robin; Cohen, Justin D; Lovelace, Patricia; Scheeren, Ferenc A; Chao, Mark P; Weiskopf, Kipp; Tang, Chad; Volkmer, Anne Kathrin; Naik, Tejaswitha J; Storm, Theresa A; Mosley, Adriane R; Edris, Badreddin; Schmid, Seraina M; Sun, Chris K; Chua, Mei-Sze; Murillo, Oihana; Rajendran, Pradeep; Cha, Adriel C; Chin, Robert K; Kim, Dongkyoon; Adorno, Maddalena; Raveh, Tal; Tseng, Diane; Jaiswal, Siddhartha; Enger, Per Øyvind; Steinberg, Gary K; Li, Gordon; So, Samuel K; Majeti, Ravindra; Harsh, Griffith R; van de Rijn, Matt; Teng, Nelson N H; Sunwoo, John B; Alizadeh, Ash A; Clarke, Michael F; Weissman, Irving L

    2012-04-24

    CD47, a "don't eat me" signal for phagocytic cells, is expressed on the surface of all human solid tumor cells. Analysis of patient tumor and matched adjacent normal (nontumor) tissue revealed that CD47 is overexpressed on cancer cells. CD47 mRNA expression levels correlated with a decreased probability of survival for multiple types of cancer. CD47 is a ligand for SIRPα, a protein expressed on macrophages and dendritic cells. In vitro, blockade of CD47 signaling using targeted monoclonal antibodies enabled macrophage phagocytosis of tumor cells that were otherwise protected. Administration of anti-CD47 antibodies inhibited tumor growth in orthotopic immunodeficient mouse xenotransplantation models established with patient tumor cells and increased the survival of the mice over time. Anti-CD47 antibody therapy initiated on larger tumors inhibited tumor growth and prevented or treated metastasis, but initiation of the therapy on smaller tumors was potentially curative. The safety and efficacy of targeting CD47 was further tested and validated in immune competent hosts using an orthotopic mouse breast cancer model. These results suggest all human solid tumor cells require CD47 expression to suppress phagocytic innate immune surveillance and elimination. These data, taken together with similar findings with other human neoplasms, show that CD47 is a commonly expressed molecule on all cancers, its function to block phagocytosis is known, and blockade of its function leads to tumor cell phagocytosis and elimination. CD47 is therefore a validated target for cancer therapies.

  14. Toxoplasma gondii induces FAK-Src-STAT3 signaling during infection of host cells that prevents parasite targeting by autophagy.

    Directory of Open Access Journals (Sweden)

    Jose-Andres C Portillo

    2017-10-01

    Full Text Available Targeting of Toxoplasma gondii by autophagy is an effective mechanism by which host cells kill the protozoan. Thus, the parasite must avoid autophagic targeting to survive. Here we show that the mammalian cytoplasmic molecule Focal Adhesion Kinase (FAK becomes activated during invasion of host cells. Activated FAK appears to accompany the formation of the moving junction (as assessed by expression the parasite protein RON4. FAK activation was inhibited by approaches that impaired β1 and β3 integrin signaling. FAK caused activation of Src that in turn mediated Epidermal Growth Factor Receptor (EGFR phosphorylation at the unique Y845 residue. Expression of Src-resistant Y845F EGFR mutant markedly inhibited ROP16-independent activation of STAT3 in host cells. Activation of FAK, Y845 EGFR or STAT3 prevented activation of PKR and eIF2α, key stimulators of autophagy. Genetic or pharmacologic inhibition of FAK, Src, EGFR phosphorylation at Y845, or STAT3 caused accumulation of the autophagy protein LC3 and LAMP-1 around the parasite and parasite killing dependent on autophagy proteins (ULK1 and Beclin 1 and lysosomal enzymes. Parasite killing was inhibited by expression of dominant negative PKR. Thus, T. gondii activates a FAK→Src→Y845-EGFR→STAT3 signaling axis within mammalian cells, thereby enabling the parasite to survive by avoiding autophagic targeting through a mechanism likely dependent on preventing activation of PKR and eIF2α.

  15. Yeast carboxypeptidase Y vacuolar targeting signal is defined by four propeptide amino acids

    DEFF Research Database (Denmark)

    Valls, L A; Winther, Jakob R.; Stevens, T H

    1990-01-01

    The amino-terminal propeptide of carboxypeptidase Y (CPY) is necessary and sufficient for targeting this glycoprotein to the vacuole of Saccharomyces cerevisiae. A 16 amino acid stretch of the propeptide was subjected to region-directed mutagenesis using randomized oligonucleotides. Mutations alt...

  16. Study of time-reversal-based signal processing applied to polarimetric GPR detection of elongated targets

    Science.gov (United States)

    Santos, Vinicius Rafael N.; Teixeira, Fernando L.

    2017-04-01

    Ground penetrating radar (GPR) is a useful sensing modality for mapping and identification of underground infrastructure networks, such as metal and concrete pipes (gas, water or sewer), phone conduits or cables, and other buried objects. Due to the polarization-dependent response of typical targets, it is of interest to investigate the optimum antenna arrangement and/or combination of arrangements that maximize the detection and classification capabilities of polarimetric GPR imaging systems. Here, we provide a preliminary study of time-reversal-based techniques applied to target detection by GPR utilizing different relative orientations of linear-polarized antenna elements (with respect to each other, as well as to the targets). We modeled three different pipe materials (metallic, plastic and concrete) and GPR systems operating at center frequencies of 100 MHz and 200 MHz. Full-wave numerical simulations are adopted to account for mutual coupling between targets. This type of assessment study may contribute to the improvement of GPR data interpretation of infrastructure networks in urban area surveys and in other engineering studies.

  17. Analysis of oncogenic signaling networks in glioblastoma identifies ASPM as a molecular target.

    Science.gov (United States)

    Horvath, S; Zhang, B; Carlson, M; Lu, K V; Zhu, S; Felciano, R M; Laurance, M F; Zhao, W; Qi, S; Chen, Z; Lee, Y; Scheck, A C; Liau, L M; Wu, H; Geschwind, D H; Febbo, P G; Kornblum, H I; Cloughesy, T F; Nelson, S F; Mischel, P S

    2006-11-14

    Glioblastoma is the most common primary malignant brain tumor of adults and one of the most lethal of all cancers. Patients with this disease have a median survival of 15 months from the time of diagnosis despite surgery, radiation, and chemotherapy. New treatment approaches are needed. Recent works suggest that glioblastoma patients may benefit from molecularly targeted therapies. Here, we address the compelling need for identification of new molecular targets. Leveraging global gene expression data from two independent sets of clinical tumor samples (n = 55 and n = 65), we identify a gene coexpression module in glioblastoma that is also present in breast cancer and significantly overlaps with the "metasignature" for undifferentiated cancer. Studies in an isogenic model system demonstrate that this module is downstream of the mutant epidermal growth factor receptor, EGFRvIII, and that it can be inhibited by the epidermal growth factor receptor tyrosine kinase inhibitor Erlotinib. We identify ASPM (abnormal spindle-like microcephaly associated) as a key gene within this module and demonstrate its overexpression in glioblastoma relative to normal brain (or body tissues). Finally, we show that ASPM inhibition by siRNA-mediated knockdown inhibits tumor cell proliferation and neural stem cell proliferation, supporting ASPM as a potential molecular target in glioblastoma. Our weighted gene coexpression network analysis provides a blueprint for leveraging genomic data to identify key control networks and molecular targets for glioblastoma, and the principle eluted from our work can be applied to other cancers.

  18. Tracking of Noncooperative Airborne Targets Using ADS-B Signal and Radar Sensing

    Directory of Open Access Journals (Sweden)

    Ming-Shih Huang

    2013-01-01

    Full Text Available As the Automatic Dependent Surveillance-Broadcast (ADS-B system has gained wide acceptance, additional exploitations of the radioed satellite-based information are topics of current interest. One such opportunity includes the augmentation of the communication ADS-B signal with a random biphase modulation for concurrent use as a radar signal. This paper addresses the formulation and analysis of a suitable noncooperative multitarget tracking method for the ADS-B radar system using radar ranging techniques and particle filter algorithms. In addition, the low-update-rate measurement due to the ADS-B system specification is discussed in order to provide acceptable estimation results. Simulation results show satisfactory tracking capability up to several kilometers with acceptable accuracy.

  19. Tocotrienols target PI3K/Akt signaling in anti-breast cancer therapy.

    Science.gov (United States)

    Sylvester, Paul W; Ayoub, Nehad M

    2013-09-01

    The PI3K/Akt signaling pathway mediates mitogen-dependent growth and survival in various types of cancer cells, and inhibition of this pathway results in tumor cell growth arrest and apoptosis. Tocotrienols are natural forms of vitamin E that displays potent anticancer activity at treatment doses that had little or no effect on normal cell viability. Mechanistic studies revealed that the anticancer effects of γ-tocotrienol were associated with a suppression in PI3K/Akt signaling. Additional studies showed that cytotoxic LD50 doses of γ-tocotrienol were 3-5-fold higher than growth inhibitory IC50 treatment doses, suggesting that cytotoxic and antiproliferative effects of γ-tocotrienol might be mediated through different mechanisms. However, γ-tocotrienol-induced caspase activation and apoptosis in mammary tumor cells was also found to be associated with suppression in intracellular PI3K/Akt signaling and subsequent down-regulation of FLIP, an endogenous inhibitor of caspase processing and activation. Since breast cancer cells are significantly more sensitive to the inhibitory effects of γ-tocotrienol on PI3K/Akt signaling than normal cells, these findings suggest that γ-tocotrienol may provide significant health benefits in reducing the risk of breast cancer in women. Studies have also shown that combined treatment of γ-tocotrienol with other chemotherapeutic agents can result in a synergistic anticancer response. Combination therapy was most effective when the anticancer mechanism of action of γ-tocotrienol is complimentary to that of the other drug and can provide significant health benefits in the prevention and/or treatment of breast cancer, while at the same time avoiding tumor resistance or toxic effects that is commonly associated with high dose monotherapy.

  20. Therapeutic Potential for Targeting the Suppressor of Cytokine Signalling-1 Pathway for the Treatment of SLE.

    Science.gov (United States)

    Sukka-Ganesh, B; Larkin, J

    2016-11-01

    Although the specific events dictating systemic lupus erythematosus (SLE) pathology remain unclear, abundant evidence indicates a critical role for dysregulated cytokine signalling in disease progression. Notably, the suppressor of cytokine signalling (SOCS) family of intracellular proteins, in particular the kinase inhibitory region (KIR) bearing SOCS1 and SOCS3, plays a critical role in regulating cytokine signalling. To assess a relationship between SOCS1/SOCS3 expression and SLE, the goals of this study were to (1) evaluate the time kinetics of SOCS1/SOCS3 message and protein expression based on SLE-associated stimulations, (2) compare levels of SOCS1 and SOCS3 present in SLE patients and healthy controls by message and protein, (3) relate SOCS1/SOCS3 expression to inflammatory markers in SLE patients and (4) correlate SOCS1/SOCS3 levels to current treatments. We found that SOCS1 and SOCS3 were most abundant in murine splenic samples at 48 h subsequent to stimulation by anti-CD3, LPS or interferon-gamma. In addition, significant reductions in SOCS1 and SOCS3 were present within PMBCs of SLE patients compared to controls by both mRNA and protein expression. We also found that decreased levels of SOCS1 in SLE patients were correlated with enhanced levels of inflammatory markers and upregulated expression of MHC class II. Finally, we show that patients receiving steroid treatment possessed higher levels SOCS1 compared to SLE patient counterparts and that steroid administration to human PBMCs upregulated SOCS1 message in a dose- and time-dependent manner. Together, these results suggest that therapeutic strategies focused on SOCS1 signalling may have efficacy in the treatment of SLE. © 2016 The Foundation for the Scandinavian Journal of Immunology.

  1. Notch signaling pathway targeted therapy suppresses tumor progression and metastatic spread in pancreatic cancer

    OpenAIRE

    Yabuuchi, Shinichi; Pai, Shweta G; Campbell, Nathaniel R.; de F. Wilde, Roeland; Oliveira,Elizabeth de; Korangath, Preethi; Streppel, Mirte; Rasheed, Zeshaan A.; Hidalgo, Manuel; Maitra, Anirban; Rajeshkumar, N.V.

    2013-01-01

    Pancreatic ductal adenocarcinoma (PDA) remains a lethal human malignancy with historically limited success in treatment. The role of aberrant Notch signaling, which requires the constitutive activation of γ-secretase, in the initiation and progression of PDA is well defined and inhibitors of this pathway are currently in clinical trials. Here we investigated the in vivo therapeutic effect of PF-03084014, a selective γ-secretase inhibitor, alone and in combination with gemcitabine in pancreati...

  2. Chemical approaches to discovery and study of sources and targets of hydrogen peroxide redox signaling through NADPH oxidase proteins.

    Science.gov (United States)

    Brewer, Thomas F; Garcia, Francisco J; Onak, Carl S; Carroll, Kate S; Chang, Christopher J

    2015-01-01

    Hydrogen peroxide (H2O2) is a prime member of the reactive oxygen species (ROS) family of molecules produced during normal cell function and in response to various stimuli, but if left unchecked, it can inflict oxidative damage on all types of biological macromolecules and lead to cell death. In this context, a major source of H2O2 for redox signaling purposes is the NADPH oxidase (Nox) family of enzymes, which were classically studied for their roles in phagocytic immune response but have now been found to exist in virtually all mammalian cell types in various isoforms with distinct tissue and subcellular localizations. Downstream of this tightly regulated ROS generation, site-specific, reversible covalent modification of proteins, particularly oxidation of cysteine thiols to sulfenic acids, represents a prominent posttranslational modification akin to phosphorylation as an emerging molecular mechanism for transforming an oxidant signal into a dynamic biological response. We review two complementary types of chemical tools that enable (a) specific detection of H2O2 generated at its sources and (b) mapping of sulfenic acid posttranslational modification targets that mediate its signaling functions, which can be used to study this important chemical signal in biological systems.

  3. The mechanical activation of mTOR signaling: an emerging role for late endosome/lysosomal targeting.

    Science.gov (United States)

    Jacobs, Brittany L; Goodman, Craig A; Hornberger, Troy A

    2014-02-01

    It is well recognized that mechanical signals play a critical role in the regulation of skeletal muscle mass, and the maintenance of muscle mass is essential for mobility, disease prevention and quality of life. Furthermore, over the last 15 years it has become established that signaling through a protein kinase called the mammalian (or mechanistic) target of rapamycin (mTOR) is essential for mechanically-induced changes in protein synthesis and muscle mass, however, the mechanism(s) via which mechanical stimuli regulate mTOR signaling have not been defined. Nonetheless, advancements are being made, and an emerging body of evidence suggests that the late endosome/lysosomal (LEL) system might play a key role in this process. Therefore, the purpose of this review is to summarize this body of evidence. Specifically, we will first explain why the Ras homologue enriched in brain (Rheb) and phosphatidic acid (PA) are considered to be direct activators of mTOR signaling. We will then describe the process of endocytosis and its involvement in the formation of LEL structures, as well as the evidence which indicates that mTOR and its direct activators (Rheb and PA) are all enriched at the LEL. Finally, we will summarize the evidence that has implicated the LEL in the regulation of mTOR by various growth regulatory inputs such as amino acids, growth factors and mechanical stimuli.

  4. Interaction networks of lithium and valproate molecular targets reveal a striking enrichment of apoptosis functional clusters and neurotrophin signaling.

    Science.gov (United States)

    Gupta, A; Schulze, T G; Nagarajan, V; Akula, N; Corona, W; Jiang, X-y; Hunter, N; McMahon, F J; Detera-Wadleigh, S D

    2012-08-01

    The overall neurobiological mechanisms by which lithium and valproate stabilize mood in bipolar disorder patients have yet to be fully defined. The therapeutic efficacy and dissimilar chemical structures of these medications suggest that they perturb both shared and disparate cellular processes. To investigate key pathways and functional clusters involved in the global action of lithium and valproate, we generated interaction networks formed by well-supported drug targets. Striking functional similarities emerged. Intersecting nodes in lithium and valproate networks highlighted a strong enrichment of apoptosis clusters and neurotrophin signaling. Other enriched pathways included MAPK, ErbB, insulin, VEGF, Wnt and long-term potentiation indicating a widespread effect of both drugs on diverse signaling systems. MAPK1/3 and AKT1/2 were the most preponderant nodes across pathways suggesting a central role in mediating pathway interactions. The convergence of biological responses unveils a functional signature for lithium and valproate that could be key modulators of their therapeutic efficacy.

  5. Characterization of sub-nuclear changes in Caenorhabditis elegans embryos exposed to brief, intermediate and long-term anoxia to analyze anoxia-induced cell cycle arrest.

    Science.gov (United States)

    Hajeri, Vinita A; Trejo, Jesus; Padilla, Pamela A

    2005-12-20

    The soil nematode C. elegans survives oxygen-deprived conditions (anoxia; animation in which cell cycle progression reversibly arrests. The majority of blastomeres of embryos exposed to anoxia arrest at interphase, prophase and metaphase. The spindle checkpoint proteins SAN-1 and MDF-2 are required for embryos to survive 24 hours of anoxia. To further investigate the mechanism of cell-cycle arrest we examined and compared sub-nuclear changes such as chromatin localization pattern, post-translational modification of histone H3, spindle microtubules, and localization of the spindle checkpoint protein SAN-1 with respect to various anoxia exposure time points. To ensure analysis of embryos exposed to anoxia and not post-anoxic recovery we fixed all embryos in an anoxia glove box chamber. Embryos exposed to brief periods to anoxia (30 minutes) contain prophase blastomeres with chromosomes in close proximity to the nuclear membrane, condensation of interphase chromatin and metaphase blastomeres with reduced spindle microtubules density. Embryos exposed to longer periods of anoxia (1-3 days) display several characteristics including interphase chromatin that is further condensed and in close proximity to the nuclear membrane, reduction in spindle structure perimeter and reduced localization of SAN-1 at the kinetochore. Additionally, we show that the spindle checkpoint protein SAN-1 is required for brief periods of anoxia-induced cell cycle arrest, thus demonstrating that this gene product is vital for early anoxia responses. In this report we suggest that the events that occur as an immediate response to brief periods of anoxia directs cell cycle arrest. From our results we conclude that the sub-nuclear characteristics of embryos exposed to anoxia depends upon exposure time as assayed using brief (30 minutes), intermediate (6 or 12 hours) or long-term (24 or 72 hours) exposures. Analyzing these changes will lead to an understanding of the mechanisms required for

  6. Characterization of sub-nuclear changes in Caenorhabditis elegans embryos exposed to brief, intermediate and long-term anoxia to analyze anoxia-induced cell cycle arrest

    Directory of Open Access Journals (Sweden)

    Trejo Jesus

    2005-12-01

    Full Text Available Abstract Background The soil nematode C. elegans survives oxygen-deprived conditions (anoxia; 2 by entering into a state of suspended animation in which cell cycle progression reversibly arrests. The majority of blastomeres of embryos exposed to anoxia arrest at interphase, prophase and metaphase. The spindle checkpoint proteins SAN-1 and MDF-2 are required for embryos to survive 24 hours of anoxia. To further investigate the mechanism of cell-cycle arrest we examined and compared sub-nuclear changes such as chromatin localization pattern, post-translational modification of histone H3, spindle microtubules, and localization of the spindle checkpoint protein SAN-1 with respect to various anoxia exposure time points. To ensure analysis of embryos exposed to anoxia and not post-anoxic recovery we fixed all embryos in an anoxia glove box chamber. Results Embryos exposed to brief periods to anoxia (30 minutes contain prophase blastomeres with chromosomes in close proximity to the nuclear membrane, condensation of interphase chromatin and metaphase blastomeres with reduced spindle microtubules density. Embryos exposed to longer periods of anoxia (1–3 days display several characteristics including interphase chromatin that is further condensed and in close proximity to the nuclear membrane, reduction in spindle structure perimeter and reduced localization of SAN-1 at the kinetochore. Additionally, we show that the spindle checkpoint protein SAN-1 is required for brief periods of anoxia-induced cell cycle arrest, thus demonstrating that this gene product is vital for early anoxia responses. In this report we suggest that the events that occur as an immediate response to brief periods of anoxia directs cell cycle arrest. Conclusion From our results we conclude that the sub-nuclear characteristics of embryos exposed to anoxia depends upon exposure time as assayed using brief (30 minutes, intermediate (6 or 12 hours or long-term (24 or 72 hours exposures

  7. Immunohistochemical Analysis of the Mechanistic Target of Rapamycin and Hypoxia Signalling Pathways in Basal Cell Carcinoma and Trichoepithelioma

    Science.gov (United States)

    Brinkhuizen, Tjinta; Weijzen, Chantal A. H.; Eben, Jonathan; Thissen, Monique R.; van Marion, Ariënne M.; Lohman, Björn G.; Winnepenninckx, Véronique J. L.; Nelemans, Patty J.; van Steensel, Maurice A. M.

    2014-01-01

    Background Basal cell carcinoma (BCC) is the most common cancer in Caucasians. Trichoepithelioma (TE) is a benign neoplasm that strongly resembles BCC. Both are hair follicle (HF) tumours. HFs are hypoxic microenvironments, therefore we hypothesized that hypoxia-induced signalling pathways could be involved in BCC and TE as they are in other human malignancies. Hypoxia-inducible factor 1 (HIF1) and mechanistic/mammalian target of rapamycin (mTOR) are key players in these pathways. Objectives To determine whether HIF1/mTOR signalling is involved in BCC and TE. Methods We used immunohistochemical staining of formalin-fixed paraffin-embedded BCC (n = 45) and TE (n = 35) samples to assess activity of HIF1, mTORC1 and their most important target genes. The percentage positive tumour cells was assessed manually in a semi-quantitative manner and categorized (0%, 80%). Results Among 45 BCC and 35 TE examined, expression levels were respectively 81% and 57% (BNIP3), 73% and 75% (CAIX), 79% and 86% (GLUT1), 50% and 19% (HIF1α), 89% and 88% (pAKT), 55% and 61% (pS6), 15% and 25% (pMTOR), 44% and 63% (PHD2) and 44% and 49% (VEGF-A). CAIX, Glut1 and PHD2 expression levels were significantly higher in TE when only samples with at least 80% expression were included. Conclusions HIF and mTORC1 signalling seems active in both BCC and TE. There are no appreciable differences between the two with respect to pathway activity. At this moment immunohistochemical analyses of HIF, mTORC1 and their target genes does not provide a reliable diagnostic tool for the discrimination of BCC and TE. PMID:25181405

  8. Immunohistochemical analysis of the mechanistic target of rapamycin and hypoxia signalling pathways in basal cell carcinoma and trichoepithelioma.

    Directory of Open Access Journals (Sweden)

    Tjinta Brinkhuizen

    Full Text Available BACKGROUND: Basal cell carcinoma (BCC is the most common cancer in Caucasians. Trichoepithelioma (TE is a benign neoplasm that strongly resembles BCC. Both are hair follicle (HF tumours. HFs are hypoxic microenvironments, therefore we hypothesized that hypoxia-induced signalling pathways could be involved in BCC and TE as they are in other human malignancies. Hypoxia-inducible factor 1 (HIF1 and mechanistic/mammalian target of rapamycin (mTOR are key players in these pathways. OBJECTIVES: To determine whether HIF1/mTOR signalling is involved in BCC and TE. METHODS: We used immunohistochemical staining of formalin-fixed paraffin-embedded BCC (n = 45 and TE (n = 35 samples to assess activity of HIF1, mTORC1 and their most important target genes. The percentage positive tumour cells was assessed manually in a semi-quantitative manner and categorized (0%, 80%. RESULTS: Among 45 BCC and 35 TE examined, expression levels were respectively 81% and 57% (BNIP3, 73% and 75% (CAIX, 79% and 86% (GLUT1, 50% and 19% (HIF1α, 89% and 88% (pAKT, 55% and 61% (pS6, 15% and 25% (pMTOR, 44% and 63% (PHD2 and 44% and 49% (VEGF-A. CAIX, Glut1 and PHD2 expression levels were significantly higher in TE when only samples with at least 80% expression were included. CONCLUSIONS: HIF and mTORC1 signalling seems active in both BCC and TE. There are no appreciable differences between the two with respect to pathway activity. At this moment immunohistochemical analyses of HIF, mTORC1 and their target genes does not provide a reliable diagnostic tool for the discrimination of BCC and TE.

  9. Pharmacological Targeting SHP-1-STAT3 Signaling Is a Promising Therapeutic Approach for the Treatment of Colorectal Cancer

    Directory of Open Access Journals (Sweden)

    Li-Ching Fan

    2015-09-01

    Full Text Available STAT3 activation is associated with poor prognosis in human colorectal cancer (CRC. Our previous data demonstrated that regorafenib (Stivarga is a pharmacological agonist of SH2 domain-containing phosphatase 1 (SHP-1 that enhances SHP-1 activity and induces apoptosis by targeting STAT3 signals in CRC. This study aimed to find a therapeutic drug that is more effective than regorafenib for CRC treatment. Here, we showed that SC-43 was more effective than regorafenib at inducing apoptosis in vitro and suppressing tumorigenesis in vivo. SC-43 significantly increased SHP-1 activity, downregulated p-STAT3Tyr705 level, and induced apoptosis in CRC cells. An SHP-1 inhibitor or knockdown of SHP-1 by siRNA both significantly rescued the SC-43–induced apoptosis and decreased p-STAT3Tyr705 level. Conversely, SHP-1 overexpression increased the effects of SC-43 on apoptosis and p-STAT3Tyr705 level. These data suggest that SC-43–induced apoptosis mediated through the loss of p-STAT3Tyr705 was dependent on SHP-1 function. Importantly, SC-43–enhanced SHP-1 activity was because of the docking potential of SC-43, which relieved the autoinhibited N-SH2 domain of SHP-1 and inhibited p-STAT3Tyr705 signals. Importantly, we observed that a significant negative correlation existed between SHP-1 and p-STAT3Tyr705expression in CRC patients (P = .038. Patients with strong SHP-1 and weak p-STAT3Tyr705 expression had significantly higher overall survival compared with patients with weak SHP-1 and strong p-STAT3Tyr705 expression (P = .029. In conclusion, SHP-1 is suitable to be a useful prognostic marker and a pharmacological target for CRC treatment. Targeting SHP-1-STAT3 signaling by SC-43 may serve as a promising pharmacotherapy for CRC.

  10. Optogenetic control of mitochondrial metabolism and Ca2+ signaling by mitochondria-targeted opsins

    OpenAIRE

    Tkatch, Tatiana; Greotti, Elisa; Baranauskas, Gytis; Pendin, Diana; Roy, Soumitra; Nita, Luliaoana I.; Wettmarshausen, Jennifer; Prigge, Matthias; Yizhar, Ofer; Shirihai, Orian S.; Fishman, Daniel; Hershfinkel, Michal; Fleidervish, Ilya A; Perocchi, Fabiana; Pozzan, Tullio

    2017-01-01

    Mitochondrial functions depend on the steep H+ electrochemical gradient (??H+) across their inner membrane. The available tools for controlling this gradient are essentially limited to inhibitors of the respiratory chain or of the H+ ATPase or to uncouplers, poisons plagued by important side effects and that lack both cell and spatial specificity. We show here that, by transfecting cells with the cDNA encoding channelrhodopsins specifically targeted to the inner mitochondrial membrane, we can...

  11. Hormonal Resistance And Metastasis ER-Coregulartor-Src Signaling Targeted Therapy

    Science.gov (United States)

    2011-09-01

    receptor family. Curr Opin Obstet Gynecol 1999 Jun;11(3):249-54. [16] Curtis HS, Couse JF, Korach KS. Estrogen receptor transcription and...breast cancer. Curr Top Med Chem 2006;6:195-216. (2) Lewis-Wambi JS, Jordan VC. Treatment of Postmenopausal Breast Cancer with Selective Estrogen...Estrogen receptors as therapeutic targets in breast cancer. Curr Top Med Chem 6:195–216 2. Lewis-Wambi JS, Jordan VC (2005) Treatment of postmeno- pausal

  12. Dysregulation of Mammalian Target of Rapamycin Signaling in Mouse Models of Autism

    OpenAIRE

    Huber, Kimberly M.; Klann, Eric; Costa-Mattioli, Mauro; Zukin, R. Suzanne

    2015-01-01

    The mammalian target of rapamycin (mTOR) is a central regulator of a diverse array of cellular processes, including cell growth, proliferation, autophagy, translation, and actin polymerization. Components of the mTOR cascade are present at synapses and influence synaptic plasticity and spine morphogenesis. A prevailing view is that the study of mTOR and its role in autism spectrum disorders (ASDs) will elucidate the molecular mechanisms by which mTOR regulates neuronal function under physiolo...

  13. Arabidopsis chromatin-associated HMGA and HMGB use different nuclear targeting signals and display highly dynamic localization within the nucleus

    DEFF Research Database (Denmark)

    Launholt, Dorte; Merkle, Thomas; Houben, Andreas

    2006-01-01

    HMGproteins appear to be involved in the regulation of transcription and other DNA-dependent processes. We have examined the subcellular localization of Arabidopsis thaliana HMGA, HMGB1, and HMGB5, revealing that they localize to the cell nucleus. They display a speckled distribution pattern throughout the chromatin...... of interphase nuclei, whereas none of the proteins associate with condensed mitotic chromosomes. HMGA is targeted to the nucleus by a monopartite nuclear localization signal, while efficient nuclear accumulation of HMGB1/5 requires large portions of the basic N-terminal part of the proteins. The acidic C......-terminal domain interferes with nucleolar targeting of HMGB1. Fluorescence recovery after photobleaching experiments revealed that HMGA and HMGB proteins are extremely dynamic in the nucleus, indicating that they bind chromatin only transiently before moving on to the next site, thereby continuously scanning...

  14. Targeting Glutamatergic Signaling for the Development of Novel Therapeutics for Mood Disorders

    Science.gov (United States)

    Machado-Vieira, R.; Salvadore, G.; Ibrahim, L.; DiazGranados, N.; Zarate, C.A.

    2009-01-01

    There have been no recent advances in drug development for mood disorders in terms of identifying drug targets that are mechanistically distinct from existing ones. As a result, existing antidepressants are based on decades-old notions of which targets are relevant to the mechanisms of antidepressant action. Low rates of remission, a delay of onset of therapeutic effects, continual residual depressive symptoms, relapses, and poor quality of life are unfortunately common in patients with mood disorders. Offering alternative options is requisite in order to reduce the individual and societal burden of these diseases. The glutamatergic system is a promising area of research in mood disorders, and likely to offer new possibilities in therapeutics. There is increasing evidence that mood disorders are associated with impairments in neuroplasticity and cellular resilience, and alterations of the glutamatergic system are known to play a major role in cellular plasticity and resilience. Existing antidepressants and mood stabilizers have prominent effects on the glutamate system, and modulating glutamatergic ionotropic or metabotropic receptors results in antidepressant-like properties in animal models. Several glutamatergic modulators targeting various glutamate components are currently being studied in the treatment of mood disorders, including release inhibitors of glutamate, N-methyl-D-aspartate (NMDA) antagonists, alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) throughput enhancers, and glutamate transporter enhancers. This paper reviews the currently available knowledge regarding the role of the glutamatergic system in the etiopathogenesis of mood disorders and putative glutamate modulators. PMID:19442176

  15. Targeting of the Hedgehog signal transduction pathway suppresses survival of malignant pleural mesothelioma cells in vitro.

    Science.gov (United States)

    You, Min; Varona-Santos, Javier; Singh, Samer; Robbins, David J; Savaraj, Niramol; Nguyen, Dao M

    2014-01-01

    The present study sought to determine whether the Hedgehog (Hh) pathway is active and regulates the cell growth of cultured malignant pleural mesothelioma (MPM) cells and to evaluate the efficacy of pathway blockade using smoothened (SMO) antagonists (SMO inhibitor GDC-0449 or the antifungal drug itraconazole [ITRA]) or Gli inhibitors (GANT61 or the antileukemia drug arsenic trioxide [ATO]) in suppressing MPM viability. Selective knockdown of SMO to inhibit Hh signaling was achieved by small interfering RNA in 3 representative MPM cells. The growth inhibitory effect of GDC-0449, ITRA, GANT61, and ATO was evaluated in 8 MPM lines, with cell viability quantified using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell death was determined by annexinV/propidium iodide staining and flow cytometry. SMO small interfering RNA mediated a two- to more than fivefold reduction of SMO and Gli1 gene expression as determined by real-time quantitative reverse-transcriptase polymerase chain reaction, indicating significant Hh pathway blockade. This was associated with significantly reduced cell viability (34% ± 7% to 61% ± 14% of nontarget small interfering RNA controls; P = .0024 to P = .043). Treating MPM cells with Hh inhibitors resulted in a 1.5- to 4-fold reduction of Gli1 expression. These 4 Hh antagonists strongly suppressed MPM cell viability. More importantly, ITRA, ATO, GANT61 induced significant apoptosis in the representative MPM cells. Hh signaling is active in MPM and regulates cell viability. ATO and ITRA were as effective as the prototypic SMO inhibitor GDC-0449 and the Gli inhibitor GANT61 in suppressing Hh signaling in MPM cells. Pharmaceutical agents Food and Drug Administration-approved for other indications but recently found to have anti-Hh activity, such as ATO or ITRA, could be repurposed to treat MPM. Copyright © 2014 The American Association for Thoracic Surgery. All rights reserved.

  16. Curcumin Rescues Diabetic Renal Fibrosis by Targeting Superoxide-Mediated Wnt Signaling Pathways.

    Science.gov (United States)

    Ho, Cheng; Hsu, Yung-Chien; Lei, Chen-Chou; Mau, Shu-Ching; Shih, Ya-Hsueh; Lin, Chun-Liang

    2016-03-01

    The purposes of this study were to investigate whether curcumin can weaken diabetic nephropathy by modulating both oxidative stress and renal injury from Wnt signaling mediation. Wnt5a/β-catenin depression and induction of superoxide synthesis are associated with high glucose (HG) induced transforming growth factor (TGF)-β1 and fibronectin expression in mesangial cells. Curcumin resumes HG depression of Wnt/β-catenin signaling and alleviates HG induction of superoxide, TGF-β1 and fibronectin expression in renal mesangial cell. Exogenous curcumin alleviated urinary total proteinuria and serum superoxide level in diabetic rats. Based on laser-captured microdissection for quantitative real-time polymerase chain reaction, it was found that diabetes significantly increased TGF-β1 and fibronectin expression in line with depressed Wnt5a expression. Curcumin treatment reduced the TGF-β1 and fibronectin activation and the inhibiting effect of diabetes on Wnt5a/β-catenin expression in renal glomeruli. Immunohistochemistry showed that curcumin treatment significantly reduced 8-hydroxy-2'-deoxyguanosine, TGF-β1 and fibronectin, and was in line with the restoration of the suppressed Wnt5a expression immunoreactivities in glomeruli of diabetic rats. Curcumin alleviated extracellular matrix accumulation in diabetic nephropathy by not only preventing the diabetes-mediated superoxide synthesis but also resuming downregulation of Wnt/β-catenin signaling. These findings suggest that regulation of Wnt activity by curcumin is a feasible alternative strategy to rescue diabetic renal injury. Copyright © 2016 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.

  17. The crucial role of protein phosphorylation in cell signaling and its use as targeted therapy (Review).

    Science.gov (United States)

    Ardito, Fatima; Giuliani, Michele; Perrone, Donatella; Troiano, Giuseppe; Lo Muzio, Lorenzo

    2017-08-01

    Protein phosphorylation is an impo-rtant cellular regulatory mechanism as many enzymes and receptors are activated/deactivated by phosphorylation and dephosphorylation events, by means of kinases and phosph-atases. In particular, the protein kinases are responsible for cellular transduction signaling and their hyperactivity, malfunction or overexpression can be found in several diseases, mostly tumors. Therefore, it is evident that the use of kinase inhibitors can be valuable for the treatment of cancer. In this review, we discuss the mechanism of action of phosphorylation, with particular attention to the importance of phosphorylation under physiological and pathological conditions. We also discuss the possibility of using kinase inhibitors in the treatment of tumors.

  18. Electrochemical detection of protein by using magnetic graphene-based target enrichment and copper nanoparticles-assisted signal amplification.

    Science.gov (United States)

    Zhao, Jing; Lv, Yun; Kang, Mingyang; Wang, Keming; Xiang, Yang

    2015-11-21

    In this paper, we propose a new method for protein detection by making use of magnetic graphene for enrichment and separation of the targets and duplex DNA-templated copper nanoparticles for amplification of electrochemical signals. Because the binding of the target protein (e.g. folate receptor) and small molecule (e.g. folate) can protect complementary DNA (cDNA) from exonuclease III-catalyzed degradation, duplex DNA from the hybridization of probe DNA and cDNA can act as the template for the formation of copper nanoparticles (CuNPs). Afterward, CuNPs-coated DNA can be enriched on the surface of magnetic graphene through the 3'-overhanging end of probe DNA, and then separated from the reaction mixture with the aid of magnet. As a result, copper ions released from acid-dissolution of CuNPs can catalyze the oxidation of o-phenylenediamine (OPD) by dissolved oxygen, resulting in an amplified electrochemical response. Our method can sensitively detect target protein over a wide linear range and with a low detection limit of 7.8 pg mL(-1), which can easily distinguish the targets even in complex serum samples. Therefore, this method may be promising for the clinical diagnosis of protein biomarkers by changing the recognition elements in the future.

  19. Hepatocyte TRAF3 promotes liver steatosis and systemic insulin resistance through targeting TAK1-dependent signalling.

    Science.gov (United States)

    Wang, Pi-Xiao; Zhang, Xiao-Jing; Luo, Pengcheng; Jiang, Xi; Zhang, Peng; Guo, Junhong; Zhao, Guang-Nian; Zhu, Xueyong; Zhang, Yan; Yang, Sijun; Li, Hongliang

    2016-02-17

    Non-alcoholic fatty liver disease (NAFLD) is characterized by hepatic steatosis, insulin resistance and a systemic pro-inflammatory response. Here we show that tumour necrosis factor receptor-associated factor 3 (TRAF3) is upregulated in mouse and human livers with hepatic steatosis. After 24 weeks on a high-fat diet (HFD), obesity, insulin resistance, hepatic steatosis and inflammatory responses are significantly ameliorated in liver-specific TRAF3-knockout mice, but exacerbated in transgenic mice overexpressing TRAF3 in hepatocytes. The detrimental effects of TRAF3 on hepatic steatosis and related pathologies are confirmed in ob/ob mice. We further show that in response to HFD, hepatocyte TRAF3 binds to TGF-β-activated kinase 1 (TAK1) to induce TAK1 ubiquitination and subsequent autophosphorylation, thereby enhancing the activation of downstream IKKβ-NF-κB and MKK-JNK-IRS1(307) signalling cascades, while disrupting AKT-GSK3β/FOXO1 signalling. The TRAF3-TAK1 interaction and TAK1 ubiquitination are indispensable for TRAF3-regulated hepatic steatosis. In conclusion, hepatocyte TRAF3 promotes HFD-induced or genetic hepatic steatosis in a TAK1-dependent manner.

  20. Hydrogen Sulfide Signaling Axis as a Target for Prostate Cancer Therapeutics

    Directory of Open Access Journals (Sweden)

    Mingzhe Liu

    2016-01-01

    Full Text Available Hydrogen sulfide (H2S was originally considered toxic at elevated levels; however just in the past decade H2S has been proposed to be an important gasotransmitter with various physiological and pathophysiological roles in the body. H2S can be generated endogenously from L-cysteine by multiple enzymes, including cystathionine gamma-lyase, cystathionine beta-synthase, and 3-mercaptopyruvate sulfurtransferase in combination with cysteine aminotransferase. Prostate cancer is a major health concern and no effective treatment for prostate cancers is available. H2S has been shown to inhibit cell survival of androgen-independent, androgen-dependent, and antiandrogen-resistant prostate cancer cells through different mechanisms. Various H2S-releasing compounds, including sulfide salts, diallyl disulfide, diallyl trisulfide, sulforaphane, and other polysulfides, also have been shown to inhibit prostate cancer growth and metastasis. The expression of H2S-producing enzyme was reduced in both human prostate cancer tissues and prostate cancer cells. Androgen receptor (AR signaling is indispensable for the development of castration resistant prostate cancer, and H2S was shown to inhibit AR transactivation and contributes to antiandrogen-resistant status. In this review, we summarized the current knowledge of H2S signaling in prostate cancer and described the molecular alterations, which may bring this gasotransmitter into the clinic in the near future for developing novel pharmacological and therapeutic interventions for prostate cancer.

  1. Peptidyl-prolyl isomerization targets rice Aux/IAAs for proteasomal degradation during auxin signalling.

    Science.gov (United States)

    Jing, Hongwei; Yang, Xiaolu; Zhang, Jian; Liu, Xuehui; Zheng, Huakun; Dong, Guojun; Nian, Jinqiang; Feng, Jian; Xia, Bin; Qian, Qian; Li, Jiayang; Zuo, Jianru

    2015-06-22

    In plants, auxin signalling is initiated by the auxin-promoted interaction between the auxin receptor TIR1, an E3 ubiquitin ligase, and the Aux/IAA transcriptional repressors, which are subsequently degraded by the proteasome. Gain-of-function mutations in the highly conserved domain II of Aux/IAAs abolish the TIR1-Aux/IAA interaction and thus cause an auxin-resistant phenotype. Here we show that peptidyl-prolyl isomerization of rice OsIAA11 catalysed by LATERAL ROOTLESS2 (LRT2), a cyclophilin-type peptidyl-prolyl cis/trans isomerase, directly regulates the stability of OsIAA11. NMR spectroscopy reveals that LRT2 efficiently catalyses the cis/trans isomerization of OsIAA11. The lrt2 mutation reduces OsTIR1-OsIAA11 interaction and consequently causes the accumulation of a higher level of OsIAA11 protein. Moreover, knockdown of the OsIAA11 expression partially rescues the lrt2 mutant phenotype in lateral root development. Together, these results illustrate cyclophilin-catalysed peptidyl-prolyl isomerization promotes Aux/IAA degradation, as a mechanism regulating auxin signalling.

  2. Honokiol suppresses metastasis of renal cell carcinoma by targeting KISS1/KISS1R signaling.

    Science.gov (United States)

    Cheng, Shujie; Castillo, Victor; Eliaz, Isaac; Sliva, Daniel

    2015-01-01

    Renal cell carcinoma (RCC) is a common urological cancer worldwide and is known to have a high risk of metastasis, which is considered responsible for more than 90% of cancer associated deaths. Honokiol is a small-molecule biphenol isolated from Magnolia spp. bark and has been shown to be a potential anticancer agent involved in multiple facets of signal transduction. In this study, we demonstrated that honokiol inhibited the invasion and colony formation of highly metastatic RCC cell line 786-0 in a dose-dependent manner. DNA-microarray data showed the significant upregulation of metastasis-suppressor gene KISS1 and its receptor, KISS1R. The upregulation was confirmed by qRT-PCR analysis. Overexpression of KISS1 and KISS1R was detected by western blotting at the translation level as well. Of note, the decreased invasive and colonized capacities were reversed by KISS1 knockdown. Taken together, the results first indicate that activation of KISS1/KISS1R signaling by honokiol suppresses multistep process of metastasis, including invasion and colony formation, in RCC cells 786-0. Honokiol may be considered as a natural agent against RCC metastasis.

  3. Targeting AGEs Signaling Ameliorates Central Nervous System Diabetic Complications in Rats

    Directory of Open Access Journals (Sweden)

    Mohamed Naguib Zakaria

    2015-01-01

    Full Text Available Diabetes is a chronic endocrine disorder associated with several complications as hypertension, advanced brain aging, and cognitive decline. Accumulation of advanced glycation end products (AGEs is an important mechanism that mediates diabetic complications. Upon binding to their receptor (RAGE, AGEs mediate oxidative stress and/or cause cross-linking with proteins in blood vessels and brain tissues. The current investigation was designed to investigate the effect of agents that decrease AGEs signaling, perindopril which increases soluble RAGE (sRAGE and alagebrium which cleaves AGEs cross-links, compared to the standard antidiabetic drug, gliclazide, on the vascular and central nervous system (CNS complications in STZ-induced (50 mg/kg, IP diabetes in rats. Perindopril ameliorated the elevation in blood pressure seen in diabetic animals. In addition, both perindopril and alagebrium significantly inhibited memory decline (performance in the Y-maze, neuronal degeneration (Fluoro-Jade staining, AGEs accumulation in serum and brain, and brain oxidative stress (level of reduced glutathione and activities of catalase and malondialdehyde. These results suggest that blockade of AGEs signaling after diabetes induction in rats is effective in reducing diabetic CNS complications.

  4. A p53-inducible microRNA-34a downregulates Ras signaling by targeting IMPDH

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Hwa-Ryeon; Roe, Jae-Seok; Lee, Ji-Eun; Hwang, In-Young [National Research Laboratory for Metabolic Checkpoint, Departments of Biomedical Sciences and Biochemistry and Molecular Biology, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); Cho, Eun-Jung [National Research Laboratory for Chromatin Dynamics, College of Pharmacy, Sungkyunkwan University, Suwon (Korea, Republic of); Youn, Hong-Duk, E-mail: hdyoun@snu.ac.kr [National Research Laboratory for Metabolic Checkpoint, Departments of Biomedical Sciences and Biochemistry and Molecular Biology, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); WCU Department of Molecular Medicine and Biopharmaceutical Sciences, Graduate School of Convergence Science, Seoul National University, Seoul (Korea, Republic of)

    2012-02-24

    Highlights: Black-Right-Pointing-Pointer p53 downregulates IMPDH. Black-Right-Pointing-Pointer p53-dependent miR-34a transactivation inhibits IMPDH transcription. Black-Right-Pointing-Pointer miR-34a-mediated inhibition of IMPDH downregulates GTP-dependent Ras signal. -- Abstract: p53 is a well-known transcription factor that controls cell cycle arrest and cell death in response to a wide range of stresses. Moreover, p53 regulates glucose metabolism and its mutation results in the metabolic switch to the Warburg effect found in cancer cells. Nucleotide biosynthesis is also critical for cell proliferation and the cell division cycle. Nonetheless, little is known about whether p53 regulates nucleotide biosynthesis. Here we demonstrated that p53-inducible microRNA-34a (miR-34a) repressed inosine 5 Prime -monophosphate dehydrogenase (IMPDH), a rate-limiting enzyme of de novo GTP biosynthesis. Treatment with anti-miR-34a inhibitor relieved the expression of IMPDH upon DNA damage. Ultimately, miR-34a-mediated inhibition of IMPDH resulted in repressed activation of the GTP-dependent Ras signaling pathway. In summary, we suggest that p53 has a novel function in regulating purine biosynthesis, aided by miR-34a-dependent IMPDH repression.

  5. Targetable signaling pathway mutations are associated with malignant phenotype in IDH-mutant gliomas.

    Science.gov (United States)

    Wakimoto, Hiroaki; Tanaka, Shota; Curry, William T; Loebel, Franziska; Zhao, Dan; Tateishi, Kensuke; Chen, Juxiang; Klofas, Lindsay K; Lelic, Nina; Kim, James C; Dias-Santagata, Dora; Ellisen, Leif W; Borger, Darrell R; Fendt, Sarah-Maria; Vander Heiden, Matthew G; Batchelor, Tracy T; Iafrate, A John; Cahill, Daniel P; Chi, Andrew S

    2014-06-01

    Isocitrate dehydrogenase (IDH) gene mutations occur in low-grade and high-grade gliomas. We sought to identify the genetic basis of malignant phenotype heterogeneity in IDH-mutant gliomas. We prospectively implanted tumor specimens from 20 consecutive IDH1-mutant glioma resections into mouse brains and genotyped all resection specimens using a CLIA-certified molecular panel. Gliomas with cancer driver mutations were tested for sensitivity to targeted inhibitors in vitro. Associations between genomic alterations and outcomes were analyzed in patients. By 10 months, 8 of 20 IDH1-mutant gliomas developed intracerebral xenografts. All xenografts maintained mutant IDH1 and high levels of 2-hydroxyglutarate on serial transplantation. All xenograft-producing gliomas harbored "lineage-defining" mutations in CIC (oligodendroglioma) or TP53 (astrocytoma), and 6 of 8 additionally had activating mutations in PIK3CA or amplification of PDGFRA, MET, or N-MYC. Only IDH1 and CIC/TP53 mutations were detected in non-xenograft-forming gliomas (P = 0.0007). Targeted inhibition of the additional alterations decreased proliferation in vitro. Moreover, we detected alterations in known cancer driver genes in 13.4% of IDH-mutant glioma patients, including PIK3CA, KRAS, AKT, or PTEN mutation or PDGFRA, MET, or N-MYC amplification. IDH/CIC mutant tumors were associated with PIK3CA/KRAS mutations whereas IDH/TP53 tumors correlated with PDGFRA/MET amplification. Presence of driver alterations at progression was associated with shorter subsequent progression-free survival (median 9.0 vs. 36.1 months; P = 0.0011). A subset of IDH-mutant gliomas with mutations in driver oncogenes has a more malignant phenotype in patients. Identification of these alterations may provide an opportunity for use of targeted therapies in these patients. Clin Cancer Res; 20(11); 2898-909. ©2014 AACR. ©2014 American Association for Cancer Research.

  6. Targeting Ligand Dependent and Ligand Independent Androgen Receptor Signaling in Prostate Cancer

    Science.gov (United States)

    2014-10-01

    Muller-Brand J, Forrer F. Targeted radio - therapy with radiolabeled somatostatin analogs. Endocrinol Metab Clin North Am 2011;40:187–204. 55...carcinogenesis1. Androgen deprivation therapy is the mainstay in treatment of advanced PCa (PCa); however, after an initial response, the disease...R1 O NO2 O R2 Scheme 1 Syntheses of bis-benzamide libraries. Reagents and conditions: (a) (COCl)2, cat . DMF, CH2Cl2, reflux, 1 h; (b) DIEA, CH2Cl2

  7. [Targeting of PP2A enzymes by viral proteins and cancer signalling].

    Science.gov (United States)

    Godet, Angélique N; Guergnon, Julien; Galioot, Amandine; Falanga, Pierre-Barthélémy; Colle, Jean-Hervé; Cayla, Xavier; Garcia, Alphonse

    2011-12-01

    Protein phosphatase 2A (PP2A) is a large family of holoenzymes that comprises 1% of total cellular proteins and accounts for the majority of Ser/Thr phosphatase activity in eukaryotic cells. PP2A proteins are made of a core dimer, composed of a catalytic (C) subunit and a structural (A) subunit, in association with a third variable -regulatory (B) subunit. Although initially considered as a constitutive housekeeping enzyme, PP2A is indeed highly regulated by post-translational modifications of its catalytic subunit or by the identity of a regulatory type B subunit, which determines substrate specificity, subcellular localization and enzymatic activity of a defined holoenzyme. During the two last decades, multiple studies of structural and functional regulation of PP2A holoenzymes by viral proteins led to the identification of critical pathways for both viral biology and tumorigenesis. To date a dozen of different viruses (ADN/ARN or retrovirus) have been identified that encode viral proteins associated to PP2A. In this review, we analyze a biological strategy, used by various viruses based on the targeting of PP2A enzymes by viral proteins, in order to specifically deregulate cellular pathways of their hosts. The impact of such PP2A targeting for biomedical search, and in further therapeutic developments against cancer, will also be discussed. © 2011 médecine/sciences – Inserm / SRMS.

  8. Pathogenic bacteria target NEDD8-conjugated cullins to hijack host-cell signaling pathways.

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    Grégory Jubelin

    2010-09-01

    Full Text Available The cycle inhibiting factors (Cif, produced by pathogenic bacteria isolated from vertebrates and invertebrates, belong to a family of molecules called cyclomodulins that interfere with the eukaryotic cell cycle. Cif blocks the cell cycle at both the G₁/S and G₂/M transitions by inducing the stabilization of cyclin-dependent kinase inhibitors p21(waf1 and p27(kip1. Using yeast two-hybrid screens, we identified the ubiquitin-like protein NEDD8 as a target of Cif. Cif co-compartmentalized with NEDD8 in the host cell nucleus and induced accumulation of NEDD8-conjugated cullins. This accumulation occurred early after cell infection and correlated with that of p21 and p27. Co-immunoprecipitation revealed that Cif interacted with cullin-RING ubiquitin ligase complexes (CRLs through binding with the neddylated forms of cullins 1, 2, 3, 4A and 4B subunits of CRL. Using an in vitro ubiquitylation assay, we demonstrate that Cif directly inhibits the neddylated CUL1-associated ubiquitin ligase activity. Consistent with this inhibition and the interaction of Cif with several neddylated cullins, we further observed that Cif modulates the cellular half-lives of various CRL targets, which might contribute to the pathogenic potential of diverse bacteria.

  9. Pathogenic bacteria target NEDD8-conjugated cullins to hijack host-cell signaling pathways.

    Science.gov (United States)

    Jubelin, Grégory; Taieb, Frédéric; Duda, David M; Hsu, Yun; Samba-Louaka, Ascel; Nobe, Rika; Penary, Marie; Watrin, Claude; Nougayrède, Jean-Philippe; Schulman, Brenda A; Stebbins, C Erec; Oswald, Eric

    2010-09-30

    The cycle inhibiting factors (Cif), produced by pathogenic bacteria isolated from vertebrates and invertebrates, belong to a family of molecules called cyclomodulins that interfere with the eukaryotic cell cycle. Cif blocks the cell cycle at both the G₁/S and G₂/M transitions by inducing the stabilization of cyclin-dependent kinase inhibitors p21(waf1) and p27(kip1). Using yeast two-hybrid screens, we identified the ubiquitin-like protein NEDD8 as a target of Cif. Cif co-compartmentalized with NEDD8 in the host cell nucleus and induced accumulation of NEDD8-conjugated cullins. This accumulation occurred early after cell infection and correlated with that of p21 and p27. Co-immunoprecipitation revealed that Cif interacted with cullin-RING ubiquitin ligase complexes (CRLs) through binding with the neddylated forms of cullins 1, 2, 3, 4A and 4B subunits of CRL. Using an in vitro ubiquitylation assay, we demonstrate that Cif directly inhibits the neddylated CUL1-associated ubiquitin ligase activity. Consistent with this inhibition and the interaction of Cif with several neddylated cullins, we further observed that Cif modulates the cellular half-lives of various CRL targets, which might contribute to the pathogenic potential of diverse bacteria.

  10. The Peroxisomal Targeting Signal 1 in sterol carrier protein 2 is autonomous and essential for receptor recognition

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    Bond Charles S

    2011-03-01

    Full Text Available Abstract Background The majority of peroxisomal matrix proteins destined for translocation into the peroxisomal lumen are recognised via a C-terminal Peroxisomal Target Signal type 1 by the cycling receptor Pex5p. The only structure to date of Pex5p in complex with a cargo protein is that of the C-terminal cargo-binding domain of the receptor with sterol carrier protein 2, a small, model peroxisomal protein. In this study, we have tested the contribution of a second, ancillary receptor-cargo binding site, which was found in addition to the characterised Peroxisomal Target Signal type 1. Results To investigate the function of this secondary interface we have mutated two key residues from the ancillary binding site and analyzed the level of binding first by a yeast-two-hybrid assay, followed by quantitative measurement of the binding affinity and kinetics of purified protein components and finally, by in vivo measurements, to determine translocation capability. While a moderate but significant reduction of the interaction was found in binding assays, we were not able to measure any significant defects in vivo. Conclusions Our data therefore suggest that at least in the case of sterol carrier protein 2 the contribution of the second binding site is not essential for peroxisomal import. At this stage, however, we cannot rule out that other cargo proteins may require this ancillary binding site.

  11. Targeting TGF-β Signaling by Antisense Oligonucleotide-mediated Knockdown of TGF-β Type I Receptor

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    Dwi U Kemaladewi

    2014-01-01

    Full Text Available Duchenne muscular dystrophy (DMD is caused by lack of functional dystrophin and results in progressive myofiber damage and degeneration. In addition, impaired muscle regeneration and fibrosis contribute to the progressive pathology of DMD. Importantly, transforming growth factor-β (TGF-β is implicated in DMD pathology and is known to stimulate fibrosis and inhibit muscle regeneration. In this study, we present a new strategy to target TGF-β signaling cascades by specifically inhibiting the expression of TGF-β type I receptor TGFBR1 (ALK5. Antisense oligonucleotides (AONs were designed to specifically induce exon skipping of mouse ALK5 transcripts. AON-induced exon skipping of ALK5 resulted in specific downregulation of full-length receptor transcripts in vitro in different cell types, repression of TGF-β activity, and enhanced C2C12 myoblast differentiation. To determine the effect of these AONs in dystrophic muscles, we performed intramuscular injections of ALK5 AONs in mdx mice, which resulted in a decrease in expression of fibrosis-related genes and upregulation of Myog expression compared to control AON-injected muscles. In summary, our study presents a novel method to target TGF-β signaling cascades with potential beneficial effects for DMD.

  12. Targeting multiple pro-apoptotic signaling pathways with curcumin in prostate cancer cells.

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    Mariela Rivera

    Full Text Available Curcumin, an extract from the turmeric rhizome (Curcuma longa, is known to exhibit anti-inflammatory, antioxidant, chemopreventive and antitumoral activities against aggressive and recurrent cancers. Accumulative data indicate that curcumin may induce cancer cell death. However, the detailed mechanism underlying its pro-apoptotic and anti-cancer effects remains to be elucidated. In the present study, we examined the signaling pathways triggered by curcumin, specifically, the exact molecular mechanisms of curcumin-induced apoptosis in highly metastatic human prostate cancer cells. The effect of curcumin was evaluated using for the first time in prostate cancer, a gel-free shotgun quantitative proteomic analysis coupled with Tandem Mass Tag isobaric labeling-based-signaling networks. Results were confirmed at the gene expression level by qRT-PCR and at the protein expression level by western blot and flow cytometry. Our findings revealed that curcumin induced an Endoplasmic Reticulum stress-mediated apoptosis in PC3. The mechanisms by which curcumin promoted cell death in these cells were associated with cell cycle arrest, increased reactive oxygen species, autophagy and the Unfolded Protein Response. Furthermore, the upregulation of ER stress was measured using key indicators of ER stress: Glucose-Regulated Protein 78, Inositol-Requiring Enzyme 1 alpha, Protein Disulfide isomerase and Calreticulin. Chronic ER stress induction was concomitant with the upregulation of pro-apoptotic markers (caspases 3,9,12 and Poly (ADP-ribose polymerase. The downregulated proteins include anti-apoptotic and anti-tumor markers, supporting their curcumin-induced pro-apoptotic role in prostate cancer cells. Taken together, these data suggest that curcumin may serve as a promising anticancer agent by inducing a chronic ER stress mediated cell death and activation of cell cycle arrest, UPR, autophagy and oxidative stress responses.

  13. Targeting p53 via JNK pathway: a novel role of RITA for apoptotic signaling in multiple myeloma.

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    Manujendra N Saha

    Full Text Available The low frequency of p53 alterations e.g., mutations/deletions (∼10% in multiple myeloma (MM makes this tumor type an ideal candidate for p53-targeted therapies. RITA is a small molecule which can induce apoptosis in tumor cells by activating the p53 pathway. We previously showed that RITA strongly activates p53 while selectively inhibiting growth of MM cells without inducing genotoxicity, indicating its potential as a drug lead for p53-targeted therapy in MM. However, the molecular mechanisms underlying the pro-apoptotic effect of RITA are largely undefined. Gene expression analysis by microarray identified a significant number of differentially expressed genes associated with stress response including c-Jun N-terminal kinase (JNK signaling pathway. By Western blot analysis we further confirmed that RITA induced activation of p53 in conjunction with up-regulation of phosphorylated ASK-1, MKK-4 and c-Jun. These results suggest that RITA induced the activation of JNK signaling. Chromatin immunoprecipitation (ChIP analysis showed that activated c-Jun binds to the activator protein-1 (AP-1 binding site of the p53 promoter region. Disruption of the JNK signal pathway by small interfering RNA (siRNA against JNK or JNK specific inhibitor, SP-600125 inhibited the activation of p53 and attenuated apoptosis induced by RITA in myeloma cells carrying wild type p53. On the other hand, p53 transcriptional inhibitor, PFT-α or p53 siRNA not only inhibited the activation of p53 transcriptional targets but also blocked the activation of c-Jun suggesting the presence of a positive feedback loop between p53 and JNK. In addition, RITA in combination with dexamethasone, known as a JNK activator, displays synergistic cytotoxic responses in MM cell lines and patient samples. Our study unveils a previously undescribed mechanism of RITA-induced p53-mediated apoptosis through JNK signaling pathway and provides the rationale for combination of p53 activating drugs with

  14. A Cardiac-enriched MicroRNA, miR-378, Blocks Cardiac Hypertrophy by Targeting Ras Signaling*

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    Nagalingam, Raghu S.; Sundaresan, Nagalingam R.; Gupta, Mahesh P.; Geenen, David L.; Solaro, R. John; Gupta, Madhu

    2013-01-01

    Understanding the regulation of cardiomyocyte growth is crucial for the management of adverse ventricular remodeling and heart failure. MicroRNA-378 (miR-378) is a newly described member of the cardiac-enriched miRNAs, which is expressed only in cardiac myocytes and not in cardiac fibroblasts. We have previously shown that miR-378 regulates cardiac growth during the postnatal period by direct targeting of IGF1R (Knezevic, I., Patel, A., Sundaresan, N. R., Gupta, M. P., Solaro, R. J., Nagalingam, R. S., and Gupta, M. (2012) J. Biol. Chem. 287, 12913–12926). Here, we report that miR-378 is an endogenous negative regulator of cardiac hypertrophy, and its levels are down-regulated during hypertrophic growth of the heart and during heart failure. In primary cultures of cardiomyocytes, overexpression of miR-378 blocked phenylephrine (PE)-stimulated Ras activity and also prevented activation of two major growth-promoting signaling pathways, PI3K-AKT and Raf1-MEK1-ERK1/2, acting downstream of Ras signaling. Overexpression of miR-378 suppressed PE-induced phosphorylation of S6 ribosomal kinase, pERK1/2, pAKT, pGSK-3β, and nuclear accumulation of NFAT. There was also suppression of the fetal gene program that was induced by PE. Experiments carried out to delineate the mechanism behind the suppression of Ras, led us to identify Grb2, an upstream component of Ras signaling, as a bona fide direct target of miR-378-mediated regulation. Deficiency of miR-378 alone was sufficient to induce fetal gene expression, which was prevented by knocking down Grb2 expression and blocking Ras activation, thus suggesting that miR-378 interferes with Ras activation by targeting Grb2. Our study demonstrates that miR-378 is an endogenous negative regulator of Ras signaling and cardiac hypertrophy and its deficiency contributes to the development of cardiac hypertrophy. PMID:23447532

  15. KSHV Entry and Trafficking in Target Cells—Hijacking of Cell Signal Pathways, Actin and Membrane Dynamics

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    Binod Kumar

    2016-11-01

    Full Text Available Kaposi’s sarcoma associated herpesvirus (KSHV is etiologically associated with human endothelial cell hyperplastic Kaposi’s sarcoma and B-cell primary effusion lymphoma. KSHV infection of adherent endothelial and fibroblast cells are used as in vitro models for infection and KSHV enters these cells by host membrane bleb and actin mediated macropinocytosis or clathrin endocytosis pathways, respectively. Infection in endothelial and fibroblast cells is initiated by the interactions between multiple viral envelope glycoproteins and cell surface associated heparan sulfate (HS, integrins (α3β1, αVβ3 and αVβ5, and EphA2 receptor tyrosine kinase (EphA2R. This review summarizes the accumulated studies demonstrating that KSHV manipulates the host signal pathways to enter and traffic in the cytoplasm of the target cells, to deliver the viral genome into the nucleus, and initiate viral gene expression. KSHV interactions with the cell surface receptors is the key platform for the manipulations of host signal pathways which results in the simultaneous induction of FAK, Src, PI3-K, Rho-GTPase, ROS, Dia-2, PKC ζ, c-Cbl, CIB1, Crk, p130Cas and GEF-C3G signal and adaptor molecules that play critical roles in the modulation of membrane and actin dynamics, and in the various steps of the early stages of infection such as entry and trafficking towards the nucleus. The Endosomal Sorting Complexes Required for Transport (ESCRT proteins are also recruited to assist in viral entry and trafficking. In addition, KSHV interactions with the cell surface receptors also induces the host transcription factors NF-κB, ERK1/2, and Nrf2 early during infection to initiate and modulate viral and host gene expression. Nuclear delivery of the viral dsDNA genome is immediately followed by the host innate responses such as the DNA damage response (DDR, inflammasome and interferon responses. Overall, these studies form the initial framework for further studies of

  16. Phosphatidylinositol-3 kinase-Akt-mammalian target of rapamycin signaling pathway mediates contractility of human endometriotic stromal cells: A promising new target for the treatment of endometriosis-associated fibrosis

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    Wakana Abe

    2014-11-01

    Conclusion: The current findings suggest that the PI3K-Akt-mTOR signaling pathway is involved in the development of endometriosis-associated fibrosis. The PI3K-Akt-mTOR signaling pathway is a promising target for the treatment of endometriosis.

  17. Targeting the Oncogenic Transcriptional Regulator MYB in Adenoid Cystic Carcinoma by Inhibition of IGF1R/AKT Signaling.

    Science.gov (United States)

    Andersson, Mattias K; Afshari, Maryam K; Andrén, Ywonne; Wick, Michael J; Stenman, Göran

    2017-09-01

    Adenoid cystic carcinoma (ACC) is an aggressive cancer with no curative treatment for patients with recurrent/metastatic disease. The MYB-NFIB gene fusion is the main genomic hallmark and a potential therapeutic target. Oncogenic signaling pathways were studied in cultured cells and/or tumors from 15 ACC patients. Phospho-receptor tyrosine kinase (RTK) arrays were used to study the activity of RTKs. Effects of RTK inhibition on cell proliferation were analyzed with AlamarBlue, sphere assays, and two ACC xenograft models (n = 4-9 mice per group). The molecular effects of MYB-NFIB knockdown and IGF1R inhibition were studied with quantitative polymerase chain reaction, immunoblot, and gene expression microarrays. All statistical tests were two-sided. The MYB-NFIB fusion drives proliferation of ACC cells and is crucial for spherogenesis. Intriguingly, the fusion is regulated through AKT-dependent signaling induced by IGF1R overexpression and is downregulated upon IGF1R-inhibition (% expression of control ± SD = 27.2 ± 1.3, P MYB-NFIB regulates genes involved in cell cycle control, DNA replication/repair, and RNA processing. The transcriptional program induced by MYB-NFIB affects critical oncogenic mediators normally controlled by MYC and is reversed by pharmacological inhibition of IGF1R. Co-activation of epidermal growth factor receptor (EGFR) and MET promoted proliferation of ACC cells, and combined targeting of IGFR1/EGFR/MET induced differentiation and synergistically inhibited the growth of patient-derived xenografted ACCs (ACCX5M1, % growth of control ± SD = 34.9 ± 20.3, P = .006; ACCX6, % growth of control ± SD = 24.1 ± 17.5, P = .04). MYB-NFIB is an oncogenic driver and a key therapeutic target in ACC that is regulated by AKT-dependent IGF1R signaling. Our studies uncover a new strategy to target an oncogenic transcriptional master regulator and provide new important insights into the biology and treatment of ACC.

  18. Atrial natriuretic peptide/natriuretic peptide receptor A (ANP/NPRA) signaling pathway: a potential therapeutic target for allergic asthma.

    Science.gov (United States)

    Ma, Libing; Xiang, Xudong

    2011-11-01

    Allergic asthma is a chronic inflammatory disease of airway and immune disorder is an acknowledged mechanism. Numerous data demonstrate Th1/Th2 cells play an important role in the development of allergic asthma. Atrial natriuretic peptide (ANP) is a multifunctional hormone secreted by cardiac atria, lung, and so forth, which has been recognized for several decades due to its general effects on cardiovascular system, and natriuretic peptide receptor A (NPRA) is the major effecting receptor for ANP. In recent years, more and more studies suggest that ANP/NPRA signaling pathway is implicated in modulation of immnue and inflammatory reaction. Moreover, there are some reports about significant changes of ANP production in peripheral blood from asthmatics in acute exacerbation compared with patients during the remission and the healthy. Nevertheless, it is unknown that why ANP shows an observable change and what role ANP plays in asthma until now. We propose that ANP/NPRA signaling pathway is involved in immune dysfunction and airway inflammation of allergic asthma based on our experimental results, which suggests ANP/NPRA signaling pathway may be a potential therapeutic target for allergic asthma. Copyright © 2011 Elsevier Ltd. All rights reserved.

  19. R-2HG Exhibits Anti-tumor Activity by Targeting FTO/m6A/MYC/CEBPA Signaling.

    Science.gov (United States)

    Su, Rui; Dong, Lei; Li, Chenying; Nachtergaele, Sigrid; Wunderlich, Mark; Qing, Ying; Deng, Xiaolan; Wang, Yungui; Weng, Xiaocheng; Hu, Chao; Yu, Mengxia; Skibbe, Jennifer; Dai, Qing; Zou, Dongling; Wu, Tong; Yu, Kangkang; Weng, Hengyou; Huang, Huilin; Ferchen, Kyle; Qin, Xi; Zhang, Bin; Qi, Jun; Sasaki, Atsuo T; Plas, David R; Bradner, James E; Wei, Minjie; Marcucci, Guido; Jiang, Xi; Mulloy, James C; Jin, Jie; He, Chuan; Chen, Jianjun

    2018-01-11

    R-2-hydroxyglutarate (R-2HG), produced at high levels by mutant isocitrate dehydrogenase 1/2 (IDH1/2) enzymes, was reported as an oncometabolite. We show here that R-2HG also exerts a broad anti-leukemic activity in vitro and in vivo by inhibiting leukemia cell proliferation/viability and by promoting cell-cycle arrest and apoptosis. Mechanistically, R-2HG inhibits fat mass and obesity-associated protein (FTO) activity, thereby increasing global N6-methyladenosine (m6A) RNA modification in R-2HG-sensitive leukemia cells, which in turn decreases the stability of MYC/CEBPA transcripts, leading to the suppression of relevant pathways. Ectopically expressed mutant IDH1 and S-2HG recapitulate the effects of R-2HG. High levels of FTO sensitize leukemic cells to R-2HG, whereas hyperactivation of MYC signaling confers resistance that can be reversed by the inhibition of MYC signaling. R-2HG also displays anti-tumor activity in glioma. Collectively, while R-2HG accumulated in IDH1/2 mutant cancers contributes to cancer initiation, our work demonstrates anti-tumor effects of 2HG in inhibiting proliferation/survival of FTO-high cancer cells via targeting FTO/m6A/MYC/CEBPA signaling. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Grape polyphenols inhibit Akt/mammalian target of rapamycin signaling and potentiate the effects of gefitinib in breast cancer.

    Science.gov (United States)

    Castillo-Pichardo, Linette; Dharmawardhane, Suranganie F

    2012-01-01

    We recently reported that a combination of dietary grape polyphenols resveratrol, quercetin, and catechin (RQC), at low concentrations, was effective at inhibiting metastatic cancer progression. Herein, we investigate the molecular mechanisms of RQC in breast cancer and explore the potential of RQC as a potentiation agent for the epidermal growth factor receptor (EGFR) therapeutic gefitinib. Our in vitro experiments showed RQC induced apoptosis in gefitinib-resistant breast cancer cells via regulation of a myriad of proapoptotic proteins. Because the Akt/mammalian target of rapamycin (mTOR) signaling pathway is often elevated during development of anti-EGFR therapy resistance, the effect of RQC on the mTOR upstream effector Akt and the negative regulator AMP kinase (AMPK) was investigated. RQC was found to reduce Akt activity, induce the activation of AMPK, and inhibit mTOR signaling in breast cancer cells. Combined RQC and gefitinib decreased gefitinib resistant breast cancer cell viability to a greater extent than RQC or gefitinib alone. Moreover, RQC inhibited Akt and mTOR and activated AMPK even in the presence of gefitinib. Our in vivo experiments showed combined RQC and gefitinib was more effective than the individual treatments at inhibiting mammary tumor growth and metastasis in nude mice. Therefore, RQC treatment inhibits breast cancer progression and may potentiate anti-EGFR therapy by inhibition of Akt/mTOR signaling.

  1. Sonic hedgehog signaling inhibition provides opportunities for targeted therapy by sulforaphane in regulating pancreatic cancer stem cell self-renewal.

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    Mariana Rodova

    Full Text Available Dysregulation of the sonic hedgehog (Shh signaling pathway has been associated with cancer stem cells (CSC and implicated in the initiation of pancreatic cancer. Pancreatic CSCs are rare tumor cells characterized by their ability to self-renew, and are responsible for tumor recurrence accompanied by resistance to current therapies. The lethality of these incurable, aggressive and invasive pancreatic tumors remains a daunting clinical challenge. Thus, the objective of this study was to investigate the role of Shh pathway in pancreatic cancer and to examine the molecular mechanisms by which sulforaphane (SFN, an active compound in cruciferous vegetables, inhibits self-renewal capacity of human pancreatic CSCs. Interestingly, we demonstrate here that Shh pathway is highly activated in pancreatic CSCs and plays important role in maintaining stemness by regulating the expression of stemness genes. Given the requirement for Hedgehog in pancreatic cancer, we investigated whether hedgehog blockade by SFN could target the stem cell population in pancreatic cancer. In an in vitro model, human pancreatic CSCs derived spheres were significantly inhibited on treatment with SFN, suggesting the clonogenic depletion of the CSCs. Interestingly, SFN inhibited the components of Shh pathway and Gli transcriptional activity. Interference of Shh-Gli signaling significantly blocked SFN-induced inhibitory effects demonstrating the requirement of an active pathway for the growth of pancreatic CSCs. SFN also inhibited downstream targets of Gli transcription by suppressing the expression of pluripotency maintaining factors (Nanog and Oct-4 as well as PDGFRα and Cyclin D1. Furthermore, SFN induced apoptosis by inhibition of BCL-2 and activation of caspases. Our data reveal the essential role of Shh-Gli signaling in controlling the characteristics of pancreatic CSCs. We propose that pancreatic cancer preventative effects of SFN may result from inhibition of the Shh pathway

  2. Chloroquine targets pancreatic cancer stem cells via inhibition of CXCR4 and hedgehog signaling

    DEFF Research Database (Denmark)

    Balic, Anamaria; Sørensen, Morten Dræby; Trabulo, Sara Maria

    2014-01-01

    Pancreatic ductal adenocarcinoma is one of the deadliest carcinomas and is characterized by highly tumorigenic and metastatic cancer stem cells (CSC). CSCs evade available therapies, which preferentially target highly proliferative and more differentiated progenies, leaving behind CSCs...... as a putative source for disease relapse. Thus, to identify potentially more effective treatment regimens, we screened established and new compounds for their ability to eliminate CSCs in primary pancreatic cancer (stem) cells in vitro and corresponding patient-derived pancreatic cancer tissue xenografts...... in vivo. Intriguingly, we found that in vitro treatment with the antimalarial agent chloroquine significantly decreased CSCs, translating into diminished in vivo tumorigenicity and invasiveness in a large panel of pancreatic cancers. In vivo treatment in combination with gemcitabine was capable of more...

  3. Targeting cardiac beta-adrenergic signaling via GRK2 inhibition for heart failure therapy

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    Alessandro eCannavo

    2013-09-01

    Full Text Available Cardiac cells, like those of the other tissues, undergo regulation through membrane-bound proteins known as G protein-coupled receptors (GPCRs. β-adrenergic receptors (βARs are key GPCRs expressed on cardiomyocytes and their role is crucial in cardiac physiology since they regulate inotropic and chronotropic responses of the sympathetic nervous system (SNS. In compromised conditions such as heart failure (HF chronic βAR hyperstimulation occurs via SNS activation resulting in receptor dysregulation and down-regulation and consequently there is a marked reduction of myocardial inotropic reserve and continued loss of pump function. Data has accumulated over the last two decades that a primary culprit in initiating and maintain βAR dysfunction in the injured and stressed heart is GPCR kinase 2 (GRK2, which was originally known as βARK1 (for βAR kinase. GRK2 is up-regulated in the failing heart due to chronic SNS activity and targeting this kinase has emerged as a novel therapeutic strategy in HF. Indeed, its inhibition or genetic deletion in several disparate animal models of HF including a pre-clinical pig model has shown that GRK2 targeting improves functional and morphological parameters of the failing heart. Moreover, non-βAR properties of GRK2 appear to also contribute to its pathological effects and thus, its inhibition will likely complement existing therapies such as βAR blockade. This review will explore recent research regarding GRK2 inhibition, in particular it will focus on the GRK2 inhibitor peptide known as βARKct, which represents new hope in the treatment against HF progression. 

  4. HH/GLI signalling as a new therapeutic target for patients with oral squamous cell carcinoma.

    Science.gov (United States)

    Yan, Ming; Wang, Lijun; Zuo, Hui; Zhang, Zhiyuan; Chen, Wantao; Mao, Li; Zhang, Ping

    2011-06-01

    Aberrant activation of HH/GLI has recently been reported in multiple cancer types, yet its role in oral squamous cell carcinoma (OSCC) has not been investigated. In this study, we aimed to determine the role of HH/GLI in OSCC. Expression of GLI1 and GLI2 was examined in OSCC samples from 136 patients by immunohistochemistry and correlated with clinicopathology parameters and clinical outcomes of the patients. Two HH/GLI specific small molecule inhibitors cyclopamine and GANT61, were used to test the potential role of HH/GLI in OSCC. We found that GLI2, one of the main transcriptional activators of HH/GLI signalling, was expressed in 60 (44%) of the 136 OSCC samples and the expression was significantly associated with poor clinical outcomes. Only 44% of the patients whose tumours expressed GLI2 survived at 5years after surgery compared to 77% of those whose tumours lacked the GLI2 expression (PHH/GLI pathway plays an important role in OSCC progression. Together with the finding that expression of GLI2 is strongly associated with a poor clinic outcome of OSCC patients, the data suggest that a subset of OSCC patients may benefit from anti-HH/GLI therapies. Copyright © 2011. Published by Elsevier Ltd.

  5. Pathophysiology of cardiac hypertrophy and heart failure: signaling pathways and novel therapeutic targets.

    Science.gov (United States)

    Tham, Yow Keat; Bernardo, Bianca C; Ooi, Jenny Y Y; Weeks, Kate L; McMullen, Julie R

    2015-09-01

    The onset of heart failure is typically preceded by cardiac hypertrophy, a response of the heart to increased workload, a cardiac insult such as a heart attack or genetic mutation. Cardiac hypertrophy is usually characterized by an increase in cardiomyocyte size and thickening of ventricular walls. Initially, such growth is an adaptive response to maintain cardiac function; however, in settings of sustained stress and as time progresses, these changes become maladaptive and the heart ultimately fails. In this review, we discuss the key features of pathological cardiac hypertrophy and the numerous mediators that have been found to be involved in the pathogenesis of cardiac hypertrophy affecting gene transcription, calcium handling, protein synthesis, metabolism, autophagy, oxidative stress and inflammation. We also discuss new mediators including signaling proteins, microRNAs, long noncoding RNAs and new findings related to the role of calcineurin and calcium-/calmodulin-dependent protein kinases. We also highlight mediators and processes which contribute to the transition from adaptive cardiac remodeling to maladaptive remodeling and heart failure. Treatment strategies for heart failure commonly include diuretics, angiotensin converting enzyme inhibitors, angiotensin II receptor blockers and β-blockers; however, mortality rates remain high. Here, we discuss new therapeutic approaches (e.g., RNA-based therapies, dietary supplementation, small molecules) either entering clinical trials or in preclinical development. Finally, we address the challenges that remain in translating these discoveries to new and approved therapies for heart failure.

  6. SOST and DKK: Antagonists of LRP Family Signaling as Targets for Treating Bone Disease

    Directory of Open Access Journals (Sweden)

    James J. Mason

    2010-01-01

    Full Text Available The study of rare human genetic disorders has often led to some of the most significant advances in biomedical research. One such example was the body of work that resulted in the identification of the Low Density Lipoprotein-Related Protein (LRP5 as a key regulator of bone mass. Point mutations were identified that encoded forms of LRP5 associated with very high bone mass (HBM. HBM patients live to a normal age and do not appear to have increased susceptibility to carcinogenesis or other disease. Thus, devising methods to mimic the molecular consequences of this mutation to treat bone diseases associated with low bone mass is a promising avenue to pursue. Two groups of agents related to putative LRP5/6 functions are under development. One group, the focus of this paper, is based on antagonizing the functions of putative inhibitors of Wnt signaling, Dickkopf-1 (DKK1, and Sclerostin (SOST. Another group of reagents under development is based on the observation that LRP5 may function to control bone mass by regulating the secretion of serotonin from the enterrochromaffin cells of the duodenum.

  7. Kaempferol inhibits cell proliferation and glycolysis in esophagus squamous cell carcinoma via targeting EGFR signaling pathway.

    Science.gov (United States)

    Yao, Shihua; Wang, Xiaowei; Li, Chunguang; Zhao, Tiejun; Jin, Hai; Fang, Wentao

    2016-08-01

    Antitumor activity of kaempferol has been studied in various tumor types, but its potency in esophagus squamous cell carcinoma is rarely known. Here, we reported the activity of kaempferol against esophagus squamous cell carcinoma as well as its antitumor mechanisms. Results of cell proliferation and colony formation assay showed that kaempferol substantially inhibited tumor cell proliferation and clone formation in vitro. Flow cytometric analysis demonstrated that tumor cells were induced G0/G1 phase arrest after kaempferol treatment, and the expression of protein involved in cell cycle regulation was dramatically changed. Except the potency on cell proliferation, we also discovered that kaempferol had a significant inhibitory effect against tumor glycolysis. With the downregulation of hexokinase-2, glucose uptake and lactate production in tumor cells were dramatically declined. Mechanism studies revealed kaempferol had a direct effect on epidermal growth factor receptor (EGFR) activity, and along with the inhibition of EGFR, its downstream signaling pathways were also markedly suppressed. Further investigations found that exogenous overexpression of EGFR in tumor cells substantially attenuated glycolysis suppression induced by kaempferol, which implied that EGFR also played an important role in kaempferol-mediated glycolysis inhibition. Finally, the antitumor activity of kaempferol was validated in xenograft model and kaempferol prominently restrained tumor growth in vivo. Meanwhile, dramatic decrease of EGFR activity and hexokinase-2 expression were observed in kaempferol-treated tumor tissue, which confirmed these findings in vitro. Briefly, these studies suggested that kaempferol, or its analogues, may serve as effective candidates for esophagus squamous cell carcinoma management.

  8. Oligomerization of Paramagnetic Substrates Result in Signal Amplification and Can be Used for MR Imaging of Molecular Targets

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    Alexei Bogdanov

    2002-01-01

    Full Text Available Magnetic resonance imaging (MRI has evolved into a sophisticated, noninvasive imaging modality capable of high-resolution anatomical and functional characterization of transgenic animals. To expand the capabilities MRI, we have developed a novel MR signal amplification (MRamp strategy based on enzyme-mediated polymerization of paramagnetic substrates into oligomers of higher magnetic relaxivity. The substrates consist of chelated gadolinium covalently bound to phenols, which then serve as electron donors during enzymatic hydrogen peroxide reduction by peroxidase. The converted monomers undergo rapid condensation into paramagnetic oligomers leading to a threefold increase in atomic relaxivity (R1/Gd. The observed relaxivity changes are largely due to an increase in the rotational correlation time τr of the lanthanide. Three applications of the developed system are demonstrated: (1 imaging of nanomolar amounts of an oxidoreductase (peroxidase; (2 detection of a model ligand using an enzyme-linked immunoadsorbent assay format; and (3 imaging of E-selectin on the surface of endothelial cells probed for with an anti-E-selectin – peroxidase conjugate. The development of “enzyme sensing” probes is expected to have utility for a number of applications including in vivo detection of specific molecular targets. One particular advantage of the MRamp technique is that the same paramagnetic substrate can be potentially used to identify different molecular targets by attaching enzymes to various antibodies or other target-seeking molecules.

  9. Sigma1 Targeting to Suppress Aberrant Androgen Receptor Signaling in Prostate Cancer.

    Science.gov (United States)

    Thomas, Jeffrey D; Longen, Charles G; Oyer, Halley M; Chen, Nan; Maher, Christina M; Salvino, Joseph M; Kania, Blase; Anderson, Kelsey N; Ostrander, William F; Knudsen, Karen E; Kim, Felix J

    2017-05-01

    Suppression of androgen receptor (AR) activity in prostate cancer by androgen depletion or direct AR antagonist treatment, although initially effective, leads to incurable castration-resistant prostate cancer (CRPC) via compensatory mechanisms including resurgence of AR and AR splice variant (ARV) signaling. Emerging evidence suggests that Sigma1 (also known as sigma-1 receptor) is a unique chaperone or scaffolding protein that contributes to cellular protein homeostasis. We reported previously that some Sigma1-selective small molecules can be used to pharmacologically modulate protein homeostasis pathways. We hypothesized that these Sigma1-mediated responses could be exploited to suppress AR protein levels and activity. Here we demonstrate that treatment with a small-molecule Sigma1 inhibitor prevented 5α- dihydrotestosterone-mediated nuclear translocation of AR and induced proteasomal degradation of AR and ARV, suppressing the transcriptional activity and protein levels of both full-length and splice-variant AR. Consistent with these data, RNAi knockdown of Sigma1 resulted in decreased AR levels and transcriptional activity. Furthermore, Sigma1 physically associated with ARV7 and ARv567es as well as full-length AR. Treatment of mice xenografted with ARV-driven CRPC tumors with a drug-like small-molecule Sigma1 inhibitor significantly inhibited tumor growth associated with elimination of AR and ARV7 in responsive tumors. Together, our data show that Sigma1 modulators can be used to suppress AR/ARV-driven prostate cancer cells via regulation of pharmacologically responsive Sigma1-AR/ARV interactions, both in vitro and in vivoCancer Res; 77(9); 2439-52. ©2017 AACR. ©2017 American Association for Cancer Research.

  10. An Epstein-Barr Virus MicroRNA Blocks Interleukin-1 (IL-1) Signaling by Targeting IL-1 Receptor 1.

    Science.gov (United States)

    Skinner, Camille M; Ivanov, Nikita S; Barr, Sarah A; Chen, Yan; Skalsky, Rebecca L

    2017-11-01

    Epstein-Barr virus (EBV) encodes >44 viral microRNAs (miRNAs) that are differentially expressed throughout infection, can be detected in Epstein-Barr virus (EBV)-positive tumors, and manipulate several biological processes, including cell proliferation, apoptosis, and immune responses. Here, we show that EBV BHRF1-2 miRNAs block NF-κB activation following treatment with proinflammatory cytokines, specifically interleukin-1β (IL-1β). Analysis of EBV PAR-CLIP miRNA targetome data sets combined with pathway analysis revealed multiple BHRF1-2 miRNA targets involved in interleukin signaling pathways. By further analyzing changes in cellular gene expression patterns, we identified the IL-1 receptor 1 (IL1R1) as a direct target of miR-BHRF1-2-5p. Targeting the IL1R1 3' untranslated region (UTR) by EBV miR-BHRF1-2-5p was confirmed using 3'-UTR luciferase reporter assays and Western blot assays. Manipulation of EBV BHRF1-2 miRNA activity in latently infected B cells altered steady-state cytokine levels and disrupted IL-1β responsiveness. These studies demonstrate functionally relevant BHRF1-2 miRNA interactions during EBV infection, which is an important step in understanding their roles in pathogenesis. IMPORTANCE IL-1 signaling plays an important role in inflammation and early activation of host innate immune responses following virus infection. Here, we demonstrate that a viral miRNA downregulates the IL-1 receptor 1 during EBV infection, which consequently alters the responsiveness of cells to IL-1 stimuli and changes the cytokine expression levels within infected cell populations. We postulate that this viral miRNA activity not only disrupts IL-1 autocrine and paracrine signaling loops that can alert effector cells to sites of infection but also provides a survival advantage by dampening excessive inflammation that may be detrimental to the infected cell. Copyright © 2017 American Society for Microbiology.

  11. Extracellular signal regulated kinase 5: A potential therapeutic target for malignant mesotheliomas

    Science.gov (United States)

    Shukla, Arti; Miller, Jill M.; Cason, Christopher; Sayan, Mutlay; MacPherson, Maximilian B.; Beuschel, Stacie L.; Hillegass, Jedd; Vacek, Pamela M.; Pass, Harvey I.; Mossman, Brooke T.

    2013-01-01

    Purpose Malignant mesothelioma (MM) is a devastating disease with a need for new treatment strategies. In the present study we demonstrated the importance of ERK5 in MM tumor growth and treatment. Experimental Design ERK5 as a target for MM therapy was verified using mesothelial and mesothelioma cell lines as well as by xenograft SCID mouse models. Results We first showed that crocidolite asbestos activated ERK5 in LP9 cells and mesothelioma cell lines exhibit constitutive activation of ERK5. Addition of doxorubicin resulted in further activation of ERK5 in MM cells. ERK5 silencing increased DOX-induced cell death and DOX retention in MM cells. In addition, shERK5 MM lines exhibited both attenuated colony formation on soft agar and invasion of MM cells in vitro that could be related to modulation of gene expression linked to cell proliferation, apoptosis, migration/invasion and drug resistance as shown by microarray analysis. Most importantly, injection of shERK5 MM cell lines into SCID mice showed significant reduction in tumor growth using both subcutaneous and intraperitoneal models. Assessment of selected human cytokine profiles in peritoneal lavage fluid from IP shERK5 and control tumor-bearing mice showed that ERK5 was critical in regulation of various proinflammatory (RANTES/CCL5, MCP-1) and angiogenesis related (IL-8, VEGF) cytokines. Finally, use of doxorubicin and cisplatin in combination with ERK5 inhibition showed further reduction in tumor weight and volume in the IP model of tumor growth. Conclusion ; ERK5 inhibition in combination with chemotherapeutic drugs is a beneficial strategy for combination therapy in MM patients. PMID:23446998

  12. Curcumin Differs from Tetrahydrocurcumin for Molecular Targets, Signaling Pathways and Cellular Responses

    Directory of Open Access Journals (Sweden)

    Bharat B. Aggarwal

    2014-12-01

    Full Text Available Curcumin (diferuloylmethane, a golden pigment from turmeric, has been linked with antioxidant, anti-inflammatory, anticancer, antiviral, antibacterial, and antidiabetic properties. Most of the these activities have been assigned to methoxy, hydroxyl, α,β-unsaturated carbonyl moiety or to diketone groups present in curcumin. One of the major metabolites of curcumin is tetrahydrocurcumin (THC, which lacks α,β-unsaturated carbonyl moiety and is white in color. Whether THC is superior to curcumin on a molecular level is unclear and thus is the focus of this review. Various studies suggest that curcumin is a more potent antioxidant than THC; curcumin (but not THC can bind and inhibit numerous targets including DNA (cytosine-5-methyltransferase-1, heme oxygenase-1, Nrf2, β-catenin, cyclooxygenase-2, NF-kappaB, inducible nitric oxide synthase, nitric oxide, amyloid plaques, reactive oxygen species, vascular endothelial growth factor, cyclin D1, glutathione, P300/CBP, 5-lipoxygenase, cytosolic phospholipase A2, prostaglandin E2, inhibitor of NF-kappaB kinase-1, -2, P38MAPK, p-Tau, tumor necrosis factor-α, forkhead box O3a, CRAC; curcumin can inhibit tumor cell growth and suppress cellular entry of viruses such as influenza A virus and hepatitis C virus much more effectively than THC; curcumin affects membrane mobility; and curcumin is also more effective than THC in suppressing phorbol-ester-induced tumor promotion. Other studies, however, suggest that THC is superior to curcumin for induction of GSH peroxidase, glutathione-S-transferase, NADPH: quinone reductase, and quenching of free radicals. Most studies have indicated that THC exhibits higher antioxidant activity, but curcumin exhibits both pro-oxidant and antioxidant properties.

  13. Estrogen enhanced cell-cell signalling in breast cancer cells exposed to targeted irradiation

    Directory of Open Access Journals (Sweden)

    Held Kathryn D

    2008-06-01

    tumour cells may offer new potential targets for radiation-based therapies in the treatment of breast cancer.

  14. HES1, a target of Notch signaling, is elevated in canine osteosarcoma, but reduced in the most aggressive tumors.

    Science.gov (United States)

    Dailey, Deanna D; Anfinsen, Kristin P; Pfaff, Liza E; Ehrhart, E J; Charles, J Brad; Bønsdorff, Tina B; Thamm, Douglas H; Powers, Barbara E; Jonasdottir, Thora J; Duval, Dawn L

    2013-07-01

    Hairy and enhancer of split 1 (HES1), a basic helix-loop-helix transcriptional repressor, is a downstream target of Notch signaling. Notch signaling and HES1 expression have been linked to growth and survival in a variety of human cancer types and have been associated with increased metastasis and invasiveness in human osteosarcoma cell lines. Osteosarcoma (OSA) is an aggressive cancer demonstrating both high metastatic rate and chemotherapeutic resistance. The current study examined expression of Notch signaling mediators in primary canine OSA tumors and canine and human osteosarcoma cell lines to assess their role in OSA development and progression. Reverse transcriptase - quantitative PCR (RT-qPCR) was utilized to quantify HES1, HEY1, NOTCH1 and NOTCH2 gene expression in matched tumor and normal metaphyseal bone samples taken from dogs treated for appendicular OSA at the Colorado State University Veterinary Teaching Hospital. Gene expression was also assessed in tumors from dogs with a disease free interval (DFI) of  300 days following treatment with surgical amputation followed by standard chemotherapy. Immunohistochemistry was performed to confirm expression of HES1. Data from RT-qPCR and immunohistochemical (IHC) experiments were analyzed using REST2009 software and survival analysis based on IHC expression employed the Kaplan-Meier method and log rank analysis. Unbiased clustered images were generated from gene array analysis data for Notch/HES1 associated genes. Gene array analysis of Notch/HES1 associated genes suggested alterations in the Notch signaling pathway may contribute to the development of canine OSA. HES1 mRNA expression was elevated in tumor samples relative to normal bone, but decreased in tumor samples from dogs with a DFI 300 days. NOTCH2 and HEY1 mRNA expression was also elevated in tumors relative to normal bone, but was not differentially expressed between the DFI tumor groups. Survival analysis confirmed an association between

  15. Quercetin inhibits Cr(VI)-induced malignant cell transformation by targeting miR-21-PDCD4 signaling pathway.

    Science.gov (United States)

    Pratheeshkumar, Poyil; Son, Young-Ok; Divya, Sasidharan Padmaja; Wang, Lei; Turcios, Lilia; Roy, Ram Vinod; Hitron, John Andrew; Kim, Donghern; Dai, Jin; Asha, Padmaja; Zhang, Zhuo; Shi, Xianglin

    2017-08-08

    Hexavalent chromium [Cr(VI)] is an important human carcinogen associated with pulmonary diseases and lung cancer. Inhibition of Cr(VI)-induced carcinogenesis by a dietary antioxidant is a novel approach. Quercetin is one of the most abundant dietary flavonoids widely present in many fruits and vegetables, possesses potent antioxidant and anticancer properties. MicroRNA-21 (miR-21) is a key oncomiR significantly elevated in the majority of human cancers that exerts its oncogenic activity by targeting the tumor suppressor gene programmed cell death 4 (PDCD4). The present study examined the effect of quercetin on the inhibition of Cr(VI)-induced malignant cell transformation and the role of miR-21-PDCD4 signaling involved. Our results showed that quercetin decreased ROS generation induced by Cr(VI) exposure in BEAS-2B cells. Chronic Cr(VI) exposure induced malignant cell transformation, increased miR-21 expression and caused inhibition of PDCD4, which were significantly inhibited by the treatment of quercetin in a dose dependent manner. Nude mice injected with BEAS-2B cells chronically exposed to Cr(VI) in the presence of quercetin showed reduced tumor incidence compared to Cr(VI) alone treated group. Stable knockdown of miR-21 and overexpression of PDCD4 or catalase in BEAS-2B cells suppressed Cr(VI)-induced malignant transformation and tumorigenesis. Taken together, these results demonstrate that quercetin is able to protect BEAS-2B cells from Cr(VI)-induced carcinogenesis by targeting miR-21-PDCD4 signaling.

  16. Angiopreventive efficacy of pure flavonolignans from milk thistle extract against prostate cancer: targeting VEGF-VEGFR signaling.

    Directory of Open Access Journals (Sweden)

    Gagan Deep

    Full Text Available The role of neo-angiogenesis in prostate cancer (PCA growth and metastasis is well established, but the development of effective and non-toxic pharmacological inhibitors of angiogenesis remains an unaccomplished goal. In this regard, targeting aberrant angiogenesis through non-toxic phytochemicals could be an attractive angiopreventive strategy against PCA. The rationale of the present study was to compare the anti-angiogenic potential of four pure diastereoisomeric flavonolignans, namely silybin A, silybin B, isosilybin A and isosilybin B, which we established previously as biologically active constituents in Milk Thistle extract. Results showed that oral feeding of these flavonolignans (50 and 100 mg/kg body weight effectively inhibit the growth of advanced human PCA DU145 xenografts. Immunohistochemical analyses revealed that these flavonolignans inhibit tumor angiogenesis biomarkers (CD31 and nestin and signaling molecules regulating angiogenesis (VEGF, VEGFR1, VEGFR2, phospho-Akt and HIF-1α without adversely affecting the vessel-count in normal tissues (liver, lung, and kidney of tumor bearing mice. These flavonolignans also inhibited the microvessel sprouting from mouse dorsal aortas ex vivo, and the VEGF-induced cell proliferation, capillary-like tube formation and invasiveness of human umbilical vein endothelial cells (HUVEC in vitro. Further studies in HUVEC showed that these diastereoisomers target cell cycle, apoptosis and VEGF-induced signaling cascade. Three dimensional growth assay as well as co-culture invasion and in vitro angiogenesis studies (with HUVEC and DU145 cells suggested the differential effectiveness of the diastereoisomers toward PCA and endothelial cells. Overall, these studies elucidated the comparative anti-angiogenic efficacy of pure flavonolignans from Milk Thistle and suggest their usefulness in PCA angioprevention.

  17. Target-of-rapamycin complex 1 (Torc1) signaling modulates cilia size and function through protein synthesis regulation

    Science.gov (United States)

    Yuan, Shiaulou; Li, Jade; Diener, Dennis R.; Choma, Michael A.; Rosenbaum, Joel L.; Sun, Zhaoxia

    2012-01-01

    The cilium serves as a cellular antenna by coordinating upstream environmental cues with numerous downstream signaling processes that are indispensable for the function of the cell. This role is supported by the revelation that defects of the cilium underlie an emerging class of human disorders, termed “ciliopathies.” Although mounting interest in the cilium has demonstrated the essential role that the organelle plays in vertebrate development, homeostasis, and disease pathogenesis, the mechanisms regulating cilia morphology and function remain unclear. Here, we show that the target-of-rapamycin (TOR) growth pathway modulates cilia size and function during zebrafish development. Knockdown of tuberous sclerosis complex 1a (tsc1a), which encodes an upstream inhibitor of TOR complex 1 (Torc1), increases cilia length. In contrast, treatment of embryos with rapamycin, an inhibitor of Torc1, shortens cilia length. Overexpression of ribosomal protein S6 kinase 1 (S6k1), which encodes a downstream substrate of Torc1, lengthens cilia. Furthermore, we provide evidence that TOR-mediated cilia assembly is evolutionarily conserved and that protein synthesis is essential for this regulation. Finally, we demonstrate that TOR signaling and cilia length are pivotal for a variety of downstream ciliary functions, such as cilia motility, fluid flow generation, and the establishment of left-right body asymmetry. Our findings reveal a unique role for the TOR pathway in regulating cilia size through protein synthesis and suggest that appropriate and defined lengths are necessary for proper function of the cilium. PMID:22308353

  18. Reduced mammalian target of rapamycin activity facilitates mitochondrial retrograde signaling and increases life span in normal human fibroblasts

    Science.gov (United States)

    Lerner, Chad; Bitto, Alessandro; Pulliam, Daniel; Nacarelli, Timothy; Konigsberg, Mina; Van Remmen, Holly; Torres, Claudio; Sell, Christian

    2017-01-01

    Summary Coordinated expression of mitochondrial and nuclear genes is required to maintain proper mitochondrial function. However, the precise mechanisms that ensure this coordination are not well defined. We find that signaling from mitochondria to the nucleus is influenced by mammalian target of rapamycin (mTOR) activity via changes in autophagy and p62/SQSTM1 turnover. Reducing mTOR activity increases autophagic flux, enhances mitochondrial membrane potential, reduces reactive oxygen species within the cell, and increases replicative life span. These effects appear to be mediated in part by an interaction between p62/SQSTM1 and Keap1. This interaction allows nuclear accumulation of the nuclear factor erythroid 2-like 2 (NFE2L2, also known as nuclear factor related factor 2 or NRF2), increased expression of the nuclear respiratory factor 1 (NRF1), and increased expression of nuclear-encoded mitochondrial genes, such as the mitochondrial transcription factor A, and mitochondrial-encoded genes involved in oxidative phosphorylation. These findings reveal a portion of the intracellular signaling network that couples mitochondrial turnover with mitochondrial renewal to maintain homeostasis within the cell and suggest mechanisms whereby a reduction in mTOR activity may enhance longevity. PMID:23795962

  19. “Eating” Cancer cells by blocking CD47 signaling: Cancer therapy by targeting the innate immune checkpoint

    Directory of Open Access Journals (Sweden)

    Yi-Rong Xiang

    2017-01-01

    Full Text Available Differing from the adaptive immune checkpoint mediated by programmed cell death-1 (PD-1 PD-1-ligand or CTLA-4, the CD47 and signal regulatory protein α (SIRPα axis is emerging as a novel innate immune checkpoint of the immune cells of myeloid lineage. A balance should be established between the dual signals, the “Don't eat me signal” of CD47-SIRPα and the “Eat me signal” of calreticulin/low-density lipoprotein receptor-related protein. The enhanced expression of CD47 molecule has been found in many cancer tissues, including malignant blood tumors (acute myeloid leukemia and solid tumors. A therapeutic value could be achieved by counteracting the expression of CD47 in cancer cells. In the recent years, great progress has been made to develop anticancer therapies by targeting CD47 (e.g., anti-CD47 antibody, in various types of cancer. However, there are a few challenges, like “antigen sink” in the clinical translation of CD47-mediated anticancer therapies, the attention to which is crucial.

  20. Lawsone derivatives target the Wnt/β-catenin signaling pathway in multidrug-resistant acute lymphoblastic leukemia cells.

    Science.gov (United States)

    Hamdoun, Sami; Fleischer, Edmond; Klinger, Anette; Efferth, Thomas

    2017-12-15

    Multidrug resistance (MDR) represents a serious problem in cancer treatment. One strategy to overcome this obstacle is to identify agents that are selectively lethal to MDR cells. The aim of this study was to discover novel compounds against MDR leukemia and to determine the molecular mechanisms behind collateral sensitivity. A library of 1162 compounds was tested against parental, drug-sensitive CCRF-CEM cells using the resazurin assay. A total of 302 compounds showed reasonable activity (less than 50% cell viability). Eleven out of 30 lawsone derivatives revealed considerable collateral sensitivity in MDR P-glycoprotein (Pgp)-overexpressing CEM/ADR5000 cells. They reduced β-catenin activity in a Wnt/β-catenin reporter cell line. Their activities significantly correlated with apolar desolvation (R = 0.819). Compound (1) (3-hydroxy-1,4-dioxo-N-phenyl-naphthalene-2-carboxamide) was the most active compound and dose-dependently down-regulated protein expression of β-catenin, c-MYC, Pgp and Frizzled 7. By molecular docking, we predicted that compound (1) bound to the palmitoyl-binding groove of the cysteine-rich domain of Frizzled-7 and Frizzled-8. Compound (1) neither stimulated ATPase activity of Pgp nor reactive oxygen species generation, both of which have been previously described as possible mechanisms of collateral sensitivity. Instead, we found that Wnt/β-catenin signaling was selectively inhibited in CEM/ADR5000 cells, but not in CCRF-CEM cells. In conclusion, we found for the first time that the inhibition of Wnt/β-catenin signaling may represent a novel molecular mechanism of collateral sensitivity in MDR cells. Wnt/β-catenin signaling, therefore, represents a potential therapeutic target for the selective killing of Pgp-mediated MDR. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Structures of the signal recognition particle receptor from the archaeon Pyrococcus furiosus: implications for the targeting step at the membrane.

    Science.gov (United States)

    Egea, Pascal F; Tsuruta, Hiro; de Leon, Gladys P; Napetschnig, Johanna; Walter, Peter; Stroud, Robert M

    2008-01-01

    In all organisms, a ribonucleoprotein called the signal recognition particle (SRP) and its receptor (SR) target nascent proteins from the ribosome to the translocon for secretion or membrane insertion. We present the first X-ray structures of an archeal FtsY, the receptor from the hyper-thermophile Pyrococcus furiosus (Pfu), in its free and GDP*magnesium-bound forms. The highly charged N-terminal domain of Pfu-FtsY is distinguished by a long N-terminal helix. The basic charges on the surface of this helix are likely to regulate interactions at the membrane. A peripheral GDP bound near a regulatory motif could indicate a site of interaction between the receptor and ribosomal or SRP RNAs. Small angle X-ray scattering and analytical ultracentrifugation indicate that the crystal structure of Pfu-FtsY correlates well with the average conformation in solution. Based on previous structures of two sub-complexes, we propose a model of the core of archeal and eukaryotic SRP*SR targeting complexes.

  2. Structures of the Signal Recognition Particle Receptor From the Archaeon Pyrococcus Furiosus: Implications for the Targeting Step at the Membrane

    Energy Technology Data Exchange (ETDEWEB)

    Egea, P.F.; Tsuruta, H.; Leon, G.P.de; Napetschnig, J.; Walter, P.; Stroud, R.M.

    2009-05-18

    In all organisms, a ribonucleoprotein called the signal recognition particle (SRP) and its receptor (SR) target nascent proteins from the ribosome to the translocon for secretion or membrane insertion. We present the first X-ray structures of an archeal FtsY, the receptor from the hyper-thermophile Pyrococcus furiosus (Pfu), in its free and GDP {center_dot} magnesium-bound forms. The highly charged N-terminal domain of Pfu-FtsY is distinguished by a long N-terminal helix. The basic charges on the surface of this helix are likely to regulate interactions at the membrane. A peripheral GDP bound near a regulatory motif could indicate a site of interaction between the receptor and ribosomal or SRP RNAs. Small angle X-ray scattering and analytical ultracentrifugation indicate that the crystal structure of Pfu-FtsY correlates well with the average conformation in solution. Based on previous structures of two sub-complexes, we propose a model of the core of archeal and eukaryotic SRP {center_dot} SR targeting complexes.

  3. Combination of Chemical Genetics and Phosphoproteomics for Kinase Signaling Analysis Enables Confident Identification of Cellular Downstream Targets*

    Science.gov (United States)

    Oppermann, Felix S.; Grundner-Culemann, Kathrin; Kumar, Chanchal; Gruss, Oliver J.; Jallepalli, Prasad V.; Daub, Henrik

    2012-01-01

    Delineation of phosphorylation-based signaling networks requires reliable data about the underlying cellular kinase-substrate interactions. We report a chemical genetics and quantitative phosphoproteomics approach that encompasses cellular kinase activation in combination with comparative replicate mass spectrometry analyses of cells expressing either inhibitor-sensitive or resistant kinase variant. We applied this workflow to Plk1 (Polo-like kinase 1) in mitotic cells and induced cellular Plk1 activity by wash-out of the bulky kinase inhibitor 3-MB-PP1, which targets a mutant kinase version with an enlarged catalytic pocket while not interfering with wild-type Plk1. We quantified more than 20,000 distinct phosphorylation sites by SILAC, approximately half of which were measured in at least two independent experiments in cells expressing mutant and wild-type Plk1. Based on replicate phosphorylation site quantifications in both mutant and wild-type Plk1 cells, our chemical genetic proteomics concept enabled stringent comparative statistics by significance analysis of microarrays, which unveiled more than 350 cellular downstream targets of Plk1 validated by full concordance of both statistical and experimental data. Our data point to hitherto poorly characterized aspects in Plk1-controlled mitotic progression and provide a largely extended resource for functional studies. We anticipate the described strategies to be of general utility for systematic and confident identification of cellular protein kinase substrates. PMID:22199227

  4. Combination of chemical genetics and phosphoproteomics for kinase signaling analysis enables confident identification of cellular downstream targets.

    Science.gov (United States)

    Oppermann, Felix S; Grundner-Culemann, Kathrin; Kumar, Chanchal; Gruss, Oliver J; Jallepalli, Prasad V; Daub, Henrik

    2012-04-01

    Delineation of phosphorylation-based signaling networks requires reliable data about the underlying cellular kinase-substrate interactions. We report a chemical genetics and quantitative phosphoproteomics approach that encompasses cellular kinase activation in combination with comparative replicate mass spectrometry analyses of cells expressing either inhibitor-sensitive or resistant kinase variant. We applied this workflow to Plk1 (Polo-like kinase 1) in mitotic cells and induced cellular Plk1 activity by wash-out of the bulky kinase inhibitor 3-MB-PP1, which targets a mutant kinase version with an enlarged catalytic pocket while not interfering with wild-type Plk1. We quantified more than 20,000 distinct phosphorylation sites by SILAC, approximately half of which were measured in at least two independent experiments in cells expressing mutant and wild-type Plk1. Based on replicate phosphorylation site quantifications in both mutant and wild-type Plk1 cells, our chemical genetic proteomics concept enabled stringent comparative statistics by significance analysis of microarrays, which unveiled more than 350 cellular downstream targets of Plk1 validated by full concordance of both statistical and experimental data. Our data point to hitherto poorly characterized aspects in Plk1-controlled mitotic progression and provide a largely extended resource for functional studies. We anticipate the described strategies to be of general utility for systematic and confident identification of cellular protein kinase substrates.

  5. Targeted metabolomics shows plasticity in the evolution of signaling lipids and uncovers old and new endocannabinoids in the plant kingdom.

    Science.gov (United States)

    Gachet, María Salomé; Schubert, Alexandra; Calarco, Serafina; Boccard, Julien; Gertsch, Jürg

    2017-01-25

    The remarkable absence of arachidonic acid (AA) in seed plants prompted us to systematically study the presence of C20 polyunsaturated fatty acids, stearic acid, oleic acid, jasmonic acid (JA), N-acylethanolamines (NAEs) and endocannabinoids (ECs) in 71 plant species representative of major phylogenetic clades. Given the difficulty of extrapolating information about lipid metabolites from genetic data we employed targeted metabolomics using LC-MS/MS and GC-MS to study these signaling lipids in plant evolution. Intriguingly, the distribution of AA among the clades showed an inverse correlation with JA which was less present in algae, bryophytes and monilophytes. Conversely, ECs co-occurred with AA in algae and in the lower plants (bryophytes and monilophytes), thus prior to the evolution of cannabinoid receptors in Animalia. We identified two novel EC-like molecules derived from the eicosatetraenoic acid juniperonic acid, an omega-3 structural isomer of AA, namely juniperoyl ethanolamide and 2-juniperoyl glycerol in gymnosperms, lycophytes and few monilophytes. Principal component analysis of the targeted metabolic profiles suggested that distinct NAEs may occur in different monophyletic taxa. This is the first report on the molecular phylogenetic distribution of apparently ancient lipids in the plant kingdom, indicating biosynthetic plasticity and potential physiological roles of EC-like lipids in plants.

  6. miR-150 suppresses the proliferation and tumorigenicity of leukemia stem cells by targeting the Nanog signaling pathway

    Directory of Open Access Journals (Sweden)

    Dan-dan Xu

    2016-11-01

    Full Text Available Proliferation, a key feature of cancer cells, accounts for the majority of cancer-related diseases resulting in mortality. MicroRNAs (miRNAs plays important post-transcriptional modulation roles by acting on multiple signaling pathways, but the underlying mechanism in proliferation and tumorigenicity is unclear. Here, we identified the role of miR-150 in proliferation and tumorigenicity in leukemia stem cells (LSCs (CD34+CD38- cells. miR-150 expression was significantly down-regulated in LSCs from leukemia cell lines and clinical samples. Functional assays demonstrated that increased miR-150 expression inhibited proliferation and clonal and clonogenic growth, enhanced chemosensitivity, and attenuated tumorigenic activity of LSCs in vitro. Transplantation animal studies revealed that miR-150 overexpression progressively abrogates tumour growth. Immunohistochemistry assays demonstrated that miR-150 overexpression enhanced caspase-3 level and reduced Ki-67 level. Moreover, luciferase reporter assays indicated Nanog is a direct and functional target of miR-150. Nanog silencing using small interfering RNA recapitulated anti-proliferation and tumorigenicity inhibition effects. Furthermore, miR-150 directly down-regulated the expression of other cancer stem cell factors including Notch2 and CTNNB1. These results provide insights into the specific biological behaviour of miR-150 in regulating LSC proliferation and tumorigenicity. Targeting this miR-150/Nanog axis would be a helpful therapeutic strategy to treat acute myeloid leukemia.

  7. Structures of the signal recognition particle receptor from the archaeon Pyrococcus furiosus: implications for the targeting step at the membrane.

    Directory of Open Access Journals (Sweden)

    Pascal F Egea

    Full Text Available In all organisms, a ribonucleoprotein called the signal recognition particle (SRP and its receptor (SR target nascent proteins from the ribosome to the translocon for secretion or membrane insertion. We present the first X-ray structures of an archeal FtsY, the receptor from the hyper-thermophile Pyrococcus furiosus (Pfu, in its free and GDP*magnesium-bound forms. The highly charged N-terminal domain of Pfu-FtsY is distinguished by a long N-terminal helix. The basic charges on the surface of this helix are likely to regulate interactions at the membrane. A peripheral GDP bound near a regulatory motif could indicate a site of interaction between the receptor and ribosomal or SRP RNAs. Small angle X-ray scattering and analytical ultracentrifugation indicate that the crystal structure of Pfu-FtsY correlates well with the average conformation in solution. Based on previous structures of two sub-complexes, we propose a model of the core of archeal and eukaryotic SRP*SR targeting complexes.

  8. Silibinin inhibits fibronectin induced motility, invasiveness and survival in human prostate carcinoma PC3 cells via targeting integrin signaling

    Energy Technology Data Exchange (ETDEWEB)

    Deep, Gagan [Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Denver, Aurora, CO (United States); University of Colorado Cancer Center, University of Colorado Denver, Aurora, CO (United States); Kumar, Rahul; Jain, Anil K. [Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Denver, Aurora, CO (United States); Agarwal, Chapla [Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Denver, Aurora, CO (United States); University of Colorado Cancer Center, University of Colorado Denver, Aurora, CO (United States); Agarwal, Rajesh, E-mail: Rajesh.agarwal@ucdenver.edu [Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Denver, Aurora, CO (United States); University of Colorado Cancer Center, University of Colorado Denver, Aurora, CO (United States)

    2014-10-15

    Highlights: • Silibinin inhibits fibronectin-induce motile morphology in PC3 cells. • Silibinin inhibits fibronectin-induced migration and invasion in PC3 cells. • Silibinin targets fibronectin-induced integrins and downstream signaling molecule. - Abstract: Prostate cancer (PCA) is the 2nd leading cause of cancer-related deaths among men in the United States. Preventing or inhibiting metastasis-related events through non-toxic agents could be a useful approach for lowering high mortality among PCA patients. We have earlier reported that natural flavonoid silibinin possesses strong anti-metastatic efficacy against PCA however, mechanism/s of its action still remains largely unknown. One of the major events during metastasis is the replacement of cell–cell interaction with integrins-based cell–matrix interaction that controls motility, invasiveness and survival of cancer cells. Accordingly, here we examined silibinin effect on advanced human PCA PC3 cells’ interaction with extracellular matrix component fibronectin. Silibinin (50–200 μM) treatment significantly decreased the fibronectin (5 μg/ml)-induced motile morphology via targeting actin cytoskeleton organization in PC3 cells. Silibinin also decreased the fibronectin-induced cell proliferation and motility but significantly increased cell death in PC3 cells. Silibinin also inhibited the PC3 cells invasiveness in Transwell invasion assays with fibronectin or cancer associated fibroblasts (CAFs) serving as chemoattractant. Importantly, PC3-luc cells cultured on fibronectin showed rapid dissemination and localized in lungs following tail vein injection in athymic male nude mice; however, in silibinin-treated PC3-luc cells, dissemination and lung localization was largely compromised. Molecular analyses revealed that silibinin treatment modulated the fibronectin-induced expression of integrins (α5, αV, β1 and β3), actin-remodeling (FAK, Src, GTPases, ARP2 and cortactin), apoptosis (cPARP and

  9. Rapamycin targeting mTOR and hedgehog signaling pathways blocks human rhabdomyosarcoma growth in xenograft murine model

    Energy Technology Data Exchange (ETDEWEB)

    Kaylani, Samer Z. [Division of Hematology and Oncology, Department of Pediatrics, University of Alabama at Birmingham, 1600 7th Avenue South, ACC 414, Birmingham, AL 35233 (United States); Xu, Jianmin; Srivastava, Ritesh K. [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, 1530 3rd Avenue South, VH 509, Birmingham, AL 35294-0019 (United States); Kopelovich, Levy [Division of Cancer Prevention, National Cancer Institute, Bethesda (United States); Pressey, Joseph G. [Division of Hematology and Oncology, Department of Pediatrics, University of Alabama at Birmingham, 1600 7th Avenue South, ACC 414, Birmingham, AL 35233 (United States); Athar, Mohammad, E-mail: mathar@uab.edu [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, 1530 3rd Avenue South, VH 509, Birmingham, AL 35294-0019 (United States)

    2013-06-14

    Graphical abstract: Intervention of poorly differentiated RMS by rapamycin: In poorly differentiated RMS, rapamycin blocks mTOR and Hh signaling pathways concomitantly. This leads to dampening in cell cycle regulation and induction of apoptosis. This study provides a rationale for the therapeutic intervention of poorly differentiated RMS by treating patients with rapamycin alone or in combination with other chemotherapeutic agents. -- Highlights: •Rapamycin abrogates RMS tumor growth by modulating proliferation and apoptosis. •Co-targeting mTOR/Hh pathways underlie the molecular basis of effectiveness. •Reduction in mTOR/Hh pathways diminish EMT leading to reduced invasiveness. -- Abstract: Rhabdomyosarcomas (RMS) represent the most common childhood soft-tissue sarcoma. Over the past few decades outcomes for low and intermediate risk RMS patients have slowly improved while patients with metastatic or relapsed RMS still face a grim prognosis. New chemotherapeutic agents or combinations of chemotherapies have largely failed to improve the outcome. Based on the identification of novel molecular targets, potential therapeutic approaches in RMS may offer a decreased reliance on conventional chemotherapy. Thus, identification of effective therapeutic agents that specifically target relevant pathways may be particularly beneficial for patients with metastatic and refractory RMS. The PI3K/AKT/mTOR pathway has been found to be a potentially attractive target in RMS therapy. In this study, we provide evidence that rapamycin (sirolimus) abrogates growth of RMS development in a RMS xenograft mouse model. As compared to a vehicle-treated control group, more than 95% inhibition in tumor growth was observed in mice receiving parenteral administration of rapamycin. The residual tumors in rapamycin-treated group showed significant reduction in the expression of biomarkers indicative of proliferation and tumor invasiveness. These tumors also showed enhanced apoptosis

  10. Pituitary adenylate cyclase activating polypeptide (PACAP signalling exerts chondrogenesis promoting and protecting effects: implication of calcineurin as a downstream target.

    Directory of Open Access Journals (Sweden)

    Tamás Juhász

    Full Text Available Pituitary adenylate cyclase activating polypeptide (PACAP is an important neurotrophic factor influencing differentiation of neuronal elements and exerting protecting role during traumatic injuries or inflammatory processes of the central nervous system. Although increasing evidence is available on its presence and protecting function in various peripheral tissues, little is known about the role of PACAP in formation of skeletal components. To this end, we aimed to map elements of PACAP signalling in developing cartilage under physiological conditions and during oxidative stress. mRNAs of PACAP and its receptors (PAC1,VPAC1, VPAC2 were detectable during differentiation of chicken limb bud-derived chondrogenic cells in micromass cell cultures. Expression of PAC1 protein showed a peak on days of final commitment of chondrogenic cells. Administration of either the PAC1 receptor agonist PACAP 1-38, or PACAP 6-38 that is generally used as a PAC1 antagonist, augmented cartilage formation, stimulated cell proliferation and enhanced PAC1 and Sox9 protein expression. Both variants of PACAP elevated the protein expression and activity of the Ca-calmodulin dependent Ser/Thr protein phosphatase calcineurin. Application of PACAPs failed to rescue cartilage formation when the activity of calcineurin was pharmacologically inhibited with cyclosporine A. Moreover, exogenous PACAPs prevented diminishing of cartilage formation and decrease of calcineurin activity during oxidative stress. As an unexpected phenomenon, PACAP 6-38 elicited similar effects to those of PACAP 1-38, although to a different extent. On the basis of the above results, we propose calcineurin as a downstream target of PACAP signalling in differentiating chondrocytes either in normal or pathophysiological conditions. Our observations imply the therapeutical perspective that PACAP can be applied as a natural agent that may have protecting effect during joint inflammation and/or may promote

  11. SEL-10 is an inhibitor of notch signaling that targets notch for ubiquitin-mediated protein degradation.

    Science.gov (United States)

    Wu, G; Lyapina, S; Das, I; Li, J; Gurney, M; Pauley, A; Chui, I; Deshaies, R J; Kitajewski, J

    2001-11-01

    Notch receptors and their ligands play important roles in both normal animal development and pathogenesis. We show here that the F-box/WD40 repeat protein SEL-10 negatively regulates Notch receptor activity by targeting the intracellular domain of Notch receptors for ubiquitin-mediated protein degradation. Blocking of endogenous SEL-10 activity was done by expression of a dominant-negative form containing only the WD40 repeats. In the case of Notch1, this block leads to an increase in Notch signaling stimulated by either an activated form of the Notch1 receptor or Jagged1-induced signaling through Notch1. Expression of dominant-negative SEL-10 leads to stabilization of the intracellular domain of Notch1. The Notch4 intracellular domain bound to SEL-10, but its activity was not increased as a result of dominant-negative SEL-10 expression. SEL-10 bound Notch4 via the WD40 repeats and bound preferentially to a phosphorylated form of Notch4 in cells. We mapped the region of Notch4 essential for SEL-10 binding to the C-terminal region downstream of the ankyrin repeats. When this C-terminal fragment of Notch4 was expressed in cells, it was highly labile but could be stabilized by the expression of dominant-negative SEL-10. Ubiquitination of Notch1 and Notch4 intracellular domains in vitro was dependent on SEL-10. Although SEL-10 interacts with the intracellular domains of both Notch1 and Notch4, these proteins respond differently to interference with SEL-10 function. Thus, SEL-10 functions to promote the ubiquitination of Notch proteins; however, the fates of these proteins may differ.

  12. Protease-activated receptors (PARs) in cancer: Novel biased signaling and targets for therapy.

    Science.gov (United States)

    Bar-Shavit, R; Maoz, M; Kancharla, A; Jaber, M; Agranovich, D; Grisaru-Granovsky, S; Uziely, B

    2016-01-01

    Despite the fact that G protein-coupled receptors (GPCRs) mediate numerous physiological processes and represent targets for therapeutics for a vast array of diseases, their role in tumor biology is under appreciated. Protease-activated receptors (PARs) form a family which belongs to GPCR class A. PAR1&2 emerge with a central role in epithelial malignancies. Although the part of PAR1&2 in cancer is on the rise, their underlying signaling events are poorly understood. We review hereby past, present, and future cancer-associated PAR biology. Mainly, their role in physiological (placenta-cytotophobalst) and patho-physiological invasion processes. The identification and characterization of signal pleckstrin homology (PH)-domain-binding motifs established critical sites for breast cancer growth in PAR1&2. Among the proteins found to harbor important PH-domains and are involved in PAR biology are Akt/PKB as also Etk/Bmx and Vav3. A point mutation in PAR2, H349A, but not R352A, abrogated PH-protein association and is sufficient to markedly reduce PAR2-instigated breast tumor growth in vivo as also placental extravillous trophoblast (EVT) invasion in vitro is markedly reduced. Similarly, the PAR1 mutant hPar1-7A, which is unable to bind PH-domain, inhibits mammary tumors and EVT invasion, endowing these motifs with physiological significance and underscoring the importance of these previously unknown PAR1 and PAR2 PH-domain-binding motifs in both pathological and physiological invasion processes. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. Targeting MTA1/HIF-1α signaling by pterostilbene in combination with histone deacetylase inhibitor attenuates prostate cancer progression.

    Science.gov (United States)

    Butt, Nasir A; Kumar, Avinash; Dhar, Swati; Rimando, Agnes M; Akhtar, Israh; Hancock, John C; Lage, Janice M; Pound, Charles R; Lewin, Jack R; Gomez, Christian R; Levenson, Anait S

    2017-10-10

    The metastasis-associated protein 1(MTA1)/histone deacetylase (HDAC) unit is a cancer progression-related epigenetic regulator, which is overexpressed in hormone-refractory and metastatic prostate cancer (PCa). In our previous studies, we found a significantly increased MTA1 expression in a prostate-specific Pten-null mouse model. We also demonstrated that stilbenes, namely resveratrol and pterostilbene (Pter), affect MTA1/HDAC signaling, including deacetylation of tumor suppressors p53 and PTEN. In this study, we examined whether inhibition of MTA1/HDAC using combination of Pter and a clinically approved HDAC inhibitor, SAHA (suberoylanilide hydroxamic acid, vorinostat), which also downregulates MTA1, could block prostate tumor progression in vivo. We generated and utilized a luciferase reporter in a prostate-specific Pten-null mouse model (Pb-Cre(+) ; Pten(f/f) ; Rosa26(Luc/+) ) to evaluate the anticancer efficacy of Pter/SAHA combinatorial approach. Our data showed that Pter sensitized tumor cells to SAHA treatment resulting in inhibiting tumor growth and additional decline of tumor progression. These effects were dependent on the reduction of MTA1-associated proangiogenic factors HIF-1α, VEGF, and IL-1β leading to decreased angiogenesis. In addition, treatment of PCa cell lines in vitro with combined Pter and low dose SAHA resulted in more potent inhibition of MTA1/HIF-1α than by high dose SAHA alone. Our study provides preclinical evidence that Pter/SAHA combination treatment inhibits MTA1/HIF-1α tumor-promoting signaling in PCa. The beneficial outcome of combinatorial strategy using a natural agent and an approved drug for higher efficacy and less toxicity supports further development of MTA1-targeted therapies in PCa. © 2017 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

  14. MicroRNA-24 promotes 3T3-L1 adipocyte differentiation by directly targeting the MAPK7 signaling

    Energy Technology Data Exchange (ETDEWEB)

    Jin, Min, E-mail: min_jin@zju.edu.cn [Division of Reproductive Medicine & Infertility, The Second Affiliated Hospital, School of Medicine, Zhejiang University, 88#, Jiefang Rd., Hangzhou, Zhejiang, 310009 (China); Wu, Yutao; Wang, Jing [School of Medicine, Zhejiang University, 288# Yuhangtang Rd, Hangzhou, Zhejiang, 310003 (China); Chen, Jian; Huang, Yiting; Rao, Jinpeng; Feng, Chun [Division of Reproductive Medicine & Infertility, The Second Affiliated Hospital, School of Medicine, Zhejiang University, 88#, Jiefang Rd., Hangzhou, Zhejiang, 310009 (China)

    2016-05-20

    Over the past years, MicroRNAs (miRNAs) act as a vital role in harmony with gene regulation and maintaining cellular homeostasis. It is well testified that miRNAshave been involved in numerous physiological and pathological processes, including embryogenesis, cell fate decision, and cellular differentiation. Adipogenesis is an organized process of cellular differentiation by which pre-adipocytes differentiate towards mature adipocytes, and it is tightly modulated by a series of transcription factors such as peroxisome proliferator-activated receptor γ (PPAR-γ) and sterol regulatory-element binding proteins 1 (SREBP1). However, the molecular mechanisms underlying the connection between miRNAs and adipogenesis-related transcription factors remain obscure. In this study, we unveiled that miR- 24 was remarkably upregulated during 3T3-L1 adipogenesis. Overexpression of miR-24 significantly promoted 3T3-L1 adipogenesis, as evidenced by its ability to increase the expression of PPAR-γ and SREBP1, lipid droplet formation and triglyceride (TG) accumulation. Furthermore, we found that neither ectopic expression of miR-24nor miR-24 inhibitor affect cell proliferation and cell cycle progression. Finally, we demonstrated that miR-24 plays the modulational role by directly repressing MAPK7, a key number in the MAPK signaling pathway. These data indicate that miR-24 is a novel positive regulator of adipocyte differentiation by targeting MAPK7, which provides new insights into the molecular mechanism of miRNA-mediated cellular differentiation. -- Highlights: •We firstly found miR-24 was upregulated in 3T3-L1 pre-adipocytes differentiation. •miR-24 promoted 3T3-L1 pre-adipocytes differentiation while silencing the expression of miR-24 had an opposite function. •miR-24 regulated 3T3-L1 differentiation by directly targeting MAPK7 signaling pathway. •miR-24did not affect 3T3-L1 pre-adipocytes cellular proliferation.

  15. CX-5461 induces autophagy and inhibits tumor growth via mammalian target of rapamycin-related signaling pathways in osteosarcoma

    Directory of Open Access Journals (Sweden)

    Li L

    2016-09-01

    Full Text Available Leiming Li,1,* Yan Li,2,* Jiansong Zhao,2 Shuli Fan,3 Liguo Wang,1 Xu Li1 1Department of Joint Surgery and Sports Medicine, The First Affiliated Hospital, 2Department of Spine and Joint Surgery, Sheng Jing Hospital, 3Department of Geriatrics, The First Affiliated Hospital, China Medical University, Shenyang, People’s Republic of China *These authors contributed equally to this work Abstract: Osteosarcoma (OS is the most common primary bone tumor, but molecular mechanisms of the disease have not been well understood, and treatment of metastatic OS remains a challenge. Rapid ribosomal RNA synthesis in cancer is transcribed by RNA polymerase I, which results in unbridled cell growth. The recent discovery of CX-5461, a selective RNA polymerase I inhibitor, exerted its inhibitory effect of ribosomal RNA synthesis and antiproliferative potency. Here, we demonstrate that CX-5461 induces G2 arrest in the cell cycle and expression of microtubule-associated protein 1 light chain 3 II isoform in OS cell lines. Autophagic vacuoles could be observed in electron microscopy and 3-methyladenine prevented cell death mediated by CX-5461. Moreover, it significantly augmented phosphorylated AMP-Activated Protein Kinases α (p-AMPK α. (Thr172 expression in U2-OS cells and decreased p-Akt (Ser473 expression in MNNG cells, respectively, which repressed their downstream effector, mammalian target of rapamycin. On the other hand, CX-5461 increased p53 accumulation and messenger RNA level of its target genes, p21, MDM2, and Sestrin1/2 in U2-OS cells. Knockdown of p53 expression markedly impaired cell death as well as the expression of light chain 3-II and p21 induced by CX-5461. It also significantly enhanced doxorubicin-mediated cytotoxic effect in vitro and in vivo together with additive expression of p53, p21, and light chain 3-II in U2-OS cells. Our data indicate that CX-5461 might induce autophagy via mammalian target of rapamycin-associated signaling pathways

  16. Signal transduction controls heterogeneous NF-κB dynamics and target gene expression through cytokine-specific refractory states

    Science.gov (United States)

    Adamson, Antony; Boddington, Christopher; Downton, Polly; Rowe, William; Bagnall, James; Lam, Connie; Maya-Mendoza, Apolinar; Schmidt, Lorraine; Harper, Claire V.; Spiller, David G.; Rand, David A.; Jackson, Dean A.; White, Michael R. H.; Paszek, Pawel

    2016-01-01

    Cells respond dynamically to pulsatile cytokine stimulation. Here we report that single, or well-spaced pulses of TNFα (>100 min apart) give a high probability of NF-κB activation. However, fewer cells respond to shorter pulse intervals (refractory state. This refractory state is established in the signal transduction network downstream of TNFR and upstream of IKK, and depends on the level of the NF-κB system negative feedback protein A20. If a second pulse within the refractory phase is IL-1β instead of TNFα, all of the cells respond. This suggests a mechanism by which two cytokines can synergistically activate an inflammatory response. Gene expression analyses show strong correlation between the cellular dynamic response and NF-κB-dependent target gene activation. These data suggest that refractory states in the NF-κB system constitute an inherent design motif of the inflammatory response and we suggest that this may avoid harmful homogenous cellular activation. PMID:27381163

  17. ITGBL1 Is a Runx2 Transcriptional Target and Promotes Breast Cancer Bone Metastasis by Activating the TGFβ Signaling Pathway.

    Science.gov (United States)

    Li, Xiao-Qing; Du, Xin; Li, Dong-Mei; Kong, Peng-Zhou; Sun, Yan; Liu, Pei-Fang; Wang, Qing-Shan; Feng, Yu-Mei

    2015-08-15

    Bone metastasis affects more than 70% of advanced breast cancer patients, but the molecular mechanisms of this process remain unclear. Here, we present clinical and experimental evidence to clarify the role of the integrin β-like 1 (ITGBL1) as a key contributor to bone metastasis of breast cancer. In an in vivo model system and in vitro experiments, ITGBL1 expression promoted formation of osteomimetic breast cancers, facilitating recruitment, residence, and growth of cancer cells in bone microenvironment along with osteoclast maturation there to form osteolytic lesions. Mechanistic investigations identified the TGFβ signaling pathway as a downstream effector of ITGBL1 and the transcription factor Runx2 as an upstream activator of ITGBL1 expression. In support of these findings, we also found that ITGBL1 was an essential mediator of Runx2-induced bone metastasis of breast cancer. Overall, our results illuminate how bone metastasis occurs in breast cancer, and they provide functional evidence for new candidate biomarkers and therapeutic targets to identify risk, to prevent, and to treat this dismal feature of advanced breast cancer. ©2015 American Association for Cancer Research.

  18. Naringin inhibits growth potential of human triple-negative breast cancer cells by targeting β-catenin signaling pathway.

    Science.gov (United States)

    Li, Hongzhong; Yang, Bing; Huang, Jing; Xiang, Tingxiu; Yin, Xuedong; Wan, Jingyuan; Luo, Fuling; Zhang, Li; Li, Hongyuan; Ren, Guosheng

    2013-07-18

    Triple-negative (ER-/PR-/HER2-) breast cancer (TNBC) is a severe clinical problem because of its relatively poorer prognosis, aggressive behavior and lack of targeted therapies. Naringin, a major flavonoid extracted from citrus fruits, has been reported to exert promising anticancer activities. However, the detailed antitumor mechanism of naringin still remains enigmatic. In this study, TNBC cell lines-based in vitro and in vivo models were used to explore the anticancer effect and mechanism of naringin. Our data demonstrated that naringin inhibited cell proliferation, and promoted cell apoptosis and G1 cycle arrest, accompanied by increased p21 and decreased survivin. Meanwhile, β-catenin signaling pathway was found to be suppressed by naringin. In contrast, over-expressing β-catenin by adenoviral vector system in TNBC cells reversed the antitumor activity of naringin, and regulated p21 and survivin. Correspondingly, the antitumor potential of naringin was also observed in naringin-treated MDA-MB-231 xenograft mice, while immunohistochemical analysis of tumors from naringin-treated mice showed higher expression of p21 and lower expression of survivin and active β-catenin. Taken together, these results indicate that naringin could inhibit growth potential of TNBC cells by modulating β-catenin pathway, which suggests naringin might be used as a potential supplement for the prevention and treatment of breast cancer. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  19. A cryptic targeting signal creates a mitochondrial FEN1 isoform with tailed R-Loop binding properties.

    Directory of Open Access Journals (Sweden)

    Lawrence Kazak

    Full Text Available A growing number of DNA transacting proteins is found in the nucleus and in mitochondria, including the DNA repair and replication protein Flap endonuclease 1, FEN1. Here we show a truncated FEN1 isoform is generated by alternative translation initiation, exposing a mitochondrial targeting signal. The shortened form of FEN1, which we term FENMIT, localizes to mitochondria, based on import into isolated organelles, immunocytochemistry and subcellular fractionation. In vitro FENMIT binds to flap structures containing a 5' RNA flap, and prefers such substrates to single-stranded RNA. FENMIT can also bind to R-loops, and to a lesser extent to D-loops. Exposing human cells to ethidium bromide results in the generation of RNA/DNA hybrids near the origin of mitochondrial DNA replication. FENMIT is recruited to the DNA under these conditions, and is released by RNase treatment. Moreover, high levels of recombinant FENMIT expression inhibit mtDNA replication, following ethidium bromide treatment. These findings suggest FENMIT interacts with RNA/DNA hybrids in mitochondrial DNA, such as those found at the origin of replication.

  20. Comparative gene expression analysis of Dtg, a novel target gene of Dpp signaling pathway in the early Drosophila melanogaster embryo.

    Science.gov (United States)

    Hodar, Christian; Zuñiga, Alejandro; Pulgar, Rodrigo; Travisany, Dante; Chacon, Carlos; Pino, Michael; Maass, Alejandro; Cambiazo, Verónica

    2014-02-10

    In the early Drosophila melanogaster embryo, Dpp, a secreted molecule that belongs to the TGF-β superfamily of growth factors, activates a set of downstream genes to subdivide the dorsal region into amnioserosa and dorsal epidermis. Here, we examined the expression pattern and transcriptional regulation of Dtg, a new target gene of Dpp signaling pathway that is required for proper amnioserosa differentiation. We showed that the expression of Dtg was controlled by Dpp and characterized a 524-bp enhancer that mediated expression in the dorsal midline, as well as, in the differentiated amnioserosa in transgenic reporter embryos. This enhancer contained a highly conserved region of 48-bp in which bioinformatic predictions and in vitro assays identified three Mad binding motifs. Mutational analysis revealed that these three motifs were necessary for proper expression of a reporter gene in transgenic embryos, suggesting that short and highly conserved genomic sequences may be indicative of functional regulatory regions in D. melanogaster genes. Dtg orthologs were not detected in basal lineages of Dipterans, which unlike D. melanogaster develop two extra-embryonic membranes, amnion and serosa, nevertheless Dtg orthologs were identified in the transcriptome of Musca domestica, in which dorsal ectoderm patterning leads to the formation of a single extra-embryonic membrane. These results suggest that Dtg was recruited as a new component of the network that controls dorsal ectoderm patterning in the lineage leading to higher Cyclorrhaphan flies, such as D. melanogaster and M. domestica. Copyright © 2013 Elsevier B.V. All rights reserved.

  1. Resveratrol inhibits Hexokinases II mediated glycolysis in non-small cell lung cancer via targeting Akt signaling pathway.

    Science.gov (United States)

    Li, Wei; Ma, Xiaoqian; Li, Na; Liu, Huasheng; Dong, Qiong; Zhang, Juan; Yang, Cejun; Liu, Yin; Liang, Qi; Zhang, Shengwang; Xu, Chang; Song, Wei; Tan, Shiming; Rong, Pengfei; Wang, Wei

    2016-12-10

    Deregulation of glycolysis was often observed in human cancer cells. In the present study, we reported resveratrol, a small polyphenol, which has been intensively studied in various tumor models, has a profound anti-tumor effect on human non-small cell lung cancer (NSCLC) via regulation of glycolysis. Resveratrol impaired hexokinase II (HK2)-mediated glycolysis, and markedly inhibited anchorage-dependent and -independent growth of NSCLC cells. Exposure to resveratrol decreased EGFR and downstream kinases Akt and ERK1/2 activation. Moreover, we revealed that resveratrol impaired glucose metabolism by mainly inhibiting expression of HK2 mediated by the Akt signaling pathway, and exogenous overexpression of constitutively activated Akt1 in NSCLC cells substantially rescued resveratrol-induced glycolysis suppression. The in vivo data indicated that resveratrol obviously suppressed tumor growth in a xenograft mouse model. Our results suggest targeting HK2 or metabolic enzymes appears to be a new approach for clinical NSCLC prevention or treatment. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. Immunohistochemical analysis of phosphorylated mammalian target of rapamycin and its downstream signaling components in invasive breast cancer.

    Science.gov (United States)

    Ma, Bin-Lin; Shan, Mei-Hui; Sun, Gang; Ren, Guang-Hui; Dong, Chao; Yao, Xuemei; Zhou, Mei

    2015-10-01

    The present study aimed to investigate whether the mammalian target of rapamycin (mTOR) signaling pathway is activated in invasive breast cancer. The expression levels of phosphorylated (p)‑mTOR at ser2448 were detected, as well as the expression levels of its downstream signaling molecules: Eukaryotic translation initiation factor 4E‑binding protein 1 (4E‑BP1), and p70 ribosomal protein S6 kinase 1 (S6K1). The correlation between p‑mTOR, p‑4E‑BP1, p‑S6K1, and the clinicopathological parameters of breast cancer were also determined. p‑mTOR, p‑4E‑BP1 and p‑S6K1 expression was detected in 285 breast cancer tumor samples and adjacent normal tissue samples using immunohistochemistry. The expression levels and the location of the proteins were analyzed and compared in the various tissue samples. Multivariate Cox regression was used to analyze the clinicopathological factors and prognosis associated with the tissue samples. The disease‑free survival rate was examined using survival analyses and Log‑rank tests. The results of the present study indicated that the expression levels of p‑mTOR, p‑4E‑BP1, and p‑S6K1 were significantly higher in breast cancer tissue, as compared with normal tissue (Pbreast cancer tissue samples, as compared with normal tissue samples (P0.05). Thus suggesting that these markers are not adequate risk factors for disease free survival (P>0.05). In conclusion, the results of the present study suggested that p‑mTOR, p‑4E‑BP1, and p‑S6K1 are activated in invasive breast cancer. In addition, the exclusive expression of p‑4E‑BP1 and p‑S6K1 in the cytoplasm may be characteristic of progressive breast cancer. However, p‑mTOR, p‑4E‑BP1, and p‑S6K1 are not prognostic factors for breast cancer.

  3. Rac1/β-Catenin Signalling Pathway Contributes to Trophoblast Cell Invasion by Targeting Snail and MMP9

    Directory of Open Access Journals (Sweden)

    Minghua Fan

    2016-03-01

    Full Text Available Background/Aims: Preeclampsia is an idiopathic and serious complication during gestation in which placental trophoblast cells differentiate into several functional subtypes, including highly invasive extravillous trophoblasts (EVTs. Although the cause and pathogenesis of preeclampsia have remained unclear, numerous studies have suggested that the inadequacy of EVT invasion leads to imperfect uterine spiral artery remodelling, which plays a crucial role in the development of preeclampsia. Rac1, or Ras-related C3 botulinum toxin substrate 1, was found to be a key regulator of the migration, invasion uand apoptosis of various tumour cells. Because EVTs share similar invasive and migratory biological behaviours with malignant cells, this study aimed to determine whether the Rac1 signalling pathway affects trophoblast invasion and is thus involved in the pathogenesis of preeclampsia. Methods: We measured the activity of Rac1 and its downstream targets, β-catenin, Snail and MMP9 in placental tissues from patients experiencing a normal pregnancy and those with preeclampsia. Furthermore, we treated HTR-8/SVneo cells with a shRNA Rac1 vector and the β-catenin inhibitor IWP-2 and explored Rac1 signalling pathway activation as well as the effects of Snail and β-catenin on trophoblast invasion. Results: In placental samples from patients experiencing a normal pregnancy and those with preeclampsia, active Rac1 levels and MMP9 protein and mRNA levels were significantly decreased in term pregnancy samples compared to early pregnancy samples. Lower levels were found in preeclampsia samples than in normal term pregnancy samples, and these levels significantly declined in severe preeclampsia samples compared with mild preeclampsia samples. Further analyses demonstrated that both Rac1 shRNA and the β-catenin inhibitor significantly suppressed MMP9 and Snail activation in trophoblasts, thus impairing trophoblast invasion. Notably, silencing Rac1 down

  4. The E3 Ubiquitin Ligase WWP1 Selectively Targets HER4 and Its Proteolytically Derived Signaling Isoforms for Degradation▿

    Science.gov (United States)

    Feng, Shu-Mang; Muraoka-Cook, Rebecca S.; Hunter, Debra; Sandahl, Melissa A.; Caskey, Laura S.; Miyazawa, Keiji; Atfi, Azeddine; Earp, H. Shelton

    2009-01-01

    In general, epidermal growth factor receptor family members stimulate cell proliferation. In contrast, at least one HER4 isoform, JM-a/Cyt1, inhibits cell growth after undergoing a two-step proteolytic cleavage that first produces a membrane-anchored 80-kDa fragment (m80HER4) and subsequently liberates a soluble 80-kDa fragment, s80HER4. Here we report that s80HER4 Cyt1 action increased the expression of WWP1 (for WW domain-containing protein 1), an E3 ubiquitin ligase, but not other members of the Nedd4 E3 ligase family. The HER4 Cyt1 isoform contains three proline-rich tyrosine (PY) WW binding motifs, while Cyt2 has only two. WWP1 binds to all three Cyt1 PY motifs; the interaction with PY2 found exclusively in Cyt1 was strongest. WWP1 ubiquitinated and caused the degradation of HER4 but not of EGFR, HER2, or HER3. The HER4-WWP1 interaction also accelerated WWP1 degradation. Membrane HER4 (full length and m80HER4, the product of the first proteolytic cleavage) were the preferred targets of WWP1, correlating with the membrane localization of WWP1. Conversely s80HER4, a poorer WWP1 substrate, was found in the cell nucleus, while WWP1 was not. Deletion of the C2 membrane association domain of WWP1 allowed more efficient s80HER4 degradation, suggesting that WWP1 is normally part of a membrane complex that regulates HER4 membrane species levels, with a predilection for the growth-inhibitory Cyt1 isoform. Finally, WWP1 expression diminished HER4 biologic activity in MCF-7 cells. We previously showed that nuclear s80HER4 is ubiquitinated and degraded by the anaphase-promoting complex, suggesting that HER4 ubiquitination within specific cellular compartments helps regulate the unique HER4 signaling capabilities. PMID:19047365

  5. Replication Study: The CD47-signal regulatory protein alpha (SIRPa) interaction is a therapeutic target for human solid tumors.

    Science.gov (United States)

    Horrigan, Stephen K

    2017-01-19

    In 2015, as part of the Reproducibility Project: Cancer Biology, we published a Registered Report (Chroscinski et al., 2015) that described how we intended to replicate selected experiments from the paper "The CD47-signal regulatory protein alpha (SIRPa) interaction is a therapeutic target for human solid tumors "(Willingham et al., 2012). Here we report the results of those experiments. We found that treatment of immune competent mice bearing orthotopic breast tumors with anti-mouse CD47 antibodies resulted in short-term anemia compared to controls, consistent with the previously described function of CD47 in normal phagocytosis of aging red blood cells and results reported in the original study (Table S4; Willingham et al., 2012). The weight of tumors after 30 days administration of anti-CD47 antibodies or IgG isotype control were not found to be statistically different, whereas the original study reported inhibition of tumor growth with anti-CD47 treatment (Figure 6A,B; Willingham et al., 2012). However, our efforts to replicate this experiment were confounded because spontaneous regression of tumors occurred in several of the mice. Additionally, the excised tumors were scored for inflammatory cell infiltrates. We found IgG and anti-CD47 treated tumors resulted in minimal to moderate lymphocytic infiltrate, while the original study observed sparse lymphocytic infiltrate in IgG-treated tumors and increased inflammatory cell infiltrates in anti-CD47 treated tumors (Figure 6C; Willingham et al., 2012). Furthermore, we observed neutrophilic infiltration was slightly increased in anti-CD47 treated tumors compared to IgG control. Finally, we report a meta-analysis of the result.

  6. Theoretical nuclear and subnuclear physics

    CERN Document Server

    Walecka, John Dirk

    1995-01-01

    This comprehensive text expertly details the numerous theoretical techniques central to the discipline of nuclear physics. It is based on lecture notes from a three-lecture series given at CEBAF (the Continuous Electron Beam Accelerator Facility), where John Dirk Walecka at the time was Scientific Director: "Graduate Quantum Mechanics", "Advanced Quantum Mechanics and Field Theory" and "Special Topics in Nuclear Physics". The primary goal of this text is pedagogical; providing a clear, logical, in-depth, and unifying treatment of many diverse aspects of modern nuclear theory ranging from the non-relativistic many-body problem to the standard model of the strong, electromagnetic, and weak interactions. Four key topics are emphasised in this text: basic nuclear structure, the relativistic nuclear many-body problem, strong-coupling QCD, and electroweak interactions with nuclei. The text is designed to provide graduate students with a basic level of understanding of modern nuclear physics so that they in turn can...

  7. An electrochemical aptasensor for multiplex antibiotics detection based on metal ions doped nanoscale MOFs as signal tracers and RecJfexonuclease-assisted targets recycling amplification.

    Science.gov (United States)

    Chen, Meng; Gan, Ning; Zhou, You; Li, Tianhua; Xu, Qing; Cao, Yuting; Chen, Yinji

    2016-12-01

    An ultrasensitive electrochemical aptasensor for simultaneous detection of oxytetracycline (OTC) and kanamycin (KAN) has been developed based on metal ions doped metal organic frame materials (MOFs) as signal tracers and RecJ f exonuclease-catalyzed targets recycling amplification. The aptasensor consists of capture beads (the anti-single-stranded DNA Antibody, as anti-ssDNA Ab, labeled on Dynabeads) and nanoscale MOF (NMOF) based signal tracers (simplified as Apts-MNM, the NMOF labeled with metal ions and the aptamers). Particularly, the MOF (UiO-66-NH 2 ), with large internal surface areas, ultrahigh porosity and abundant amine groups in the pores, was employed as substrates to carry plenty of metal ions (Pb 2+ or Cd 2+ ) and label aptamers of OTC or KAN. Thus, the aptasensor is formed by the specific recognition between anti-ssDNA Ab and aptamers. In the presence of targets (OTC and KAN), aptamers prefer to form targets-Apts-MNM complexes in lieu of anti-ssDNA Ab-aptamer complexes, which results in the dissociation of Apts-MNM from capture beads. With the employment of RecJ f exonuclease, targets-Apts-MNM in supernatant was digested into mononucleotides and liberated the target, which can further participate in the next reaction cycling to produce more signal tracers. After magnetic separation, the enhanced square wave voltammetry (SWV) signals were produced from signal tracers. The aptasensor exhibited a linear correlation in the range from 0.5pM to 50nM, with detection limits of 0.18pM and 0.15pM (S/N=3) toward OTC and KAN respectively. This strategy provides specificity and sensitive approach for multiplex antibiotics detection and has promising applications in food analysis. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. miR-130b targets NKD2 and regulates the Wnt signaling to promote proliferation and inhibit apoptosis in osteosarcoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Li, Zhi [Department of Human Anatomy and Histoembryology, College of Basic Medical Sciences, Jilin University (China); Li, Youjun, E-mail: liyoujunn@126.com [Department of Human Anatomy and Histoembryology, College of Basic Medical Sciences, Jilin University (China); Wang, Nan; Yang, Lifeng; Zhao, Wei; Zeng, Xiandong [Central Hospital Affiliated to Shenyang Medical College (China)

    2016-03-18

    miR-130b was significantly up-regulated in osteosarcoma (OS) cells. Naked cuticle homolog 2 (NKD2) inhibited tumor growth and metastasis in OS by suppressing Wnt signaling. We used three miRNA target analysis tools to identify potential targets of miR-130b, and found that NKD2 is a potential target of miR-130b. Based on these findings, we hypothesize that miR-130b might target NKD2 and regulate the Wnt signaling to promote OS growth. We detected the expression of miR-130b and NKD2 mRNA and protein by quantitative Real-Time PCR (qRT-PCR) and western blot assays, respectively, and found up-regulation of miR-130b and down-regulation of NKD2 mRNA and protein exist in OS cell lines. MTT and flow cytometry assays showed that miR-130b inhibitors inhibit proliferation and promote apoptosis in OS cells. Furthermore, we showed that NKD2 is a direct target of miR-130b, and miR-130b regulated proliferation and apoptosis of OS cells by targeting NKD2. We further investigated whether miR-130b and NKD2 regulate OS cell proliferation and apoptosis by inhibiting Wnt signaling, and the results confirmed our speculation that miR-130b targets NKD2 and regulates the Wnt signaling to promote proliferation and inhibit apoptosis of OS cells. These findings will offer new clues for OS development and progression, and novel potential therapeutic targets for OS. - Highlights: • miR-130b is up-regulated and NKD2 is down-regulated in osteosarcoma cell lines. • Down-regulation of miR-130b inhibits proliferation of osteosarcoma cells. • Down-regulation of miR-130b promotes apoptosis of osteosarcoma cells. • miR-130b directly targets NKD2. • NKD2 regulates OS cell proliferation and apoptosis by inhibiting the Wnt signaling.

  9. Striated muscle activator of Rho signalling (STARS) is reduced in ageing human skeletal muscle and targeted by miR-628-5p.

    Science.gov (United States)

    Russell, A P; Wallace, M A; Kalanon, M; Zacharewicz, E; Della Gatta, P A; Garnham, A; Lamon, S

    2017-06-01

    The striated muscle activator of Rho signalling (STARS) is a muscle-specific actin-binding protein. The STARS signalling pathway is activated by resistance exercise and is anticipated to play a role in signal mechanotransduction. Animal studies have reported a negative regulation of STARS signalling with age, but such regulation has not been investigated in humans. Ten young (18-30 years) and 10 older (60-75 years) subjects completed an acute bout of resistance exercise. Gene and protein expression of members of the STARS signalling pathway and miRNA expression of a subset of miRNAs, predicted or known to target members of STARS signalling pathway, were measured in muscle biopsies collected pre-exercise and 2 h post-exercise. For the first time, we report a significant downregulation of the STARS protein in older subjects. However, there was no effect of age on the magnitude of STARS activation in response to an acute bout of exercise. Finally, we established that miR-628-5p, a miRNA regulated by age and exercise, binds to the STARS 3'UTR to directly downregulate its transcription. This study describes for the first time the resistance exercise-induced regulation of STARS signalling in skeletal muscle from older humans and identifies a new miRNA involved in the transcriptional control of STARS. © 2016 The Authors. Acta Physiologica published by John Wiley & Sons Ltd on behalf of Scandinavian Physiological Society.

  10. Kinase activity profiling reveals active signal transduction pathways in pediatric acute lymphoblastic leukemia : A new approach for target discovery

    NARCIS (Netherlands)

    van der Sligte, Naomi E.; Scherpen, Frank J. G.; Meeuwsen-de Boer, Tiny G. J.; Lourens, Harm Jan; ter Elst, Arja; Diks, Sander H.; Guryev, Victor; Peppelenbosch, Maikel P.; van Leeuwen, Frank N.; de Bont, Eveline S. J. M.

    Still about 20% of patients with acute lymphoblastic leukemia (ALL) struggle with relapse, despite intensive chemotherapy. We and others have shown that kinase activity profiling is able to give more insights in active signal transduction pathways and point out interesting signaling hubs as well as

  11. Kinase activity profiling reveals active signal transduction pathways in pediatric acute lymphoblastic leukemia: A new approach for target discovery

    NARCIS (Netherlands)

    Sligte, N.E. van der; Scherpen, F.J.; Meeuwsen-de Boer, T.G.; Lourens, H.J.; Elst, A. Ter; Diks, S.H.; Guryev, V.; Peppelenbosch, M.P.; Leeuwen, F.N. van; Bont, E.S. de

    2015-01-01

    Still about 20% of patients with acute lymphoblastic leukemia (ALL) struggle with relapse, despite intensive chemotherapy. We and others have shown that kinase activity profiling is able to give more insights in active signal transduction pathways and point out interesting signaling hubs as well as

  12. Intermittent compressive stress regulates Notch target gene expression via transforming growth factor-β signaling in murine pre-osteoblast cell line.

    Science.gov (United States)

    Manokawinchoke, Jeeranan; Pavasant, Prasit; Osathanon, Thanaphum

    2017-10-01

    Different mechanical stimuli regulate behaviors of various cell types, including osteoblasts, osteocytes, and periodontal ligament fibroblasts. Notch signaling participates in the mechanical stress-regulated cell responses. The present study investigated the regulation of Notch target gene and sclerostin (Sost) expression in murine pre-osteoblast cell line (MC3T3-E1) under intermittent compressive stress. MC3T3-E1 were subjected to the intermittent compressive force under the computerized controlled machine. In some experiments, cells were pretreated with chemical inhibitors for Notch and transforming growth factor (TGF)-β signaling prior to mechanical stimuli. To evaluate role of Notch signaling in MC3T3-E1 cells under unloaded condition, cells were seeded on indirect immobilized Notch ligand (Jagged1). Gene expression was determined using real-time quantitative polymerase chain reaction. The intermittent compressive stress significantly upregulated Notch target gene expression (Hes Family BHLH transcription factor 1; Hes1 and Hairy/enhancer-of-split related with YRPW motif protein1; Hey1). The intermittent stress-induced Hes1 and Hey1 mRNA expression could be inhibited by a γ-secretase inhibitor (DAPT) or a TGF-β superfamily type I activing receptor-like kinase receptors inhibitor (SB431542). The results imply that intermittent compressive stress regulates Notch signaling via TGF-β pathway. Further, the intermittent compressive stress reduced Sost mRNA expression and this phenomenon could be rescued by a DAPT pretreatment, implying the involvement of Notch signaling. However, activation of Notch signaling under the unloaded condition resulted in the increase of Sost expression and the reduction of osteogenic marker genes. These results imply the involvement of Notch signaling in the homeostasis maintaining of osteogenic cells under mechanical stress stimuli. Copyright © 2017 Elsevier Ltd. All rights reserved.

  13. Agrin-LRP4-MuSK signaling as a therapeutic target for myasthenia gravis and other neuromuscular disorders.

    Science.gov (United States)

    Ohno, Kinji; Ohkawara, Bisei; Ito, Mikako

    2017-10-01

    Signal transduction at the neuromuscular junction (NMJ) is compromised in a diverse array of diseases including myasthenia gravis, Lambert-Eaton myasthenic syndrome, Isaacs' syndrome, congenital myasthenic syndromes, Fukuyama-type congenital muscular dystrophy, amyotrophic lateral sclerosis, and sarcopenia. Except for sarcopenia, all are orphan diseases. In addition, the NMJ signal transduction is impaired by tetanus, botulinum, curare, α-bungarotoxin, conotoxins, organophosphate, sarin, VX, and soman to name a few. Areas covered: This review covers the agrin-LRP4-MuSK signaling pathway, which drives clustering of acetylcholine receptors (AChRs) and ensures efficient signal transduction at the NMJ. We also address diseases caused by autoantibodies against the NMJ molecules and by germline mutations in genes encoding the NMJ molecules. Expert opinion: Representative small compounds to treat the defective NMJ signal transduction are cholinesterase inhibitors, which exert their effects by increasing the amount of acetylcholine at the synaptic space. Another possible therapeutic strategy to enhance the NMJ signal transduction is to increase the number of AChRs, but no currently available drug has this functionality.

  14. Subcellular Localization of a Plant Catalase-Phenol Oxidase, AcCATPO, from Amaranthus and Identification of a Non-canonical Peroxisome Targeting Signal.

    Science.gov (United States)

    Chen, Ning; Teng, Xiao-Lu; Xiao, Xing-Guo

    2017-01-01

    AcCATPO is a plant catalase-phenol oxidase recently identified from red amaranth. Its physiological function remains unexplored. As the starting step of functional analysis, here we report its subcellular localization and a non-canonical targeting signal. Commonly used bioinformatics programs predicted a peroxisomal localization for AcCATPO, but failed in identification of canonical peroxisomal targeting signals (PTS). The C-terminal GFP tagging led the fusion protein AcCATPO-GFP to the cytosol and the nucleus, but N-terminal tagging directed the GFP-AcCATPO to peroxisomes and nuclei, in transgenic tobacco. Deleting the tripeptide (PTM) at the extreme C-terminus almost ruled out the peroxisomal localization of GFP-AcCATPOΔ3, and removing the C-terminal decapeptide completely excluded peroxisomes as the residence of GFP-AcCATPOΔ10. Furthermore, this decapeptide as a targeting signal could import GFP-10aa to the peroxisome exclusively. Taken together, these results demonstrate that AcCATPO is localized to the peroxisome and the nucleus, and its peroxisomal localization is attributed to a non-canonical PTS1, the C-terminal decapeptide which contains an internal SRL motif and a conserved tripeptide P-S/T-I/M at the extreme of C-terminus. This work may further the study as to the physiological function of AcCATPO, especially clarify its involvement in betalain biosynthesis, and provide a clue to elucidate more non-canonic PTS.

  15. Subcellular Localization of a Plant Catalase-Phenol Oxidase, AcCATPO, from Amaranthus and Identification of a Non-canonical Peroxisome Targeting Signal

    Science.gov (United States)

    Chen, Ning; Teng, Xiao-Lu; Xiao, Xing-Guo

    2017-01-01

    AcCATPO is a plant catalase-phenol oxidase recently identified from red amaranth. Its physiological function remains unexplored. As the starting step of functional analysis, here we report its subcellular localization and a non-canonical targeting signal. Commonly used bioinformatics programs predicted a peroxisomal localization for AcCATPO, but failed in identification of canonical peroxisomal targeting signals (PTS). The C-terminal GFP tagging led the fusion protein AcCATPO-GFP to the cytosol and the nucleus, but N-terminal tagging directed the GFP-AcCATPO to peroxisomes and nuclei, in transgenic tobacco. Deleting the tripeptide (PTM) at the extreme C-terminus almost ruled out the peroxisomal localization of GFP-AcCATPOΔ3, and removing the C-terminal decapeptide completely excluded peroxisomes as the residence of GFP-AcCATPOΔ10. Furthermore, this decapeptide as a targeting signal could import GFP-10aa to the peroxisome exclusively. Taken together, these results demonstrate that AcCATPO is localized to the peroxisome and the nucleus, and its peroxisomal localization is attributed to a non-canonical PTS1, the C-terminal decapeptide which contains an internal SRL motif and a conserved tripeptide P-S/T-I/M at the extreme of C-terminus. This work may further the study as to the physiological function of AcCATPO, especially clarify its involvement in betalain biosynthesis, and provide a clue to elucidate more non-canonic PTS. PMID:28824680

  16. Subcellular Localization of a Plant Catalase-Phenol Oxidase, AcCATPO, from Amaranthus and Identification of a Non-canonical Peroxisome Targeting Signal

    Directory of Open Access Journals (Sweden)

    Ning Chen

    2017-08-01

    Full Text Available AcCATPO is a plant catalase-phenol oxidase recently identified from red amaranth. Its physiological function remains unexplored. As the starting step of functional analysis, here we report its subcellular localization and a non-canonical targeting signal. Commonly used bioinformatics programs predicted a peroxisomal localization for AcCATPO, but failed in identification of canonical peroxisomal targeting signals (PTS. The C-terminal GFP tagging led the fusion protein AcCATPO-GFP to the cytosol and the nucleus, but N-terminal tagging directed the GFP-AcCATPO to peroxisomes and nuclei, in transgenic tobacco. Deleting the tripeptide (PTM at the extreme C-terminus almost ruled out the peroxisomal localization of GFP-AcCATPOΔ3, and removing the C-terminal decapeptide completely excluded peroxisomes as the residence of GFP-AcCATPOΔ10. Furthermore, this decapeptide as a targeting signal could import GFP-10aa to the peroxisome exclusively. Taken together, these results demonstrate that AcCATPO is localized to the peroxisome and the nucleus, and its peroxisomal localization is attributed to a non-canonical PTS1, the C-terminal decapeptide which contains an internal SRL motif and a conserved tripeptide P-S/T-I/M at the extreme of C-terminus. This work may further the study as to the physiological function of AcCATPO, especially clarify its involvement in betalain biosynthesis, and provide a clue to elucidate more non-canonic PTS.

  17. Targeting deregulated epigenetic control in cancer.

    Science.gov (United States)

    Zaidi, Sayyed K; Van Wijnen, Andre J; Lian, Jane B; Stein, Janet L; Stein, Gary S

    2013-11-01

    Cancer is a multifaceted disease that involves acquisition of genetic mutations, deletions, and amplifications as well as deregulation of epigenetic mechanisms that fine-tune gene regulation. Key epigenetic mechanisms that include histone modifications, DNA methylation, and non-coding RNA-mediated gene silencing are often deregulated in a variety of cancers. Subnuclear localization of key proteins in the interphase nucleus and bookmarking of genes by lineage commitment factors in mitosis-a new dimension to epigenetic control of fundamental biological processes-is also modified in cancer. In this review, we discuss the various aspects of epigenetic control that are operative in a variety of cancers and their potential for risk assessment, early detection, targeted therapy, and personalized medicine. Copyright © 2013 Wiley Periodicals, Inc.

  18. Klotho-beta overexpression as a novel target for suppressing proliferation and fibroblast growth factor receptor-4 signaling in hepatocellular carcinoma

    Directory of Open Access Journals (Sweden)

    Poh Weijie

    2012-03-01

    Full Text Available Abstract Background We had previously demonstrated overexpression of fibroblast growth factor receptor-4 (FGFR4 in hepatocellular carcinoma (HCC. However, additional molecular mechanisms resulting in amplified FGFR4 signaling in HCC remain under-studied. Here, we studied the mechanistic role of its co-receptor klotho-beta (KLB in driving elevated FGFR4 activity in HCC progression. Results Quantitative real-time PCR analysis identified frequent elevation of KLB gene expression in HCC tumors relative to matched non-tumor tissue, with a more than two-fold increase correlating with development of multiple tumors in patients. KLB-silencing in Huh7 cells decreased cell proliferation and suppressed FGFR4 downstream signaling. While transient repression of KLB-FGFR4 signaling decreased protein expression of alpha-fetoprotein (AFP, a HCC diagnostic marker, prolonged inhibition enriched for resistant HCC cells exhibiting increased liver stemness. Conclusions Elevated KLB expression in HCC tissues provides further credence to the oncogenic role of increased FGFR4 signaling in HCC progression and represents a novel biomarker to identify additional patients amenable to anti-FGFR4 therapy. The restricted tissue expression profile of KLB, together with the anti-proliferative effect observed with KLB-silencing, also qualifies it as a specific and potent therapeutic target for HCC patients. The enrichment of a liver stem cell-like population in response to extended KLB-FGFR4 repression necessitates further investigation to target the development of drug resistance.

  19. Direct Targeting of β-Catenin by a Small Molecule Stimulates Proteasomal Degradation and Suppresses Oncogenic Wnt/β-Catenin Signaling

    Directory of Open Access Journals (Sweden)

    So-Young Hwang

    2016-06-01

    Full Text Available The Wnt/β-catenin signaling pathway plays a major role in tissue homeostasis, and its dysregulation can lead to various human diseases. Aberrant activation of β-catenin is oncogenic and is a critical driver in the development and progression of human cancers. Despite the significant potential of targeting the oncogenic β-catenin pathway for cancer therapy, the development of specific inhibitors remains insufficient. Using a T cell factor (TCF-dependent luciferase-reporter system, we screened for small-molecule compounds that act against Wnt/β-catenin signaling and identified MSAB (methyl 3-{[(4-methylphenylsulfonyl]amino}benzoate as a selective inhibitor of Wnt/β-catenin signaling. MSAB shows potent anti-tumor effects selectively on Wnt-dependent cancer cells in vitro and in mouse cancer models. MSAB binds to β-catenin, promoting its degradation, and specifically downregulates Wnt/β-catenin target genes. Our findings might represent an effective therapeutic strategy for cancers addicted to the Wnt/β-catenin signaling pathway.

  20. Evasion of anti-growth signaling: A key step in tumorigenesis and potential target for treatment and prophylaxis by natural compounds.

    Science.gov (United States)

    Amin, A R M Ruhul; Karpowicz, Phillip A; Carey, Thomas E; Arbiser, Jack; Nahta, Rita; Chen, Zhuo G; Dong, Jin-Tang; Kucuk, Omer; Khan, Gazala N; Huang, Gloria S; Mi, Shijun; Lee, Ho-Young; Reichrath, Joerg; Honoki, Kanya; Georgakilas, Alexandros G; Amedei, Amedeo; Amin, Amr; Helferich, Bill; Boosani, Chandra S; Ciriolo, Maria Rosa; Chen, Sophie; Mohammed, Sulma I; Azmi, Asfar S; Keith, W Nicol; Bhakta, Dipita; Halicka, Dorota; Niccolai, Elena; Fujii, Hiromasa; Aquilano, Katia; Ashraf, S Salman; Nowsheen, Somaira; Yang, Xujuan; Bilsland, Alan; Shin, Dong M

    2015-12-01

    The evasion of anti-growth signaling is an important characteristic of cancer cells. In order to continue to proliferate, cancer cells must somehow uncouple themselves from the many signals that exist to slow down cell growth. Here, we define the anti-growth signaling process, and review several important pathways involved in growth signaling: p53, phosphatase and tensin homolog (PTEN), retinoblastoma protein (Rb), Hippo, growth differentiation factor 15 (GDF15), AT-rich interactive domain 1A (ARID1A), Notch, insulin-like growth factor (IGF), and Krüppel-like factor 5 (KLF5) pathways. Aberrations in these processes in cancer cells involve mutations and thus the suppression of genes that prevent growth, as well as mutation and activation of genes involved in driving cell growth. Using these pathways as examples, we prioritize molecular targets that might be leveraged to promote anti-growth signaling in cancer cells. Interestingly, naturally occurring phytochemicals found in human diets (either singly or as mixtures) may promote anti-growth signaling, and do so without the potentially adverse effects associated with synthetic chemicals. We review examples of naturally occurring phytochemicals that may be applied to prevent cancer by antagonizing growth signaling, and propose one phytochemical for each pathway. These are: epigallocatechin-3-gallate (EGCG) for the Rb pathway, luteolin for p53, curcumin for PTEN, porphyrins for Hippo, genistein for GDF15, resveratrol for ARID1A, withaferin A for Notch and diguelin for the IGF1-receptor pathway. The coordination of anti-growth signaling and natural compound studies will provide insight into the future application of these compounds in the clinical setting. Copyright © 2015 Elsevier Ltd. All rights reserved.

  1. Transcriptomic-anatomic analysis of the mouse habenula uncovers a high molecular heterogeneity among neurons in the lateral complex, while gene expression in the medial complex largely obeys subnuclear boundaries.

    Science.gov (United States)

    Wagner, Franziska; French, Leon; Veh, Rüdiger W

    2016-01-01

    The mammalian habenula with its medial and lateral complexes has gained much interest in recent years, while knowledge on the detailed biological functions of these nuclei is still scarce. Novel strategies to differentiate and identify habenular cell types are required. Such attempts have largely failed, most likely due to the lack of appropriate molecular markers. One important tool to approach this dilemma is available in form of the Allen Brain Atlas (ABA), which provides detailed expression patterns of many genes in the mouse brain. In the present report, ABA tools in combination with visual inspection of ISH images were used to detect transcripts, which are strongly expressed in medial (MHb) and lateral (LHb) habenular complexes. Against our expectations, most transcripts were differentially distributed throughout the LHb, disregarding boundaries of subnuclear areas. Nine distinct distribution patterns were recognized. Yet, several transcripts could not be attributed to one of these, suggesting a high diversity of neuron types in the LHb. In the MHb, in contrast, many transcripts tended to obey subnuclear boundaries. The differential distribution of others like Adcyap1, Chrna3, or Trp53i11 suggests the presence of a novel subfield adjacent to the region of the MHbVm, which now is termed intermediate field of the ventral MHb. In addition, the localizations of Amigo2, Adcyap1, and a couple of other transcripts suggest a lateral extension of the MHb, which is here, termed HbX area. Apparently, this area is composed of intermingled MHb and LHb neurons and may allow functional interaction between the both habenular complexes.

  2. Targeting of pro-apoptotic TLR adaptor SARM to mitochondria: definition of the critical region and residues in the signal sequence.

    Science.gov (United States)

    Panneerselvam, Porkodi; Singh, Laishram Pradeepkumar; Ho, Bow; Chen, Jianzhu; Ding, Jeak Ling

    2012-03-01

    The fifth and the most well-conserved member of the TLR (Toll-like receptor) adaptor, SARM (sterile α- and HEAT/armadillo-motif-containing protein), has been reported to be an important mediator of apoptosis. However, the exact cellular localization of SARM with respect to its role is unclear. In the present study we show that SARM specifically co-localizes with mitochondria. Endogenous SARM is mainly found in the mitochondria. We demonstrate that the N-terminal 27 amino acids (S27) of SARM, which is hydrophobic and polybasic, acts as a mitochondria-targeting signal sequence, associating SARM to the mitochondria. The S27 peptide has an inherent ability to bind to lipids and mitochondria. This sequence effectively translocates the soluble EGFP (enhanced green fluorescence protein) reporter into the mitochondria. Positioning S27 downstream of the EGFP abrogates its mitochondria-targeting ability. Transmission electron microscopy confirms the ability of S27 to import EGFP into the mitochondria. Importantly, by mutagenesis study, we delineated the specificity of the mitochondria-targeting ability to the arginine residue at the 14th position. The R14A SARM mutant also showed reduced apoptotic potential when compared with the wild-type. Taken together, S27, which is a bona fide signal sequence that targets SARM to the mitochondria, explains the pro-apoptotic activity of SARM.

  3. Green tea polyphenol, (−)-epigallocatechin-3-gallate, induces toxicity in human skin cancer cells by targeting β-catenin signaling

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Tripti [Department of Dermatology, University of Alabama at Birmingham, Birmingham, AL 35294 (United States); Katiyar, Santosh K., E-mail: skatiyar@uab.edu [Department of Dermatology, University of Alabama at Birmingham, Birmingham, AL 35294 (United States); Comprehensive Cancer Center, University of Alabama at Birmingham, Birmingham, AL 35294 (United States); Birmingham Veterans Affairs Medical Center, Birmingham, AL 35233 (United States)

    2013-12-01

    The green tea polyphenol, (−)-epigallocatechin-3-gallate (EGCG), has been shown to have anti-carcinogenic effects in several skin tumor models, and efforts are continued to investigate the molecular targets responsible for its cytotoxic effects to cancer cells. Our recent observation that β-catenin is upregulated in skin tumors suggested the possibility that the anti-skin carcinogenic effects of EGCG are mediated, at least in part, through its effects on β-catenin signaling. We have found that treatment of the A431 and SCC13 human skin cancer cell lines with EGCG resulted in reduced cell viability and increased cell death and that these cytotoxic effects were associated with inactivation of β-catenin signaling. Evidence of EGCG-induced inactivation of β-catenin included: (i) reduced accumulation of nuclear β-catenin; (ii) enhanced levels of casein kinase1α, reduced phosphorylation of glycogen synthase kinase-3β, and increased phosphorylation of β-catenin on critical serine{sup 45,33/37} residues; and (iii) reduced levels of matrix metalloproteinase (MMP)-2 and MMP-9, which are down-stream targets of β-catenin. Treatment of cells with prostaglandin E2 (PGE{sub 2}) enhanced the accumulation of β-catenin and enhanced β-catenin signaling. Treatment with either EGCG or an EP2 antagonist (AH6809) reduced the PGE{sub 2}-enhanced levels of cAMP, an upstream regulator of β-catenin. Inactivation of β-catenin by EGCG resulted in suppression of cell survival signaling proteins. siRNA knockdown of β-catenin in A431 and SCC13 cells reduced cell viability. Collectively, these data suggest that induction of cytotoxicity in skin cancer cells by EGCG is mediated by targeting of β-catenin signaling and that the β-catenin signaling is upregulated by inflammatory mediators. - Highlights: • EGCG inhibits cancer cell viability through inactivation of β-catenin signaling. • Inactivation of β-catenin involves the downregulation of inflammatory mediators. • EGCG

  4. A method for predicting target drug efficiency in cancer based on the analysis of signaling pathway activation.

    Science.gov (United States)

    Artemov, Artem; Aliper, Alexander; Korzinkin, Michael; Lezhnina, Ksenia; Jellen, Leslie; Zhukov, Nikolay; Roumiantsev, Sergey; Gaifullin, Nurshat; Zhavoronkov, Alex; Borisov, Nicolas; Buzdin, Anton

    2015-10-06

    A new generation of anticancer therapeutics called target drugs has quickly developed in the 21st century. These drugs are tailored to inhibit cancer cell growth, proliferation, and viability by specific interactions with one or a few target proteins. However, despite formally known molecular targets for every "target" drug, patient response to treatment remains largely individual and unpredictable. Choosing the most effective personalized treatment remains a major challenge in oncology and is still largely trial and error. Here we present a novel approach for predicting target drug efficacy based on the gene expression signature of the individual tumor sample(s). The enclosed bioinformatic algorithm detects activation of intracellular regulatory pathways in the tumor in comparison to the corresponding normal tissues. According to the nature of the molecular targets of a drug, it predicts whether the drug can prevent cancer growth and survival in each individual case by blocking the abnormally activated tumor-promoting pathways or by reinforcing internal tumor suppressor cascades. To validate the method, we compared the distribution of predicted drug efficacy scores for five drugs (Sorafenib, Bevacizumab, Cetuximab, Sorafenib, Imatinib, Sunitinib) and seven cancer types (Clear Cell Renal Cell Carcinoma, Colon cancer, Lung adenocarcinoma, non-Hodgkin Lymphoma, Thyroid cancer and Sarcoma) with the available clinical trials data for the respective cancer types and drugs. The percent of responders to a drug treatment correlated significantly (Pearson's correlation 0.77 p = 0.023) with the percent of tumors showing high drug scores calculated with the current algorithm.

  5. Targeting the PDGF-B/PDGFR-β Interface with Destruxin A5 to Selectively Block PDGF-BB/PDGFR-ββ Signaling and Attenuate Liver Fibrosis

    Directory of Open Access Journals (Sweden)

    Xingqi Wang

    2016-05-01

    Full Text Available PDGF-BB/PDGFR-ββ signaling plays very crucial roles in the process of many diseases such as liver fibrosis. However, drug candidates with selective affinities for PDGF-B/PDGFR-β remain deficient. Here, we identified a natural cyclopeptide termed destruxin A5 that effectively inhibits PDGF-BB-induced PDGFR-β signaling. Interestingly and importantly, the inhibitory mechanism is distinct from the mechanism of tyrosine kinase inhibitors because destruxin A5 does not have the ability to bind to the ATP-binding pocket of PDGFR-β. Using Biacore T200 technology, thermal shift technology, microscale thermophoresis technology and computational analysis, we confirmed that destruxin A5 selectively targets the PDGF-B/PDGFR-β interaction interface to block this signaling. Additionally, the inhibitory effect of destruxin A5 on PDGF-BB/PDGFR-ββ signaling was verified using in vitro, ex vivo and in vivo models, in which the extent of liver fibrosis was effectively alleviated by destruxin A5. In summary, destruxin A5 may represent an efficacious and more selective inhibitor of PDGF-BB/PDGFR-ββ signaling.

  6. Analysis of Maneuvering Targets with Complex Motions by Two-Dimensional Product Modified Lv’s Distribution for Quadratic Frequency Modulation Signals

    Directory of Open Access Journals (Sweden)

    Fulong Jing

    2017-06-01

    Full Text Available For targets with complex motion, such as ships fluctuating with oceanic waves and high maneuvering airplanes, azimuth echo signals can be modeled as multicomponent quadratic frequency modulation (QFM signals after migration compensation and phase adjustment. For the QFM signal model, the chirp rate (CR and the quadratic chirp rate (QCR are two important physical quantities, which need to be estimated. For multicomponent QFM signals, the cross terms create a challenge for detection, which needs to be addressed. In this paper, by employing a novel multi-scale parametric symmetric self-correlation function (PSSF and modified scaled Fourier transform (mSFT, an effective parameter estimation algorithm is proposed—referred to as the Two-Dimensional product modified Lv’s distribution (2D-PMLVD—for QFM signals. The 2D-PMLVD is simple and can be easily implemented by using fast Fourier transform (FFT and complex multiplication. These measures are analyzed in the paper, including the principle, the cross term, anti-noise performance, and computational complexity. Compared to the other three representative methods, the 2D-PMLVD can achieve better anti-noise performance. The 2D-PMLVD, which is free of searching and has no identifiability problems, is more suitable for multicomponent situations. Through several simulations and analyses, the effectiveness of the proposed estimation algorithm is verified.

  7. 10-Hydroxy-2-decenoic Acid, the Major Lipid Component of Royal Jelly, Extends the Lifespan of Caenorhabditis elegans through Dietary Restriction and Target of Rapamycin Signaling

    Directory of Open Access Journals (Sweden)

    Yoko Honda

    2015-01-01

    Full Text Available Royal jelly (RJ produced by honeybees has been reported to possess diverse health-beneficial properties and has been implicated to have a function in longevity across diverse species as well as honeybees. 10-Hydroxy-2-decenoic acid (10-HDA, the major lipid component of RJ produced by honeybees, was previously shown to increase the lifespan of Caenorhabditis elegans. The objective of this study is to elucidate signaling pathways that are involved in the lifespan extension by 10-HDA. 10-HDA further extended the lifespan of the daf-2 mutants, which exhibit long lifespan through reducing insulin-like signaling (ILS, indicating that 10-HDA extended lifespan independently of ILS. On the other hand, 10-HDA did not extend the lifespan of the eat-2 mutants, which show long lifespan through dietary restriction caused by a food-intake defect. This finding indicates that 10-HDA extends lifespan through dietary restriction signaling. We further found that 10-HDA did not extend the lifespan of the long-lived mutants in daf-15, which encodes Raptor, a target of rapamycin (TOR components, indicating that 10-HDA shared some longevity control mechanisms with TOR signaling. Additionally, 10-HDA was found to confer tolerance against thermal and oxidative stress. 10-HDA increases longevity not through ILS but through dietary restriction and TOR signaling in C. elegans.

  8. Cortical axons, isolated in channels, display activity-dependent signal modulation as a result of targeted stimulation

    Directory of Open Access Journals (Sweden)

    Marta K. Lewandowska

    2016-03-01

    Full Text Available Mammalian cortical axons are extremely thin processes that are difficult to study as a result of their small diameter: they are too narrow to patch while intact, and super-resolution microscopy is needed to resolve single axons. We present a method for studying axonal physiology by pairing a high-density microelectrode array with a microfluidic axonal isolation device, and use it to study activity-dependent modulation of axonal signal propagation evoked by stimulation near the soma. Up to three axonal branches from a single neuron, isolated in different channels, were recorded from simultaneously using 10-20 electrodes per channel. The axonal channels amplified spikes such that propagations of individual signals along tens of electrodes could easily be discerned with high signal to noise. Stimulation from 10 Hz up to 160 Hz demonstrated similar qualitative results from all of the cells studied: extracellular action potential characteristics changed drastically in response to stimulation. Spike height decreased, spike width increased, and latency increased, as a result of reduced propagation velocity, as the number of stimulations and the stimulation frequencies increased. Quantitatively, the strength of these changes manifested itself differently in cells at different frequencies of stimulation. Some cells’ signal fidelity fell to 80% already at 10 Hz, while others maintained 80% signal fidelity at 80 Hz. Differences in modulation by axonal branches of the same cell were also seen for many different stimulation frequencies, starting at 10 Hz. Potassium ion concentration changes altered the behavior of the cells causing propagation failures at lower concentrations and improving signal fidelity at higher concentrations.

  9. Macrophages are targets of retinoic acid signaling during the wound-healing process after thulium laser resection of the prostate

    OpenAIRE

    Yu, Dian-Jun; Wang, Xing-Jie; Shi, Yun-Feng; Jiang, Chen-Yi; Zhao, Rui-Zhe; Zhu, Yi-Ping; Chen, Li; Yang, Yuan-Qing; Sun, Xiao-Wen; Xia, Shu-Jie

    2017-01-01

    BACKGROUND The wound-healing process is very important for reducing complications after thulium laser resection of the prostate (TmLRP). The retinoic acid (RA) signaling pathway has been well studied in the wound-healing process of the skin and other organs. The goals of this study were to identify the role of RA signaling in the repair of the prostate after TmLRP and to investigate the molecular mechanism of this process. RESULTS Retinoic acid receptors (RARs) were present in the prostate, a...

  10. microRNA miR-34a regulates cytodifferentiation and targets multi-signaling pathways in human dental papilla cells.

    Directory of Open Access Journals (Sweden)

    Mian Wan

    Full Text Available Odontogenesis relies on the reciprocal signaling interactions between dental epithelium and neural crest-derived mesenchyme, which is regulated by several signaling pathways. Subtle changes in the activity of these major signaling pathways can have dramatic effects on tooth development. An important regulator of such subtle changes is the fine tuning function of microRNAs (miRNAs. However, the underlying mechanism by which miRNAs regulate tooth development remains elusive. This study determined the expression of miRNAs during cytodifferentiation in the human tooth germ and studied miR-34a as a regulator of dental papilla cell differentiation. Using microarrays, miRNA expression profiles were established at selected times during development (early bell stage or late bell stage of the human fetal tooth germ. We identified 29 differentially expressed miRNAs from early bell stage/late bell stage comparisons. Out of 6 miRNAs selected for validation by qPCR, all transcripts were confirmed to be differentially expressed. miR-34a was selected for further investigation because it has been previously reported to regulate organogenesis. miR-34a mimics and inhibitors were transfected into human fetal dental papilla cells, mRNA levels of predicted target genes were detected by quantitative real-time PCR, and levels of putative target proteins were examined by western blotting. ALP and DSPP expression were also tested by qPCR, western blotting, and immunofluorescence. Findings from these studies suggested that miR-34a may play important roles in dental papilla cell differentiation during human tooth development by targeting NOTCH and TGF-beta signaling.

  11. microRNA miR-34a Regulates Cytodifferentiation and Targets Multi-signaling Pathways in Human Dental Papilla Cells

    Science.gov (United States)

    Sun, Feifei; Tang, Yin; Ye, Ling; Fan, Yi; Klein, Ophir D.; Zhou, Xuedong; Zheng, Liwei

    2012-01-01

    Odontogenesis relies on the reciprocal signaling interactions between dental epithelium and neural crest-derived mesenchyme, which is regulated by several signaling pathways. Subtle changes in the activity of these major signaling pathways can have dramatic effects on tooth development. An important regulator of such subtle changes is the fine tuning function of microRNAs (miRNAs). However, the underlying mechanism by which miRNAs regulate tooth development remains elusive. This study determined the expression of miRNAs during cytodifferentiation in the human tooth germ and studied miR-34a as a regulator of dental papilla cell differentiation. Using microarrays, miRNA expression profiles were established at selected times during development (early bell stage or late bell stage) of the human fetal tooth germ. We identified 29 differentially expressed miRNAs from early bell stage/late bell stage comparisons. Out of 6 miRNAs selected for validation by qPCR, all transcripts were confirmed to be differentially expressed. miR-34a was selected for further investigation because it has been previously reported to regulate organogenesis. miR-34a mimics and inhibitors were transfected into human fetal dental papilla cells, mRNA levels of predicted target genes were detected by quantitative real-time PCR, and levels of putative target proteins were examined by western blotting. ALP and DSPP expression were also tested by qPCR, western blotting, and immunofluorescence. Findings from these studies suggested that miR-34a may play important roles in dental papilla cell differentiation during human tooth development by targeting NOTCH and TGF-beta signaling. PMID:23226240

  12. The Triangle of Death in Alzheimer's Disease Brain: The Aberrant Cross-Talk Among Energy Metabolism, Mammalian Target of Rapamycin Signaling, and Protein Homeostasis Revealed by Redox Proteomics.

    Science.gov (United States)

    Di Domenico, Fabio; Barone, Eugenio; Perluigi, Marzia; Butterfield, D Allan

    2017-03-10

    Alzheimer's disease (AD) is a multifactorial neurodegenerative disorder and represents one of the most disabling conditions. AD shares many features in common with systemic insulin resistance diseases, suggesting that it can be considered as a metabolic disease, characterized by reduced insulin-stimulated growth and survival signaling, increased oxidative stress (OS), proinflammatory cytokine activation, mitochondrial dysfunction, impaired energy metabolism, and altered protein homeostasis. Recent Advances: Reduced glucose utilization and energy metabolism in AD have been associated with the buildup of amyloid-β peptide and hyperphosphorylated tau, increased OS, and the accumulation of unfolded/misfolded proteins. Mammalian target of rapamycin (mTOR), which is aberrantly activated in AD since early stages, plays a key role during AD neurodegeneration by, on one side, inhibiting insulin signaling as a negative feedback mechanism and, on the other side, regulating protein homeostasis (synthesis/clearance). It is likely that the concomitant and mutual alterations of energy metabolism-mTOR signaling-protein homeostasis might represent a self-sustaining triangle of harmful events that trigger the degeneration and death of neurons and the development and progression of AD. Intriguingly, the altered cross-talk between the components of such a triangle of death, beyond altering the redox homeostasis of the neuron, is further exacerbated by increased levels of OS that target and impair key components of the pathways involved. Redox proteomic studies in human samples and animal models of AD-like dementia led to identification of oxidatively modified components of the pathways composing the triangle of death, therefore revealing the crucial role of OS in fueling this aberrant vicious cycle. The identification of compounds able to restore the function of the pathways targeted by oxidative damage might represent a valuable therapeutic approach to slow or delay AD. Antioxid

  13. Target-induced formation of gold amalgamation on DNA-based sensing platform for electrochemical monitoring of mercury ion coupling with cycling signal amplification strategy

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Jinfeng; Tang, Juan; Zhou, Jun; Zhang, Lan; Chen, Guonan; Tang, Dianping, E-mail: dianping.tang@fzu.edu.cn

    2014-01-31

    Graphical abstract: -- Highlights: •We report a new electrochemical sensing protocol for the detection of mercury ion. •Gold amalgamation on DNA-based sensing platform was used as nanocatalyst. •The signal was amplified by cycling signal amplification strategy. -- Abstract: Heavy metal ion pollution poses severe risks in human health and environmental pollutant, because of the likelihood of bioaccumulation and toxicity. Driven by the requirement to monitor trace-level mercury ion (Hg{sup 2+}), herein we construct a new DNA-based sensor for sensitive electrochemical monitoring of Hg{sup 2+} by coupling target-induced formation of gold amalgamation on DNA-based sensing platform with gold amalgamation-catalyzed cycling signal amplification strategy. The sensor was simply prepared by covalent conjugation of aminated poly-T{sub (25)} oligonucleotide onto the glassy carbon electrode by typical carbodiimide coupling. Upon introduction of target analyte, Hg{sup 2+} ion was intercalated into the DNA polyion complex membrane based on T–Hg{sup 2+}–T coordination chemistry. The chelated Hg{sup 2+} ion could induce the formation of gold amalgamation, which could catalyze the p-nitrophenol with the aid of NaBH{sub 4} and Ru(NH{sub 3}){sub 6}{sup 3+} for cycling signal amplification. Experimental results indicated that the electronic signal of our system increased with the increasing Hg{sup 2+} level in the sample, and has a detection limit of 0.02 nM with a dynamic range of up to 1000 nM Hg{sup 2+}. The strategy afforded exquisite selectivity for Hg{sup 2+} against other environmentally related metal ions. In addition, the methodology was evaluated for the analysis of Hg{sup 2+} in spiked tap-water samples, and the recovery was 87.9–113.8%.

  14. A protein kinase target of a PDK1 signalling pathway is involved in root hair growth in Arabidopsis

    NARCIS (Netherlands)

    Anthony, R.G.; Henriques, R.; Helfer, A.; Mészáros, T.; Rios, G.; Testerink, C.; Munnik, T.; Deák, M.; Koncz, C.; Bögre, L.

    2004-01-01

    Here we report on a lipid-signalling pathway in plants that is downstream of phosphatidic acid and involves the Arabidopsis protein kinase, AGC2-1, regulated by the 3'-phosphoinositide-dependent kinase-1 (AtPDK1). AGC2-1 specifically interacts with AtPDK1 through a conserved C-terminal hydrophobic

  15. A phylogenetic mosaic plastid proteome and unusual plastid-targeting signals in the green-colored dinoflagellate Lepidodinium chlorophorum

    Directory of Open Access Journals (Sweden)

    Inagaki Yuji

    2010-06-01

    Full Text Available Abstract Background Plastid replacements through secondary endosymbioses include massive transfer of genes from the endosymbiont to the host nucleus and require a new targeting system to enable transport of the plastid-targeted proteins across 3-4 plastid membranes. The dinoflagellates are the only eukaryotic lineage that has been shown to have undergone several plastid replacement events, and this group is thus highly relevant for studying the processes involved in plastid evolution. In this study, we analyzed the phylogenetic origin and N-terminal extensions of plastid-targeted proteins from Lepidodinium chlorophorum, a member of the only dinoflagellate genus that harbors a green secondary plastid rather than the red algal-derived, peridinin-containing plastid usually found in photosynthetic dinoflagellates. Results We sequenced 4,746 randomly picked clones from a L. chlorophorum cDNA library. 22 of the assembled genes were identified as genes encoding proteins functioning in plastids. Some of these were of green algal origin. This confirms that genes have been transferred from the plastid to the host nucleus of L. chlorophorum and indicates that the plastid is fully integrated as an organelle in the host. Other nuclear-encoded plastid-targeted protein genes, however, are clearly not of green algal origin, but have been derived from a number of different algal groups, including dinoflagellates, streptophytes, heterokonts, and red algae. The characteristics of N-terminal plastid-targeting peptides of all of these genes are substantially different from those found in peridinin-containing dinoflagellates and green algae. Conclusions L. chlorophorum expresses plastid-targeted proteins with a range of different origins, which probably arose through endosymbiotic gene transfer (EGT and horizontal gene transfer (HGT. The N-terminal extension of the genes is different from the extensions found in green alga and other dinoflagellates (peridinin- and

  16. Targeting Mortalin by Embelin Causes Activation of Tumor Suppressor p53 and Deactivation of Metastatic Signaling in Human Breast Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Nupur Nigam

    Full Text Available Embelin, a natural quinone found in the fruits of Embelia ribes, is commonly used in Ayurvedic home medicine for a variety of therapeutic potentials including anti-inflammation, anti-fever, anti-bacteria and anti-cancer. Molecular mechanisms of these activities and cellular targets have not been clarified to-date. We demonstrate that the embelin inhibits mortalin-p53 interactions, and activates p53 protein in tumor cells. We provide bioinformatics, molecular docking and experimental evidence to the binding affinity of embelin with mortalin and p53. Binding of embelin with mortalin/p53 abrogates their complex resulted in nuclear translocation and transcriptional activation function of p53 causing growth arrest in cancer cells. Furthermore, analyses of growth factors and metastatic signaling using antibody membrane array revealed their downregulation in embelin-treated cells. We also found that the embelin causes transcriptional attenuation of mortalin and several other proteins involved in metastatic signaling in cancer cells. Based on these molecular dynamics and experimental data, it is concluded that the anticancer activity of embelin involves targeting of mortalin, activation of p53 and inactivation of metastatic signaling.

  17. Target Area Extension in Synthetic Aperture Array Signal Processing for High-Frame-Rate Estimate of Two-Dimensional Motion Vector In vivo

    Science.gov (United States)

    Yagi, Shin-ichi; Yokoyama, Ryouta; Tamura, Kiyoshi; Sato, Masakazu

    2011-07-01

    A strategic synthetic aperture radar (SAR) along a flight path has been developed including a potential extensibility in a wide-range target area and an excellent spatial resolution by utilizing two-way range stacking, matched filtering, and chirp signal transmission. For the simultaneous ultrahigh-frame-rate ultrasonic imaging of microdynamics in a living tissue, a one-way synthetic aperture array processing of a real-time-received two-dimensional (2D) echo signal followed by a successive transmission is indispensable, in which the range stacking in SAR should be modified toward the pulsed ultrasonic irradiation generated by the array transducer. Therefore, the modification of the range stacking was proposed for pulsed radiation from a flexible point ultrasonic source. Firstly, a one-way receiving range stacking algorithm was described in a spatiotemporal frequency domain, and it was consequently extended to account for the forward-range- and cross-range-dependent time delay of the 2D echo signal in each range bin for the reconstruction of the target area. The overall system performance for the linear array transducer having 256 elements with a 3.0 MHz center frequency and a 0.25 mm pitch was verified for the reconstructed images in a numerical simulation and a hardware experiment.

  18. Role of adenosine 5'-monophosphate-activated protein kinase subunits in skeletal muscle mammalian target of rapamycin signaling

    DEFF Research Database (Denmark)

    Deshmukh, Atul S.; Treebak, Jonas Thue; Long, Yun Chau

    2008-01-01

    AMP-activated protein kinase (AMPK) is an important energy-sensing protein in skeletal muscle. Mammalian target of rapamycin (mTOR) mediates translation initiation and protein synthesis through ribosomal S6 kinase 1 (S6K1) and eukaryotic initiation factor 4E-binding protein 1 (4E-BP1). AMPK activ...

  19. Targeting Intracellular Calcium Signaling ([Ca2+]i to Overcome Acquired Multidrug Resistance of Cancer Cells: A Mini-Overview

    Directory of Open Access Journals (Sweden)

    Dietrich Büsselberg

    2017-05-01

    Full Text Available Cancer is a main public health problem all over the world. It affects millions of humans no matter their age, gender, education, or social status. Although chemotherapy is the main strategy for the treatment of cancer, a major problem limiting its success is the intrinsic or acquired drug resistance. Therefore, cancer drug resistance is a major impediment in medical oncology resulting in a failure of a successful cancer treatment. This mini-overview focuses on the interdependent relationship between intracellular calcium ([Ca2+]i signaling and multidrug resistance of cancer cells, acquired upon treatment of tumors with anticancer drugs. We propose that [Ca2+]i signaling modulates gene expression of multidrug resistant (MDR genes which in turn can be modulated by epigenetic factors which in turn leads to modified protein expression in drug resistant tumor cells. A precise knowledge of these mechanisms will help to develop new therapeutic strategies for drug resistant tumors and will improve current chemotherapy.

  20. Transforming growth factor beta signal transduction: a potential target for maintenance/restoration of transparency of the cornea.

    Science.gov (United States)

    Saika, Shizuya; Yamanaka, Osamu; Sumioka, Takayoshi; Okada, Yuka; Miyamoto, Takeshi; Shirai, Kumi; Kitano, Ai; Tanaka, Sai-ichi

    2010-09-01

    Maintenance of the transparency and regular shape of the cornea are essential to the normal vision, whereas opacification of the tissue impairs vision. Fibrogenic reaction leading to scarring in an injured cornea is characterized by appearance of myofibroblasts, the key player of the fibrogenic reaction, and excess accumulation of fibrous extracellular matrix. Inflammatory/fibrogenic growth factors/cytokines produced by inflammatory cells play a pivotal role in fibrogenic response. Signaling systems involved in myofibroblast formation and fibrogenesis are activated by various growth factors, i.e., transforming growth factor beta or others. Modulation of transforming growth factor beta signal transduction molecules, e.g., Smad and mitogen-activated protein kinases, by gene transfer and other technology provides a new concept of prevention/treatment of unfavorable fibrogenesis in the cornea.

  1. The anabolic steroid methandienone targets the hypothalamic-pituitary-testicular axis and myostatin signaling in a rat training model.

    Science.gov (United States)

    Mosler, Stephanie; Pankratz, Carlos; Seyfried, Alexis; Piechotta, Marion; Diel, Patrick

    2012-01-01

    There is increasing evidence that the biological activity of myostatin (MSTN), a negative regulator of muscle growth, is affected by training but also anabolic steroids. In this study, we analyzed the effects of the frequently abused anabolic steroid methandienone (Md) on the hypothalamic-pituitary-testicular axis and androgen-sensitive tissues in intact rats performing a treadmill training to simulate the situation of abusing athletes. The anabolic effects were correlated with the expression of members of the MSTN signaling cascade. Md treatment resulted in a significant stimulation of anabolic activity of the levator ani muscle, which was further increased by training, while prostate and seminal vesicle weights decreased in conformance with hormone concentrations of LH and testosterone. In gastrocnemius muscle, mRNA expression of genes of the MSTN signaling cascade (MSTN, Smad7 and MyoD) was reduced by training but not after Md treatment, in soleus muscle MSTN and its inhibitors, follistatin (FLST) and Smad-7 were only affected after training in combination with Md treatment. In summary, our data demonstrate that Md treatment of intact rats results in anabolic effects which are enhanced in combination with physical activity. Interestingly, the anabolic activity on the levator ani was increased in combination with training, although the levator ani muscle was not specifically stimulated by our training protocol. In the m. gastrocnemius and soleus, the anabolic effects correlate with changes in the expression patterns of genes involved in MSTN signaling. Our data provide evidence that the decrease in the weight of androgen-sensitive sexual glands, observed after Md treatment, is caused by a suppression of endogenous testosterone synthesis. These observations provide new insights into the molecular mechanisms of the interaction between anabolic steroids, training and MSTN signaling during skeletal muscle adaptation.

  2. The role of diacylglycerol kinase ζ and phosphatidic acid in the mechanical activation of mammalian target of rapamycin (mTOR) signaling and skeletal muscle hypertrophy.

    Science.gov (United States)

    You, Jae-Sung; Lincoln, Hannah C; Kim, Chan-Ran; Frey, John W; Goodman, Craig A; Zhong, Xiao-Ping; Hornberger, Troy A

    2014-01-17

    The activation of mTOR signaling is essential for mechanically induced changes in skeletal muscle mass, and previous studies have suggested that mechanical stimuli activate mTOR (mammalian target of rapamycin) signaling through a phospholipase D (PLD)-dependent increase in the concentration of phosphatidic acid (PA). Consistent with this conclusion, we obtained evidence which further suggests that mechanical stimuli utilize PA as a direct upstream activator of mTOR signaling. Unexpectedly though, we found that the activation of PLD is not necessary for the mechanically induced increases in PA or mTOR signaling. Motivated by this observation, we performed experiments that were aimed at identifying the enzyme(s) that promotes the increase in PA. These experiments revealed that mechanical stimulation increases the concentration of diacylglycerol (DAG) and the activity of DAG kinases (DGKs) in membranous structures. Furthermore, using knock-out mice, we determined that the ζ isoform of DGK (DGKζ) is necessary for the mechanically induced increase in PA. We also determined that DGKζ significantly contributes to the mechanical activation of mTOR signaling, and this is likely driven by an enhanced binding of PA to mTOR. Last, we found that the overexpression of DGKζ is sufficient to induce muscle fiber hypertrophy through an mTOR-dependent mechanism, and this event requires DGKζ kinase activity (i.e. the synthesis of PA). Combined, these results indicate that DGKζ, but not PLD, plays an important role in mechanically induced increases in PA and mTOR signaling. Furthermore, this study suggests that DGKζ could be a fundamental component of the mechanism(s) through which mechanical stimuli regulate skeletal muscle mass.

  3. The Role of Diacylglycerol Kinase ζ and Phosphatidic Acid in the Mechanical Activation of Mammalian Target of Rapamycin (mTOR) Signaling and Skeletal Muscle Hypertrophy*

    Science.gov (United States)

    You, Jae-Sung; Lincoln, Hannah C.; Kim, Chan-Ran; Frey, John W.; Goodman, Craig A.; Zhong, Xiao-Ping; Hornberger, Troy A.

    2014-01-01

    The activation of mTOR signaling is essential for mechanically induced changes in skeletal muscle mass, and previous studies have suggested that mechanical stimuli activate mTOR (mammalian target of rapamycin) signaling through a phospholipase D (PLD)-dependent increase in the concentration of phosphatidic acid (PA). Consistent with this conclusion, we obtained evidence which further suggests that mechanical stimuli utilize PA as a direct upstream activator of mTOR signaling. Unexpectedly though, we found that the activation of PLD is not necessary for the mechanically induced increases in PA or mTOR signaling. Motivated by this observation, we performed experiments that were aimed at identifying the enzyme(s) that promotes the increase in PA. These experiments revealed that mechanical stimulation increases the concentration of diacylglycerol (DAG) and the activity of DAG kinases (DGKs) in membranous structures. Furthermore, using knock-out mice, we determined that the ζ isoform of DGK (DGKζ) is necessary for the mechanically induced increase in PA. We also determined that DGKζ significantly contributes to the mechanical activation of mTOR signaling, and this is likely driven by an enhanced binding of PA to mTOR. Last, we found that the overexpression of DGKζ is sufficient to induce muscle fiber hypertrophy through an mTOR-dependent mechanism, and this event requires DGKζ kinase activity (i.e. the synthesis of PA). Combined, these results indicate that DGKζ, but not PLD, plays an important role in mechanically induced increases in PA and mTOR signaling. Furthermore, this study suggests that DGKζ could be a fundamental component of the mechanism(s) through which mechanical stimuli regulate skeletal muscle mass. PMID:24302719

  4. TGF-β1/Smad3 Pathway Targets PP2A-AMPK-FoxO1 Signaling to Regulate Hepatic Gluconeogenesis.

    Science.gov (United States)

    Yadav, Hariom; Devalaraja, Samir; Chung, Stephanie T; Rane, Sushil G

    2017-02-24

    Maintenance of glucose homeostasis is essential for normal physiology. Deviation from normal glucose levels, in either direction, increases susceptibility to serious medical complications such as hypoglycemia and diabetes. Maintenance of glucose homeostasis is achieved via functional interactions among various organs: liver, skeletal muscle, adipose tissue, brain, and the endocrine pancreas. The liver is the primary site of endogenous glucose production, especially during states of prolonged fasting. However, enhanced gluconeogenesis is also a signature feature of type 2 diabetes (T2D). Thus, elucidating the signaling pathways that regulate hepatic gluconeogenesis would allow better insight into the process of normal endogenous glucose production as well as how this process is impaired in T2D. Here we demonstrate that the TGF-β1/Smad3 signaling pathway promotes hepatic gluconeogenesis, both upon prolonged fasting and during T2D. In contrast, genetic and pharmacological inhibition of TGF-β1/Smad3 signals suppressed endogenous glucose production. TGF-β1 and Smad3 signals achieved this effect via the targeting of key regulators of hepatic gluconeogenesis, protein phosphatase 2A (PP2A), AMP-activated protein kinase (AMPK), and FoxO1 proteins. Specifically, TGF-β1 signaling suppressed the LKB1-AMPK axis, thereby facilitating the nuclear translocation of FoxO1 and activation of key gluconeogenic genes, glucose-6-phosphatase and phosphoenolpyruvate carboxykinase. These findings underscore an important role of TGF-β1/Smad3 signaling in hepatic gluconeogenesis, both in normal physiology and in the pathophysiology of metabolic diseases such as diabetes, and are thus of significant medical relevance. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. TGF-β1/Smad3 Pathway Targets PP2A-AMPK-FoxO1 Signaling to Regulate Hepatic Gluconeogenesis*

    Science.gov (United States)

    Yadav, Hariom; Devalaraja, Samir; Chung, Stephanie T.; Rane, Sushil G.

    2017-01-01

    Maintenance of glucose homeostasis is essential for normal physiology. Deviation from normal glucose levels, in either direction, increases susceptibility to serious medical complications such as hypoglycemia and diabetes. Maintenance of glucose homeostasis is achieved via functional interactions among various organs: liver, skeletal muscle, adipose tissue, brain, and the endocrine pancreas. The liver is the primary site of endogenous glucose production, especially during states of prolonged fasting. However, enhanced gluconeogenesis is also a signature feature of type 2 diabetes (T2D). Thus, elucidating the signaling pathways that regulate hepatic gluconeogenesis would allow better insight into the process of normal endogenous glucose production as well as how this process is impaired in T2D. Here we demonstrate that the TGF-β1/Smad3 signaling pathway promotes hepatic gluconeogenesis, both upon prolonged fasting and during T2D. In contrast, genetic and pharmacological inhibition of TGF-β1/Smad3 signals suppressed endogenous glucose production. TGF-β1 and Smad3 signals achieved this effect via the targeting of key regulators of hepatic gluconeogenesis, protein phosphatase 2A (PP2A), AMP-activated protein kinase (AMPK), and FoxO1 proteins. Specifically, TGF-β1 signaling suppressed the LKB1-AMPK axis, thereby facilitating the nuclear translocation of FoxO1 and activation of key gluconeogenic genes, glucose-6-phosphatase and phosphoenolpyruvate carboxykinase. These findings underscore an important role of TGF-β1/Smad3 signaling in hepatic gluconeogenesis, both in normal physiology and in the pathophysiology of metabolic diseases such as diabetes, and are thus of significant medical relevance. PMID:28069811

  6. Porcine bocavirus NP1 negatively regulates interferon signaling pathway by targeting the DNA-binding domain of IRF9

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Ruoxi [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China); Fang, Liurong, E-mail: fanglr@mail.hzau.edu.cn [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China); Wang, Dang; Cai, Kaimei; Zhang, Huan [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China); Xie, Lilan; Li, Yi [College of Life Science and Technology, Wuhan Institute of Bioengineering, Wuhan 430415 (China); Chen, Huanchun; Xiao, Shaobo [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China)

    2015-11-15

    To subvert host antiviral immune responses, many viruses have evolved countermeasures to inhibit IFN signaling pathway. Porcine bocavirus (PBoV), a newly identified porcine parvovirus, has received attention because it shows clinically high co-infection prevalence with other pathogens in post-weaning multisystemic wasting syndrome (PWMS) and diarrheic piglets. In this study, we screened the structural and non-structural proteins encoded by PBoV and found that the non-structural protein NP1 significantly suppressed IFN-stimulated response element (ISRE) activity and subsequent IFN-stimulated gene (ISG) expression. However, NP1 affected neither the activation and translocation of STAT1/STAT2, nor the formation of the heterotrimeric transcription factor complex ISGF3 (STAT1/STAT2/IRF9). Detailed analysis demonstrated that PBoV NP1 blocked the ISGF3 DNA-binding activity by combining with the DNA-binding domain (DBD) of IRF9. In summary, these results indicate that PBoV NP1 interferes with type I IFN signaling pathway by blocking DNA binding of ISGF3 to attenuate innate immune responses. - Highlights: • Porcine bocavirus (PBoV) NP1 interferes with the IFN α/β signaling pathway. • PBoV NP1 does not prevent STAT1/STAT2 phosphorylation and nuclear translocation. • PBoV NP1 inhibits the DNA-binding activity of ISGF3. • PBoV NP1 interacts with the DNA-binding domain of IRF9.

  7. Oleosome-Associated Protein of the Oleaginous Diatom Fistulifera solaris Contains an Endoplasmic Reticulum-Targeting Signal Sequence

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    Yoshiaki Maeda

    2014-06-01

    Full Text Available Microalgae tend to accumulate lipids as an energy storage material in the specific organelle, oleosomes. Current studies have demonstrated that lipids derived from microalgal oleosomes are a promising source of biofuels, while the oleosome formation mechanism has not been fully elucidated. Oleosome-associated proteins have been identified from several microalgae to elucidate the fundamental mechanisms of oleosome formation, although understanding their functions is still in infancy. Recently, we discovered a diatom-oleosome-associated-protein 1 (DOAP1 from the oleaginous diatom, Fistulifera solaris JPCC DA0580. The DOAP1 sequence implied that this protein might be transported into the endoplasmic reticulum (ER due to the signal sequence. To ensure this, we fused the signal sequence to green fluorescence protein. The fusion protein distributed around the chloroplast as like a meshwork membrane structure, indicating the ER localization. This result suggests that DOAP1 could firstly localize at the ER, then move to the oleosomes. This study also demonstrated that the DOAP1 signal sequence allowed recombinant proteins to be specifically expressed in the ER of the oleaginous diatom. It would be a useful technique for engineering the lipid synthesis pathways existing in the ER, and finally controlling the biofuel quality.

  8. Advances in revealing the molecular targets downstream of oxidative stress-induced proapoptotic kinase signaling in diabetic embryopathy.

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    Wang, Fang; Reece, E Albert; Yang, Peixin

    2015-08-01

    Preexisting maternal diabetes is a high-risk factor of diabetic embryopathy, such as neural tube defects and congenital heart defects. Maternal diabetes significantly increases the production of reactive oxygen species, resulting in oxidative stress and diabetic embryopathy. Multiple cellular and metabolic factors contribute to these processes. Forkhead box O (FoxO)-3a has been demonstrated as a key transcription factor in the signaling transduction pathways responsible for maternal diabetes-induced birth defects. Apoptosis signal-regulating kinase 1 (ASK1) activated by oxidative stress stimulates nuclear translocation of FoxO3a, resulting in the overexpression of tumor necrosis factor receptor 1-associated death domain protein, which, in turn, leads to caspase-8 activation and apoptosis. Maternal diabetes-activated c-Jun N-terminal kinase (JNK)-1/2, downstream effectors of ASK1, can be blocked by superoxide dismutase-1 overexpression, suggesting that oxidative stress is responsible for JNK1/2 signaling activation. Deletion of JNK1/2 significantly suppressed the activity of FoxO3a. These observations indicate that maternal diabetes-induced oxidative stress stimulates the activation of ASK1, JNK1/2, FoxO3a, tumor necrosis factor receptor 1-associated death domain protein, caspase-8 cleavage, and finally, apoptosis and diabetic embryopathy. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. The role of nutraceuticals in pancreatic cancer prevention and therapy: Targeting cellular signaling, miRNAs and epigenome

    Science.gov (United States)

    Li, Yiwei; Go, Vay Liang W.; Sarkar, Fazlul H.

    2014-01-01

    Pancreatic cancer is one of the most aggressive malignancies in US adults. The experimental studies have found that antioxidant nutrients could reduce oxidative DNA damage, suggesting that these antioxidants may protect against pancreatic carcinogenesis. Several epidemiologic studies showed that dietary intake of antioxidants was inversely associated with the risk of pancreatic cancer, demonstrating the inhibitory effects of antioxidants on pancreatic carcinogenesis. Moreover, nutraceuticals, the anti-cancer agents from diet or natural plants, have been found to inhibit the development and progression of pancreatic cancer through the regulation of cellular signaling pathways. Importantly, nutraceuticals also up-regulate the expression of tumor suppressive miRNAs and down-regulate the expression of oncogenic miRNAs, leading to the inhibition of pancreatic cancer cell growth and pancreatic Cancer Stem Cell (CSC) self-renewal through modulation of cellular signaling network. Furthermore, nutraceuticals also regulate epigenetically deregulated DNAs and miRNAs, leading to the normalization of altered cellular signaling in pancreatic cancer cells. Therefore, nutraceuticals could have much broader use in the prevention and/or treatment of pancreatic cancer in combination with conventional chemotherapeutics. However, more in vitro mechanistic experiments, in vivo animal studies, and clinical trials are needed to realize the true value of nutraceuticals in the prevention and/or treatment of pancreatic cancer. PMID:25493373

  10. Macrophages are targets of retinoic acid signaling during the wound-healing process after thulium laser resection of the prostate.

    Science.gov (United States)

    Yu, Dian-Jun; Wang, Xing-Jie; Shi, Yun-Feng; Jiang, Chen-Yi; Zhao, Rui-Zhe; Zhu, Yi-Ping; Chen, Li; Yang, Yuan-Qing; Sun, Xiao-Wen; Xia, Shu-Jie

    2017-09-22

    The wound-healing process is very important for reducing complications after thulium laser resection of the prostate (TmLRP). The retinoic acid (RA) signaling pathway has been well studied in the wound-healing process of the skin and other organs. The goals of this study were to identify the role of RA signaling in the repair of the prostate after TmLRP and to investigate the molecular mechanism of this process. Retinoic acid receptors (RARs) were present in the prostate, and their expression was increased after TmLRP. RARβ was expressed in the macrophages and may be related to the role of stromal cells in the wound-healing process. In vitro, RA enhanced the function of anti-inflammatory macrophages and promoted stromal cell activation and angiogenesis. Arg1 was also increased via RARβ after treatment with RA. The expression of RARs was analyzed in vivo by immunohistochemistry (IHC), real time qPCR, and western blot analysis. THP-1 cells were co-treated with or without RA and stimulating factor and then assessed by ELISA and qPCR. The supernatants from these cells were cultured with stromal cells and vascular endothelial cells, and the effects on these cells were analyzed. We found that RA signaling was involved in the wound-healing process of the prostate after TmLRP. RA treated macrophages activated stromal cells and promoted angiogenesis. RARβ was the key isoform in this process.

  11. Lactobacillus rhamnosus ATCC 7469 exopolysaccharides synergizes with low level ionizing radiation to modulate signaling molecular targets in colorectal carcinogenesis in rats.

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    Zahran, Walid E; Elsonbaty, Sawsan M; Moawed, Fatma S M

    2017-08-01

    Combination therapy that targets cellular signaling pathway represents an alternative therapy for the treatment of colon cancer (CRC). The present study was therefore aimed to investigate the probable interaction of Lactobacillus rhamnosus ATCC 7469 exopolysaccharides (EPS) with low level ionizing γ radiation (γ-R) exposure against dimethylhydrazine (DMH)- induced colorectal carcinogenesis in rats. Colon cancer was induced with 20mg DMH/kg BW. Rats received daily by gastric gavage 100mg EPS/Kg BW concomitant with 1Gy γ-R over two months. Colonic oxidative and inflammatory stresses were assessed. The change in the expression of p-p38 MAPK, p-STAT3, β-catenin, NF-kB, COX-2 and iNOS was evaluated by western blotting and q-PCR. It was found that DMH treatment significantly induced colon oxidative injury accompanied by inflammatory disturbance along with increased protein expression of the targeted signaling factors p-p38 MAPK, p-STAT3 and β-catenin. The mRNA gene expression of NF-kB, COX-2 and iNOS was significantly higher in DMH-treated animals. It's worthy to note that colon tissues with DMH treatment showed significant dysplasia and anaplasia of the glandular mucosal lining epithelium with loses of goblet cells formation, pleomorphism in the cells and hyperchromachia in nuclei. Interestingly, EPS treatment with γ-R exposure showed statistically significant amelioration of the oxidative and inflammatory biomarkers with modulated signaling molecular factors accompanied by improved histological structure against DMH-induced CRC. In conclusion, our findings showed that Lactobacillus rhamnosus ATCC 7469 EPS with low level γ-R in synergistic interaction are efficacious control against CRC progression throughout the modulation of key signaling growth factors associated with inflammation via antioxidant mediated anti-inflammatory and anti-proliferative activities. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  12. HER2 signaling pathway activation and response of breast cancer cells to HER2-targeting agents is dependent strongly on the 3D microenvironment

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    Weigelt, Britta; Lo, Alvin T; Park, Catherine C; Gray, Joe W; Bissell, Mina J

    2009-07-27

    Development of effective and durable breast cancer treatment strategies requires a mechanistic understanding of the influence of the microenvironment on response. Previous work has shown that cellular signaling pathways and cell morphology are dramatically influenced by three-dimensional (3D) cultures as opposed to traditional two-dimensional (2D) monolayers. Here, we compared 2D and 3D culture models to determine the impact of 3D architecture and extracellular matrix (ECM) on HER2 signaling and on the response of HER2-amplified breast cancer cell lines to the HER2-targeting agents Trastuzumab, Pertuzumab and Lapatinib. We show that the response of the HER2-amplified AU565, SKBR3 and HCC1569 cells to these anti-HER2 agents was highly dependent on whether the cells were cultured in 2D monolayer or 3D laminin-rich ECM gels. Inhibition of {beta}1 integrin, a major cell-ECM receptor subunit, significantly increased the sensitivity of the HER2-amplified breast cancer cell lines to the humanized monoclonal antibodies Trastuzumab and Pertuzumab when grown in a 3D environment. Finally, in the absence of inhibitors, 3D cultures had substantial impact on HER2 downstream signaling and induced a switch between PI3K-AKT- and RAS-MAPKpathway activation in all cell lines studied, including cells lacking HER2 amplification and overexpression. Our data provide direct evidence that breast cancer cells are able to rapidly adapt to different environments and signaling cues by activating alternative pathways that regulate proliferation and cell survival, events that may play a significant role in the acquisition of resistance to targeted therapies.

  13. Identification and functional analysis of NOL7 nuclear and nucleolar localization signals

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    Lingen Mark W

    2010-09-01

    Full Text Available Abstract Background NOL7 is a candidate tumor suppressor that localizes to a chromosomal region 6p23. This locus is frequently lost in a number of malignancies, and consistent loss of NOL7 through loss of heterozygosity and decreased mRNA and protein expression has been observed in tumors and cell lines. Reintroduction of NOL7 into cells resulted in significant suppression of in vivo tumor growth and modulation of the angiogenic phenotype. Further, NOL7 was observed to localize to the nucleus and nucleolus of cells. However, the mechanisms regulating its subcellular localization have not been elucidated. Results An in vitro import assay demonstrated that NOL7 requires cytosolic machinery for active nuclear transport. Using sequence homology and prediction algorithms, four putative nuclear localization signals (NLSs were identified. NOL7 deletion constructs and cytoplasmic pyruvate kinase (PK fusion proteins confirmed the functionality of three of these NLSs. Site-directed mutagenesis of PK fusions and full-length NOL7 defined the minimal functional regions within each NLS. Further characterization revealed that NLS2 and NLS3 were critical for both the rate and efficiency of nuclear targeting. In addition, four basic clusters within NLS2 and NLS3 were independently capable of nucleolar targeting. The nucleolar occupancy of NOL7 revealed a complex balance of rapid nucleoplasmic shuttling but low nucleolar mobility, suggesting NOL7 may play functional roles in both compartments. In support, targeting to the nucleolar compartment was dependent on the presence of RNA, as depletion of total RNA or rRNA resulted in a nucleoplasmic shift of NOL7. Conclusions These results identify the minimal sequences required for the active targeting of NOL7 to the nucleus and nucleolus. Further, this work characterizes the relative contribution of each sequence to NOL7 nuclear and nucleolar dynamics, the subnuclear constituents that participate in this targeting, and

  14. Spinal CCL1/CCR8 signaling interplay as a potential therapeutic target - Evidence from a mouse diabetic neuropathy model.

    Science.gov (United States)

    Zychowska, Magdalena; Rojewska, Ewelina; Piotrowska, Anna; Kreiner, Grzegorz; Nalepa, Irena; Mika, Joanna

    2017-11-01

    Chemokine signaling has been implicated in the pathogenesis of diabetic neuropathy; however, the involvement of the chemokine CC motif ligand 1 (CCL1)-chemokine CC motif receptor 8 (CCR8) interaction remains unknown. The goal of this study was to examine the role of CCL1-CCR8 signaling interplay in the development of hypersensitivity and in opioid effectiveness in diabetic neuropathy. Primary glial cell cultures and a streptozotocin (STZ; 200mg/kg, intraperitoneal)-induced mouse model of diabetic neuropathy were used. Analysis of mRNA/protein expression of glial markers and CCL1/CCR8 was performed by qRT-PCR, Western blotting and/or protein arrays. The co-localization of CCL1/CCR8 with neural/glial cells was visualized by immunofluorescence. The pharmacological tools were injected intrathecally, and pain behavior was evaluated by von Frey/cold plate tests. Single STZ injection increased blood glucose levels and induced the development of hypersensitivity as measured on days 7-21. On day 7 after STZ, the protein levels of CCL1 and IBA1 but not of CCR8 or GFAP were elevated. Immunofluorescent staining revealed that CCR8 was predominantly localized in neurons, which are also the main source of spinal CCL1. Lipopolysaccharide stimulation of primary microglial cultures resulted in decreases in the levels of CCL1 and CCR8. Single intrathecal injection of CCL1 (10-500ng) induced the development of hypersensitivity, whereas on day 7 after STZ, a CCL1-neutralizing antibody dose-dependently (2-8μg) delayed pain behavior. Repeated administration of the CCL1-neutralizing antibody (4μg) also enhanced the effectiveness of morphine and buprenorphine (1μg). These results reveal that CCL1/CCR8 neuronal signaling plays an important role in the development of diabetic neuropathy and the effectiveness of opioids. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Orf virus 002 protein targets ovine protein S100A4 and inhibits NF-kappa B signaling

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    Daxiang Chen

    2016-09-01

    Full Text Available Orf virus (ORFV, a member of Parapoxvirus, has evolved various strategies to modulate the immune responses of host cells. The ORFV-encoded protein ORFV002, a regulator factor, has been found to inhibit the acetylation of NF-κB-p65 by blocking phosphorylation of NF-kB-p65 at Ser276 and also to disrupt the binding of NF-kB-p65 and p300. To explore the mechanism by which ORFV002 regulates NF-κB signaling, the understanding of ORFV002 potential binding partners in host cells is critical. In this study, ovine S100 calcium binding protein A4 (S100A4, prolylendopeptidase-like (PREPL and NADH dehydrogenase (ubiquinone 1 alpha subcomplex 8 (NDUFA8 were found to interact with ORFV002 based on the yeast two-hybrid (Y2H assay using a cDNA library derived from primary ovine fetal turbinate cells (OFTu. GST pull-down and bidirectional co-immunoprecipitation assay results demonstrate that ORFV002 interacts with S100A4 directly. Following the pEGFP-ORFV002 (p002GFP transfection, we found that cytoplasmic S100A4 translocates into the nucleus and co-localizes with ORFV002. Furthermore, the inhibitory effect of ORFV002 on NF-κB signaling was significantly restored by S100A4 knock-down phenotype, suggesting ovine S100A4 participating in the ORFV002-mediated NF-κB signaling. These data demonstrate that ORFV002 inhibits the NF-κB activation through its interaction with S100A4 along with its nucleus translocation.

  16. Expressed miRNAs target feather related mRNAs involved in cell signaling, cell adhesion and structure during chicken epidermal development.

    Science.gov (United States)

    Bao, Weier; Greenwold, Matthew J; Sawyer, Roger H

    2016-10-15

    MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression at the post-transcriptional level. Previous studies have shown that miRNA regulation contributes to a diverse set of processes including cellular differentiation and morphogenesis which leads to the creation of different cell types in multicellular organisms and is thus key to animal development. Feathers are one of the most distinctive features of extant birds and are important for multiple functions including flight, thermal regulation, and sexual selection. However, the role of miRNAs in feather development has been woefully understudied despite the identification of cell signaling pathways, cell adhesion molecules and structural genes involved in feather development. In this study, we performed a microarray experiment comparing the expression of miRNAs and mRNAs among three embryonic stages of development and two tissues (scutate scale and feather) of the chicken. We combined this expression data with miRNA target prediction tools and a curated list of feather related genes to produce a set of 19 miRNA-mRNA duplexes. These targeted mRNAs have been previously identified as important cell signaling and cell adhesion genes as well as structural genes involved in feather and scale morphogenesis. Interestingly, the miRNA target site of the cell signaling pathway gene, Aldehyde Dehydrogenase 1 Family, Member A3 (ALDH1A3), is unique to birds indicating a novel role in Aves. The identified miRNA target site of the cell adhesion gene, Tenascin C (TNC), is only found in specific chicken TNC splice variants that are differentially expressed in developing scutate scale and feather tissue indicating an important role of miRNA regulation in epidermal differentiation. Additionally, we found that β-keratins, a major structural component of avian and reptilian epidermal appendages, are targeted by multiple miRNA genes. In conclusion, our work provides quantitative expression data on miRNAs and m

  17. Global MEF2 target gene analysis in cardiac and skeletal muscle reveals novel regulation of DUSP6 by p38MAPK-MEF2 signaling

    Science.gov (United States)

    Wales, Stephanie; Hashemi, Sara; Blais, Alexandre; McDermott, John C.

    2014-01-01

    MEF2 plays a profound role in the regulation of transcription in cardiac and skeletal muscle lineages. To define the overlapping and unique MEF2A genomic targets, we utilized ChIP-exo analysis of cardiomyocytes and skeletal myoblasts. Of the 2783 and 1648 MEF2A binding peaks in skeletal myoblasts and cardiomyocytes, respectively, 294 common binding sites were identified. Genomic targets were compared to differentially expressed genes in RNA-seq analysis of MEF2A depleted myogenic cells, revealing two prominent genetic networks. Genes largely associated with muscle development were down-regulated by loss of MEF2A while up-regulated genes reveal a previously unrecognized function of MEF2A in suppressing growth/proliferative genes. Several up-regulated (Tprg, Mctp2, Kitl, Prrx1, Dusp6) and down-regulated (Atp1a2, Hspb7, Tmem182, Sorbs2, Lmod3) MEF2A target genes were chosen for further investigation. Interestingly, siRNA targeting of the MEF2A/D heterodimer revealed a somewhat divergent role in the regulation of Dusp6, a MAPK phosphatase, in cardiac and skeletal myogenic lineages. Furthermore, MEF2D functions as a p38MAPK-dependent repressor of Dusp6 in myoblasts. These data illustrate that MEF2 orchestrates both common and non-overlapping programs of signal-dependent gene expression in skeletal and cardiac muscle lineages. PMID:25217591

  18. The Induction of Recombinant Protein Bodies in Different Subcellular Compartments Reveals a Cryptic Plastid-Targeting Signal in the 27-kDa γ-Zein Sequence.

    Science.gov (United States)

    Hofbauer, Anna; Peters, Jenny; Arcalis, Elsa; Rademacher, Thomas; Lampel, Johannes; Eudes, François; Vitale, Alessandro; Stoger, Eva

    2014-01-01

    Naturally occurring storage proteins such as zeins are used as fusion partners for recombinant proteins because they induce the formation of ectopic storage organelles known as protein bodies (PBs) where the proteins are stabilized by intermolecular interactions and the formation of disulfide bonds. Endogenous PBs are derived from the endoplasmic reticulum (ER). Here, we have used different targeting sequences to determine whether ectopic PBs composed of the N-terminal portion of mature 27 kDa γ-zein added to a fluorescent protein could be induced to form elsewhere in the cell. The addition of a transit peptide for targeting to plastids causes PB formation in the stroma, whereas in the absence of any added targeting sequence PBs were typically associated with the plastid envelope, revealing the presence of a cryptic plastid-targeting signal within the γ-zein cysteine-rich domain. The subcellular localization of the PBs influences their morphology and the solubility of the stored recombinant fusion protein. Our results indicate that the biogenesis and budding of PBs does not require ER-specific factors and therefore, confirm that γ-zein is a versatile fusion partner for recombinant proteins offering unique opportunities for the accumulation and bioencapsulation of recombinant proteins in different subcellular compartments.

  19. The induction of recombinant protein bodies in different subcellular compartments reveals a cryptic plastid-targeting signal in the 27 kD γ-zein sequence

    Directory of Open Access Journals (Sweden)

    Anna eHofbauer

    2014-12-01

    Full Text Available Naturally-occurring storage proteins such as zeins are used as fusion partners for recombinant proteins because they induce the formation of ectopic storage organelles known as protein bodies (PBs where the proteins are stabilized by intermolecular interactions and the formation of disulfide bonds. Endogenous PBs are derived from the endoplasmic reticulum (ER. Here we have used different targeting sequences to determine whether ectopic PBs composed of the N-terminal portion of mature 27 kD γ-zein added to a fluorescent protein could be induced to form elsewhere in the cell. The addition of a transit peptide for targeting to plastids causes PB formation in the stroma, whereas in the absence of any added targeting sequence PBs were typically associated with the plastid envelope, revealing the presence of a cryptic plastid targeting signal within the γ-zein cysteine-rich domain. The subcellular localization of the PBs influences their morphology and the solubility of the stored recombinant fusion protein. Our results indicate that the biogenesis and budding of PBs does not require ER-specific factors and therefore confirm that γ-zein is a versatile fusion partner for recombinant proteins offering unique opportunities for the accumulation and bioencapsulation of recombinant proteins in different subcellular compartments.

  20. Dasatinib synergizes with both cytotoxic and signal transduction inhibitors in heterogeneous breast cancer cell lines--lessons for design of combination targeted therapy.

    Science.gov (United States)

    Park, Brian J; Whichard, Zakary L; Corey, Seth J

    2012-07-01

    Molecularly targeted therapies have emerged as the leading theme in cancer therapeutics. Multi-cytotoxic drug regimens have been highly successful, yet many studies in targeted therapeutics have centered on a single agent. We investigated whether the Src/Abl kinase inhibitor dasatinib displays synergy with other agents in molecularly heterogeneous breast cancer cell lines. MCF-7, SKBR-3, and MDA-MB-231 display different signaling and gene signatures profiles due to expression of the estrogen receptor, ErbB2, or neither. Cell proliferation was measured following treatment with dasatinib±cytotoxic (paclitaxel, ixabepilone) or molecularly targeted agents (tamoxifen, rapamycin, sorafenib, pan PI3K inhibitor LY294002, and MEK/ERK inhibitor U0126). Dose-responses for single or combination drugs were calculated and analyzed by the Chou-Talalay method. The drugs with the greatest level of synergy with dasatinib were rapamycin, ixabepilone, and sorafenib, for the MDA-MB-231, MCF-7, and SK-BR-3 cell lines respectively. However, dasatinib synergized with both cytotoxic and molecularly targeted agents in all three molecularly heterogeneous breast cancer cell lines. These results suggest that effectiveness of rationally designed therapies may not entirely rest on precise identification of gene signatures or molecular profiling. Since a systems analysis that reveals emergent properties cannot be easily performed for each cancer case, multi-drug regimens in the near future will still involve empirical design. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  1. Particle bombardment-mediated transient expression to identify localization signals in plant disease resistance proteins and target sites for the proteolytic activity of pathogen effectors.

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    Takemoto, Daigo; Jones, David A

    2014-01-01

    Plant pathogens, including fungi, oomycetes, bacteria, aphids, and nematodes, produce a variety of effector proteins to counter plant disease resistance mechanisms. After delivery into the cytosol of the plant cell, effectors may target proteins localized to different compartments within the plant cell. Plants, in turn, have evolved disease resistance (R) proteins to recognize the action of effectors. Elucidation of the subcellular localization of pathogen effectors, the plant proteins they target, and plant disease resistance proteins is essential to fully understand their interactions during pathogen challenge. In recent years, expression of fluorescent protein fusions has been widely used to determine the subcellular localization of plant proteins and pathogen effectors. Use of fluorescent proteins enables researchers to monitor the dynamic behavior of proteins in living cells. Among various methods available for the introduction of genes into plant cells, particle bombardment-mediated transient expression is the most rapid method suitable for both the identification of localization signals in proteins of interest and their dissection via amino acid substitutions generated using site-directed mutagenesis. This chapter describes a rapid procedure for particle bombardment-mediated transient expression in leaf epidermal cells. This method is also applicable to detection of pathogen effector protease activities directed against target proteins in the plant cell and analysis of protease recognition sites within these target proteins.

  2. Dual small-molecule targeting of SMAD signaling stimulates human induced pluripotent stem cells toward neural lineages.

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    Methichit Wattanapanitch

    Full Text Available Incurable neurological disorders such as Parkinson's disease (PD, Huntington's disease (HD, and Alzheimer's disease (AD are very common and can be life-threatening because of their progressive disease symptoms with limited treatment options. To provide an alternative renewable cell source for cell-based transplantation and as study models for neurological diseases, we generated induced pluripotent stem cells (iPSCs from human dermal fibroblasts (HDFs and then differentiated them into neural progenitor cells (NPCs and mature neurons by dual SMAD signaling inhibitors. Reprogramming efficiency was improved by supplementing the histone deacethylase inhibitor, valproic acid (VPA, and inhibitor of p160-Rho associated coiled-coil kinase (ROCK, Y-27632, after retroviral transduction. We obtained a number of iPS colonies that shared similar characteristics with human embryonic stem cells in terms of their morphology, cell surface antigens, pluripotency-associated gene and protein expressions as well as their in vitro and in vivo differentiation potentials. After treatment with Noggin and SB431542, inhibitors of the SMAD signaling pathway, HDF-iPSCs demonstrated rapid and efficient differentiation into neural lineages. Six days after neural induction, neuroepithelial cells (NEPCs were observed in the adherent monolayer culture, which had the ability to differentiate further into NPCs and neurons, as characterized by their morphology and the expression of neuron-specific transcripts and proteins. We propose that our study may be applied to generate neurological disease patient-specific iPSCs allowing better understanding of disease pathogenesis and drug sensitivity assays.

  3. Molecular biology of human epidermal receptors, signaling pathways and targeted therapy against cancers: new evidences and old challenges

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    Paulo Michel Pinheiro Ferreira

    2017-07-01

    Full Text Available ABSTRACT Human epidermal receptors (HER1/2/3/4 belong to the class of receptor-type tyrosine kinases. After binding a ligand, dimerization, it will ocurr activation of intracellular kinases after two-dimensional and cytoplasmic tail reciprocal transphosphorylation. This transphosphorylation recruits signaling pathways such as Ras/Raf/MEK/Erk1-2, PI3-K/AKT and JAK/STAT, which can affect the cell cycle, cytoskeleton reorganization, apoptosis, metastasis, differentiation, angiogenesis and transcription. HER deregulation is found in epithelial, mesenchymal and nervous neoplasms and is associated with poor prognosis and tumor severity. Since HER are promiscuous proteins when subjected to mutations, resultant modifications confer cellular metabolic superiority and activate complex, interconnected and overlapping networks of cytoplasmic signaling. Moreover, overexpression of HER1/2 is involved in tumor resistance to radiation and anti-hormone therapies. Indeed, HER2 expression is up to 100-fold higher in 25-30% of invasive breast cancers. These characteristics support the development of resistance to anti-HER1/2 chemotherapy such as monoclonal antibodies and tyrosine kinase inhibitors. Then, the challenges in research with HER-positive cancers include planning therapeutic strategies against known resistance mechanisms and identifying novel mechanisms as a way to overcome and control cell growth and malignant progression.

  4. Moderate mammalian target of rapamycin inhibition induces autophagy in HTR8/SVneo cells via O-linked β-N-acetylglucosamine signaling.

    Science.gov (United States)

    Zhang, Qiuxia; Na, Quan; Song, Weiwei

    2017-10-01

    Autophagy, a highly regulated process with a dual role (pro-survival or pro-death), has been implicated in adverse pregnancy outcomes. The aim of this study was to explore the mechanism whereby mammalian target of rapamycin (mTOR) signaling regulates autophagy by modulating protein O-GlcNAcylation in human trophoblasts. HTR8/SVneo cells were incubated in serum-free medium for different time intervals or treated with varying doses of Torin1. Protein expression and cell apoptosis were detected by immunoblotting and flow cytometry, respectively. Short-term serum starvation or slight suppression of mTOR signaling promoted autophagy and decreased apoptosis in HTR8/SVneo cells. Conversely, prolonged serum starvation or excessive inhibition of mTOR reduced autophagy and enhanced cell apoptosis. Both serum starvation and mTOR signaling suppression reduced protein O-GlcNAcylation. Upregulation and downregulation of O-linked β-N-acetylglucosamine (O-GlcNAc) levels attenuated and augmented autophagy, respectively. Moderate mTOR inhibition-induced autophagy was blocked by upregulation of protein O-GlcNAcylation. Furthermore, immunoprecipitation studies revealed that Beclin1 and synaptosome associated protein 29 (SNAP29) could be O-GlcNAcylated, and that slight mTOR inhibition resulted in decreased O-GlcNAc modification of Beclin1 and SNAP29. Notably, we observed an inverse correlation between phosphorylation (Ser15) and O-GlcNAcylation of Beclin1. mTOR signaling inhibition played dual roles in regulating autophagy and apoptosis in HTR8/SVneo cells. Moderate mTOR suppression might induce autophagy via modulating O-GlcNAcylation of Beclin1 and SNAP29. Moreover, the negative interplay between Beclin1 O-GlcNAcylation and phosphorylation (Ser15) may be involved in autophagy regulation by mTOR signaling. © 2017 Japan Society of Obstetrics and Gynecology.

  5. The Bacterial Effector HopX1 Targets JAZ Transcriptional Repressors to Activate Jasmonate Signaling and Promote Infection in Arabidopsis

    Science.gov (United States)

    Gimenez-Ibanez, Selena; Boter, Marta; Fernández-Barbero, Gemma; Chini, Andrea; Rathjen, John P.; Solano, Roberto

    2014-01-01

    Pathogenicity of Pseudomonas syringae is dependent on a type III secretion system, which secretes a suite of virulence effector proteins into the host cytoplasm, and the production of a number of toxins such as coronatine (COR), which is a mimic of the plant hormone jasmonate-isoleuce (JA-Ile). Inside the plant cell, effectors target host molecules to subvert the host cell physiology and disrupt defenses. However, despite the fact that elucidating effector action is essential to understanding bacterial pathogenesis, the molecular function and host targets of the vast majority of effectors remain largely unknown. Here, we found that effector HopX1 from Pseudomonas syringae pv. tabaci (Pta) 11528, a strain that does not produce COR, interacts with and promotes the degradation of JAZ proteins, a key family of JA-repressors. We show that hopX1 encodes a cysteine protease, activity that is required for degradation of JAZs by HopX1. HopX1 associates with JAZ proteins through its central ZIM domain and degradation occurs in a COI1-independent manner. Moreover, ectopic expression of HopX1 in Arabidopsis induces the expression of JA-dependent genes, represses salicylic acid (SA)-induced markers, and complements the growth of a COR-deficient P. syringae pv. tomato (Pto) DC3000 strain during natural bacterial infections. Furthermore, HopX1 promoted susceptibility when delivered by the natural type III secretion system, to a similar extent as the addition of COR, and this effect was dependent on its catalytic activity. Altogether, our results indicate that JAZ proteins are direct targets of bacterial effectors to promote activation of JA-induced defenses and susceptibility in Arabidopsis. HopX1 illustrates a paradigm of an alternative evolutionary solution to COR with similar physiological outcome. PMID:24558350

  6. The bacterial effector HopX1 targets JAZ transcriptional repressors to activate jasmonate signaling and promote infection in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Selena Gimenez-Ibanez

    2014-02-01

    Full Text Available Pathogenicity of Pseudomonas syringae is dependent on a type III secretion system, which secretes a suite of virulence effector proteins into the host cytoplasm, and the production of a number of toxins such as coronatine (COR, which is a mimic of the plant hormone jasmonate-isoleuce (JA-Ile. Inside the plant cell, effectors target host molecules to subvert the host cell physiology and disrupt defenses. However, despite the fact that elucidating effector action is essential to understanding bacterial pathogenesis, the molecular function and host targets of the vast majority of effectors remain largely unknown. Here, we found that effector HopX1 from Pseudomonas syringae pv. tabaci (Pta 11528, a strain that does not produce COR, interacts with and promotes the degradation of JAZ proteins, a key family of JA-repressors. We show that hopX1 encodes a cysteine protease, activity that is required for degradation of JAZs by HopX1. HopX1 associates with JAZ proteins through its central ZIM domain and degradation occurs in a COI1-independent manner. Moreover, ectopic expression of HopX1 in Arabidopsis induces the expression of JA-dependent genes, represses salicylic acid (SA-induced markers, and complements the growth of a COR-deficient P. syringae pv. tomato (Pto DC3000 strain during natural bacterial infections. Furthermore, HopX1 promoted susceptibility when delivered by the natural type III secretion system, to a similar extent as the addition of COR, and this effect was dependent on its catalytic activity. Altogether, our results indicate that JAZ proteins are direct targets of bacterial effectors to promote activation of JA-induced defenses and susceptibility in Arabidopsis. HopX1 illustrates a paradigm of an alternative evolutionary solution to COR with similar physiological outcome.

  7. Precision Medicine Approach to Anaplastic Thyroid Cancer: Advances in Targeted Drug Therapy Based on Specific Signaling Pathways.

    Science.gov (United States)

    Samimi, Hilda; Fallah, Parviz; Naderi Sohi, Alireza; Tavakoli, Rezvan; Naderi, Mahmood; Soleimani, Masoud; Larijani, Bagher; Haghpanah, Vahid

    2017-03-01

    Personalized medicine is a set of diagnostic, prognostic and therapeutic approaches in which medical interventions are carried out based on individual patient characteristics. As life expectancy increases in developed and developing countries, the incidence of diseases such as cancer goes up among people in the community. Cancer is a disease that the response to treatment varies from one person to another and also it is costly for individuals, families, and society. Among thyroid cancers, anaplastic thyroid carcinoma (ATC) is the most aggressive, lethal and unresponsive form of the disease. Unfortunately, current drugs are not targetable, and therefore they have restricted role in ATC treatment. Consequently, mortality of this cancer, despite advances in the field of diagnosis and treatment, is one of the most important challenges in medicine. Cellular, molecular and genetic evidences play an important role in finding more effective diagnostic and therapeutic approaches. Review of these evidences confirms the application of personalized medicine in cancer treatment including ATC. A growing body of evidence has elucidated that cellular and molecular mechanisms of cancer would pave the way for defining new biomarkers for targeted therapy, taking into account individual differences. It should be noted that this approach requires further progress in the fields of basic sciences, pharmacogenetics and drug design. An overview of the most important aspects in individualized anaplastic thyroid cancer treatment will be discussed in this review.

  8. Eriocalyxin B Inhibits STAT3 Signaling by Covalently Targeting STAT3 and Blocking Phosphorylation and Activation of STAT3.

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    Xiaokui Yu

    Full Text Available Activated STAT3 plays an important role in oncogenesis by stimulating cell proliferation and resisting apoptosis. STAT3 therefore is an attractive target for cancer therapy. We have screened a traditional Chinese herb medicine compound library and found Eriocalyxin B (EB, a diterpenoid from Isodon eriocalyx, as a specific inhibitor of STAT3. EB selectively inhibited constitutive as well as IL-6-induced phosphorylation of STAT3 and induced apoptosis of STAT3-dependent tumor cells. EB did not affect the upstream protein tyrosine kinases or the phosphatase (PTPase of STAT3, but rather interacted directly with STAT3. The effects of EB could be abolished by DTT or GSH, suggesting a thiol-mediated covalent linkage between EB and STAT3. Site mutagenesis of cysteine in and near the SH2 domain of STAT3 identified Cys712 to be the critical amino acid for the EB-induced inactivation of STAT3. Furthermore, LC/MS/MS analyses demonstrated that an α, β-unsaturated carbonyl of EB covalently interacted with the Cys712 of STAT3. Computational modeling analyses also supported a direct interaction between EB and the Cys712 of STAT3. These data strongly suggest that EB directly targets STAT3 through a covalent linkage to inhibit the phosphorylation and activation of STAT3 and induces apoptosis of STAT3-dependent tumor cells.

  9. Targeted mutagenesis of the P22 portal protein reveals the mechanism of signal transmission during DNA packaging.

    Science.gov (United States)

    Bedwell, Gregory J; Prevelige, Peter E

    2017-05-01

    The portal vertex in dsDNA bacteriophage serves as the site for genome encapsidation and release. In several of these viruses, efficient termination of DNA packaging has been shown to be dependent on the density of packaged DNA. The portal protein has been implicated as being part of the sensor that regulates packaging termination through DNA-dependent conformational changes during packaging. The mechanism by which DNA induces these conformational changes remains unknown. In this study, we explore how point mutants in the portal core can result in changes in genome packaging density in P22. Mutations in the portal core that subtly alter the structure or dynamics of the protein result in an increase in the amount of DNA packaged. The magnitude of the change is amino acid and location specific. Our findings suggest a mechanism wherein compression of the portal core is an essential aspect of signal transmission during packaging. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. RIP4 is a target of multiple signal transduction pathways in keratinocytes: Implications for epidermal differentiation and cutaneous wound repair

    Energy Technology Data Exchange (ETDEWEB)

    Adams, Stephanie [Charite, University Medicine Berlin, Institute of Physiology, Arnimallee 22, D-14195 Berlin (Germany); Munz, Barbara, E-mail: barbara.munz@charite.de [Charite, University Medicine Berlin, Institute of Physiology, Arnimallee 22, D-14195 Berlin (Germany)

    2010-01-01

    Receptor interacting protein 4 (RIP4) is an important regulator of epidermal morphogenesis during embryonic development. We could previously show that expression of the rip4 gene is strongly downregulated in cutaneous wound repair, which might be initiated by a broad variety of growth factors and cytokines. Here, we demonstrate that in keratinocytes, rip4 expression is controlled by a multitude of different signal transduction pathways, such as the p38 mitogen-activated protein kinase (MAPK) and the nuclear factor kappa B (NF-{kappa}B) cascade, in a unique and specific manner. Furthermore, we show that the steroid dexamethasone abolishes the physiological rip4 downregulation after injury and might thus contribute to the phenotype of reduced and delayed wound reepithelialization seen in glucocorticoid-treated patients. As a whole, our data indicate that rip4 expression is regulated in a complex manner, which might have therapeutic implications.

  11. Peroxisome proliferator-activated receptor gamma is a novel target of the nerve growth factor signaling pathway in PC12 cells.

    Science.gov (United States)

    Fuenzalida, Karen M; Aguilera, Mauricio C; Piderit, Daniela G; Ramos, Patricio C; Contador, David; Quiñones, Verónica; Rigotti, Atilio; Bronfman, Francisca C; Bronfman, Miguel

    2005-03-11

    Peroxisome proliferator-activated receptor gamma (PPARgamma), a member of the nuclear receptor superfamily, is subject to considerable interest because of its role in adipocyte differentiation, metabolic control, and anti-inflammatory action. PPARgamma research in brain cells is presently focused on glial PPARgamma because of its potential as a pharmacological target in the treatment of neurodegenerative diseases with an inflammatory component. In neurons PPARgamma function is far from clear, and PPARgamma agonist-dependent and -independent effects on cell survival or differentiation have been reported. We used PC12 cells, widely used to study neuronal signaling, such as nerve growth factor (NGF)-induced differentiation and survival or epidermal growth factor-dependent cell proliferation to dissect the possible involvement of PPARgamma in these pathways. We show that NGF but not epidermal growth factor increases the transcriptional activity of PPARgamma, and modulates the expression of this transcription factor. Because NGF signals through the tyrosine kinase (TrkA) NGF receptor and/or the p75NTR receptor, we used rescue experiments with a PC12 cell mutant lacking TrkA to show that NGF-induced PPARgamma activation is dependent on TrkA activation. Our results point out PPARgamma as a novel target of the TrkA-mediated neuronal cell survival and differentiating pathway and suggest a potential new inflammatory-independent therapeutic approach for pharmacological intervention in neurological disorders.

  12. Pirarubicin induces an autophagic cytoprotective response through suppression of the mammalian target of rapamycin signaling pathway in human bladder cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Li, Kuiqing; Chen, Xu [Department of Urology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120 (China); Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120 (China); Liu, Cheng [Department of Urology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120 (China); Gu, Peng; Li, Zhuohang; Wu, Shaoxu [Department of Urology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120 (China); Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120 (China); Xu, Kewei [Department of Urology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120 (China); Lin, Tianxin, E-mail: tianxinl@sina.com [Department of Urology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120 (China); Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120 (China); Huang, Jian, E-mail: urolhj@sina.com [Department of Urology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120 (China)

    2015-05-01

    Pirarubicin is widely used in intravesical chemotherapy for bladder cancer, but its efficacy is limited due to drug resistance; the mechanism has not been well studied. Emerging evidence shows that autophagy can be a novel target for cancer therapy. This study aimed to investigate the role of autophagy in pirarubicin-treated bladder cancer cells. Bladder cancer cells EJ and J82 were treated with pirarubicin, siRNA, 3-methyladenine or hydroxychloroquine. Cell proliferation and apoptosis were tested by cell survival assay and flow cytometric analysis, respectively. Autophagy was evaluated by immunoblotting before and after the treatments. The phosphorylated mammalian target of rapamycin, serine/threonine kinase p70 S6 kinase, and eukaryotic translation initiation factor 4E binding protein 1 were also investigated by immunoblotting. We found that pirarubicin could induce autophagy in bladder cancer cells. Inhibition of autophagy by 3-methyladenine, hydroxychloroquine or knockdown of autophagy related gene 3 significantly increased apoptosis in pirarubicin-treated bladder cancer cells. Pirarubicin-induced autophagy was mediated via the mTOR/p70S6K/4E-BP1 signaling pathway. In conclusion, autophagy induced by pirarubicin plays a cytoprotective role in bladder cancer cells, suggesting that inhibition of autophagy may improve efficacy over traditional pirarubicin chemotherapy in bladder cancer patients. - Highlights: • Pirarubicin induced autophagy in bladder cancer cells. • Inhibition of autophagy enhanced pirarubicin-induced apoptosis. • Pirarubicin induced autophagy through inhibition of mTOR signaling pathway.

  13. Feedback Regulation of ABA Signaling and Biosynthesis by a bZIP Transcription Factor Targets Drought-Resistance-Related Genes1[OPEN

    Science.gov (United States)

    Tang, Ning; Yang, Jun; Peng, Lei; Ma, Siqi; Xu, Yan; Li, Guoliang

    2016-01-01

    The OsbZIP23 transcription factor has been characterized for its essential role in drought resistance in rice (Oryza sativa), but the mechanism is unknown. In this study, we first investigated the transcriptional activation of OsbZIP23. A homolog of SnRK2 protein kinase (SAPK2) was found to interact with and phosphorylate OsbZIP23 for its transcriptional activation. SAPK2 also interacted with OsPP2C49, an ABI1 homolog, which deactivated the SAPK2 to inhibit the transcriptional activation activity of OsbZIP23. Next, we performed genome-wide identification of OsbZIP23 targets by immunoprecipitation sequencing and RNA sequencing analyses in the OsbZIP23-overexpression, osbzip23 mutant, and wild-type rice under normal and drought stress conditions. OsbZIP23 directly regulates a large number of reported genes that function in stress response, hormone signaling, and developmental processes. Among these targets, we found that OsbZIP23 could positively regulate OsPP2C49, and overexpression of OsPP2C49 in rice resulted in significantly decreased sensitivity of the abscisic acid (ABA) response and rapid dehydration. Moreover, OsNCED4 (9-cis-epoxycarotenoid dioxygenase4), a key gene in ABA biosynthesis, was also positively regulated by OsbZIP23. Together, our results suggest that OsbZIP23 acts as a central regulator in ABA signaling and biosynthesis, and drought resistance in rice. PMID:27325665

  14. Carbon nanoparticles induce ceramide- and lipid raft-dependent signalling in lung epithelial cells: a target for a preventive strategy against environmentally-induced lung inflammation

    Directory of Open Access Journals (Sweden)

    Peuschel Henrike

    2012-12-01

    Full Text Available Abstract Background Particulate air pollution in lung epithelial cells induces pathogenic endpoints like proliferation, apoptosis, and pro-inflammatory reactions. The activation of the epidermal growth factor receptor (EGFR is a key event responsible for signalling events involving mitogen activated protein kinases specific for these endpoints. The molecular events leading to receptor activation however are not well understood. These events are relevant for the toxicological evaluation of inhalable particles as well as for potential preventive strategies in situations when particulate air pollution cannot be avoided. The current study therefore had the objective to elucidate membrane-coupled events leading to EGFR activation and the subsequent signalling cascade in lung epithelial cells. Furthermore, we aimed to identify the molecular target of ectoine, a biophysical active substance which we described to prevent carbon nanoparticle-induced lung inflammation. Methods Membrane signalling events were investigated in isolated lipid rafts from lung epithelial cells with regard to lipid and protein content of the signalling platforms. Using positive and negative intervention approaches, lipid raft changes, subsequent signalling events, and lung inflammation were investigated in vitro in lung epithelial cells (RLE-6TN and in vivo in exposed animals. Results Carbon nanoparticle treatment specifically led to an accumulation of ceramides in lipid rafts. Detailed analyses demonstrated a causal link of ceramides and subsequent EGFR activation coupled with a loss of the receptor in the lipid raft fractions. In vitro and in vivo investigations demonstrate the relevance of these events for carbon nanoparticle-induced lung inflammation. Moreover, the compatible solute ectoine was able to prevent ceramide-mediated EGFR phosphorylation and subsequent signalling as well as lung inflammation in vivo. Conclusion The data identify a so far unknown event in pro

  15. Combined genome-wide expression profiling and targeted RNA interference in primary mouse macrophages reveals perturbation of transcriptional networks associated with interferon signalling

    Directory of Open Access Journals (Sweden)

    Craigon Marie

    2009-08-01

    Full Text Available Abstract Background Interferons (IFNs are potent antiviral cytokines capable of reprogramming the macrophage phenotype through the induction of interferon-stimulated genes (ISGs. Here we have used targeted RNA interference to suppress the expression of a number of key genes associated with IFN signalling in murine macrophages prior to stimulation with interferon-gamma. Genome-wide changes in transcript abundance caused by siRNA activity were measured using exon-level microarrays in the presence or absence of IFNγ. Results Transfection of murine bone-marrow derived macrophages (BMDMs with a non-targeting (control siRNA and 11 sequence-specific siRNAs was performed using a cationic lipid transfection reagent (Lipofectamine2000 prior to stimulation with IFNγ. Total RNA was harvested from cells and gene expression measured on Affymetrix GeneChip Mouse Exon 1.0 ST Arrays. Network-based analysis of these data revealed six siRNAs to cause a marked shift in the macrophage transcriptome in the presence or absence IFNγ. These six siRNAs targeted the Ifnb1, Irf3, Irf5, Stat1, Stat2 and Nfkb2 transcripts. The perturbation of the transcriptome by the six siRNAs was highly similar in each case and affected the expression of over 600 downstream transcripts. Regulated transcripts were clustered based on co-expression into five major groups corresponding to transcriptional networks associated with the type I and II IFN response, cell cycle regulation, and NF-KB signalling. In addition we have observed a significant non-specific immune stimulation of cells transfected with siRNA using Lipofectamine2000, suggesting use of this reagent in BMDMs, even at low concentrations, is enough to induce a type I IFN response. Conclusion Our results provide evidence that the type I IFN response in murine BMDMs is dependent on Ifnb1, Irf3, Irf5, Stat1, Stat2 and Nfkb2, and that siRNAs targeted to these genes results in perturbation of key transcriptional networks associated

  16. miR-625-3p regulates oxaliplatin resistance by targeting MAP2K6-p38 signalling in human colorectal adenocarcinoma cells

    DEFF Research Database (Denmark)

    Rasmussen, Mads Heilskov; Lyskjær, Iben; Jersie-Christensen, Rosa Rakownikow

    2016-01-01

    Oxaliplatin resistance in colorectal cancers (CRC) is a major medical problem, and predictive markers are urgently needed. Recently, miR-625-3p was reported as a promising predictive marker. Herein, we show that miR-625-3p functionally induces oxaliplatin resistance in CRC cells, and identify...... the signalling networks affected by miR-625-3p. We show that the p38 MAPK activator MAP2K6 is a direct target of miR-625-3p, and, accordingly, is downregulated in non-responder patients of oxaliplatin therapy. miR-625-3p-mediated resistance is reversed by anti-miR-625-3p treatment and ectopic expression of a mi......R-625-3p insensitive MAP2K6 variant. In addition, reduction of p38 signalling by using siRNAs, chemical inhibitors or expression of a dominant-negative MAP2K6 protein induces resistance to oxaliplatin. Transcriptome, proteome and phosphoproteome profiles confirm inactivation of MAP2K6-p38 signalling...

  17. Killing two birds with one stone: dual blockade of integrin and FGF signaling through targeting syndecan-4 in postoperative capsular opacification.

    Science.gov (United States)

    Qin, Yingyan; Zhu, Yi; Luo, Furong; Chen, Chuan; Chen, Xiaoyun; Wu, Mingxing

    2017-07-13

    The most common complication after cataract surgery is postoperative capsular opacification, which includes anterior capsular opacification (ACO) and posterior capsular opacification (PCO). Increased adhesion of lens epithelial cells (LECs) to the intraocular lens material surface promotes ACO formation, whereas proliferation and migration of LECs to the posterior capsule lead to the development of PCO. Cell adhesion is mainly mediated by the binding of integrin to extracellular matrix proteins, while cell proliferation and migration are regulated by fibroblast growth factor (FGF). Syndecan-4 (SDC-4) is a co-receptor for both integrin and FGF signaling pathways. Therefore, SDC-4 may be an ideal therapeutic target for the prevention and treatment of postoperative capsular opacification. However, how SDC-4 contributes to FGF-mediated proliferation, migration, and integrin-mediated adhesion of LECs is unclear. Here, we found that downregulation of SDC-4 inhibited FGF signaling through the blockade of ERK1/2 and PI3K/Akt/mTOR activation, thus suppressing cell proliferation and migration. In addition, downregulation of SDC-4 suppressed integrin-mediated cell adhesion through inhibiting focal adhesion kinase (FAK) phosphorylation. Moreover, SDC-4 knockout mice exhibited normal lens morphology, but had significantly reduced capsular opacification after injury. Finally, SDC-4 expression level was increased in the anterior capsule LECs of age-related cataract patients. Taken together, we for the first time characterized the key regulatory role of SDC-4 in FGF and integrin signaling in human LECs, and provided the basis for future pharmacological interventions of capsular opacification.

  18. 2,5-Dimethyl-Celecoxib Extends Drosophila Life Span via a Mechanism That Requires Insulin and Target of Rapamycin Signaling.

    Science.gov (United States)

    Wu, Qi; Lian, Ting; Fan, Xiaolan; Song, Chaochun; Gaur, Uma; Mao, Xueping; Yang, Deying; Piper, Matthew D W; Yang, Mingyao

    2017-10-01

    The search for antiaging drugs is a key component of gerontology research. A few drugs with positive effects on life span in model organisms have been found. Here, we report that 2,5-dimethyl-celecoxib, a derivative of the anti-inflammatory drug celecoxib, can extend Drosophila life span and delay aging by a mechanism involving insulin signaling and target of rapamycin signaling. Importantly, its positive effects were apparent when the treatment window was restricted to the beginning of life or the later half. 2,5-Dimethyl-celecoxib-induced longevity was also associated with improvements in physical activity, intestinal integrity, and increased autophagy. In addition, 2,5-dimethyl-celecoxib exhibited protective effects against several kinds of stress such as starvation and heat. The generally positive effects of 2,5-dimethyl-celecoxib on both health and life span, combined with its mode of action via evolutionarily conserved signaling pathways, indicate that it has the potential to become an effective antiaging drug. © The Author 2016. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  19. The Role of Wnt/β-Catenin Signaling Pathway in Disrupted Hippocampal Neurogenesis of Temporal Lobe Epilepsy: A Potential Therapeutic Target?

    Science.gov (United States)

    Huang, Cheng; Fu, Xiang-Hui; Zhou, Dong; Li, Jin-Mei

    2015-07-01

    Temporal lobe epilepsy is one of the most common clinical neurological disorders. One of the major pathological findings in temporal lobe epilepsy is hippocampal sclerosis, characterized by massive neuronal loss and severe gliosis. The epileptogenesis process of temporal lobe epilepsy usually starts with initial precipitating insults, followed by neurodegeneration, abnormal hippocampus circuitry reorganization, and the formation of hypersynchronicity. Experimental and clinical evidence strongly suggests that dysfunctional neurogenesis is involved in the epileptogenesis. Recent data demonstrate that neurogenesis is induced by acute seizures or precipitating insults, whereas the capacity of neuronal recruitment and proliferation substantially decreases in the chronic phase of epilepsy. Participation of the Wnt/β-catenin signaling pathway in neurogenesis reveals its importance in epileptogenesis; its dysfunction contributes to the structural and functional abnormality of temporal lobe epilepsy, while rescuing this pathway exerts neuroprotective effects. Here, we summarize data supporting the involvement of Wnt/β-catenin signaling in the epileptogenesis of temporal lobe epilepsy. We also propose that the Wnt/β-catenin signaling pathway may serve as a promising therapeutic target for temporal lobe epilepsy treatment.

  20. Improving nelarabine efficacy in T cell acute lymphoblastic leukemia by targeting aberrant PI3K/AKT/mTOR signaling pathway

    Directory of Open Access Journals (Sweden)

    Annalisa Lonetti

    2016-10-01

    Full Text Available Abstract Background Although in recent years, the introduction of novel chemotherapy protocols has improved the outcome of T cell acute lymphoblastic leukemia (T-ALL patients, refractory and/or relapsing disease remains a foremost concern. In this context, a major contribution was provided by the introduction of the nucleoside analog nelarabine, approved for salvage treatment of T-ALL patients with refractory/relapsed disease. However, nelarabine could induce a life-threatening, dose-dependent neurotoxicity. To improve nelarabine efficacy, we have analyzed its molecular targets, testing selective inhibitors of such targets in combination with nelarabine. Methods The effectiveness of nelarabine as single agent or in combination with PI3K, Bcl2, and MEK inhibitors was evaluated on human T-ALL cell lines and primary T-ALL refractory/relapsed lymphoblasts. The efficacy of signal modulators in terms of cytotoxicity, induction of apoptosis, and changes in gene and protein expression was assessed by flow cytometry, western blotting, and quantitative real-time PCR in T-ALL settings. Results Treatment with nelarabine as a single agent identified two groups of T-ALL cell lines, one sensitive and one resistant to the drug. Whereas sensitive T-ALL cells showed a significant increase of apoptosis and a strong down-modulation of PI3K signaling, resistant T-ALL cells showed a hyperactivation of AKT and MEK/ERK1/2 signaling pathways, not caused by differences in the expression of nelarabine transporters or metabolic activators. We then studied the combination of nelarabine with the PI3K inhibitors (both pan and dual γ/δ inhibitors, with the Bcl2 specific inhibitor ABT199, and with the MEK inhibitor trametinib on both T-ALL cell lines and patient samples at relapse, which displayed constitutive activation of PI3K signaling and resistance to nelarabine alone. The combination with the pan PI3K inhibitor ZSTK-474 was the most effective in inhibiting the growth of

  1. A peptide targeting an interaction interface disrupts the dopamine D1-D2 receptor heteromer to block signaling and function in vitro and in vivo: effective selective antagonism

    Science.gov (United States)

    Hasbi, Ahmed; Perreault, Melissa L.; Shen, Maurice Y. F.; Zhang, Lucia; To, Ryan; Fan, Theresa; Nguyen, Tuan; Ji, Xiaodong; O'Dowd, Brian F.; George, Susan R.

    2014-01-01

    Although the dopamine D1-D2 receptor heteromer has emerging physiological relevance and a postulated role in different neuropsychiatric disorders, such as drug addiction, depression, and schizophrenia, there is a need for pharmacological tools that selectively target such receptor complexes in order to analyze their biological and pathophysiological functions. Since no selective antagonists for the D1-D2 heteromer are available, serial deletions and point mutations were used to precisely identify the amino acids involved in an interaction interface between the receptors, residing within the carboxyl tail of the D1 receptor that interacted with the D2 receptor to form the D1-D2 receptor heteromer. It was determined that D1 receptor carboxyl tail residues 404Glu and 405Glu were critical in mediating the interaction with the D2 receptor. Isolated mutation of these residues in the D1 receptor resulted in the loss of agonist activation of the calcium signaling pathway mediated through the D1-D2 receptor heteromer. The physical interaction between the D1 and D2 receptor could be disrupted, as shown by coimmunoprecipitation and BRET analysis, by a small peptide generated from the D1 receptor sequence that contained these amino acids, leading to a switch in G-protein affinities and loss of calcium signaling, resulting in the inhibition of D1-D2 heteromer function. The use of the D1-D2 heteromer-disrupting peptide in vivo revealed a pathophysiological role for the D1-D2 heteromer in the modulation of behavioral despair. This peptide may represent a novel pharmacological tool with potential therapeutic benefits in depression treatment.—Hasbi, A., Perreault, M. L., Shen, M. Y. F., Zhang, L., To, R., Fan, T., Nguyen, T., Ji, X., O'Dowd, B. F., George, S. R. A peptide targeting an interaction interface disrupts the dopamine D1-D2 receptor heteromer to block signaling and function in vitro and in vivo: effective selective antagonism. PMID:25063849

  2. Three non-autonomous signals collaborate for nuclear targeting of CrMYC2, a Catharanthus roseus bHLH transcription factor

    Directory of Open Access Journals (Sweden)

    Gantet Pascal

    2010-11-01

    Full Text Available Abstract Background CrMYC2 is an early jasmonate-responsive bHLH transcription factor involved in the regulation of the expression of the genes of the terpenic indole alkaloid biosynthesis pathway in Catharanthus roseus. In this paper, we identified the amino acid domains necessary for the nuclear targeting of CrMYC2. Findings We examined the intracellular localization of whole CrMYC2 and of various deletion mutants, all fused with GFP, using a transient expression assay in onion epidermal cells. Sequence analysis of this protein revealed the presence of four putative basic nuclear localization signals (NLS. Assays showed that none of the predicted NLS is active alone. Further functional dissection of CrMYC2 showed that the nuclear targeting of this transcription factor involves the cooperation of three domains located in the C-terminal region of the protein. The first two domains are located at amino acid residues 454-510 and 510-562 and contain basic classical monopartite NLSs; these regions are referred to as NLS3 (KRPRKR and NLS4 (EAERQRREK, respectively. The third domain, between residues 617 and 652, is rich in basic amino acids that are well conserved in other phylogenetically related bHLH transcription factors. Our data revealed that these three domains are inactive when isolated but act cooperatively to target CrMYC2 to the nucleus. Conclusions This study identified three amino acid domains that act in cooperation to target the CrMYC2 transcription factor to the nucleus. Further fine structure/function analysis of these amino acid domains will allow the identification of new NLS domains and will allow the investigation of the related molecular mechanisms involved in the nuclear targeting of the CrMYC2 bHLH transcription factor.

  3. Redirecting the Cyanobacterial Bicarbonate Transporters BicA and SbtA to the Chloroplast Envelope: Soluble and Membrane Cargos Need Different Chloroplast Targeting Signals in Plants.

    Directory of Open Access Journals (Sweden)

    Vivien eRolland

    2016-02-01

    Full Text Available Most major crops used for human consumption are C3 plants, which yields are limited by photosynthetic inefficiency. To circumvent this, it has been proposed to implement the cyanobacterial CO2-concentrating mechanism (CCM, principally consisting of bicarbonate transporters and carboxysomes, into plant chloroplasts. As it is currently not possible to recover homoplasmic transplastomic monocots, foreign genes must be introduced in these plants via nuclear transformation. Consequently, it is paramount to ensure that resulting proteins reach the appropriate sub-cellular compartment, which for cyanobacterial transporters BicA and SbtA, is the chloroplast inner-envelope membrane (IEM. At present, targeting signals to redirect large transmembrane proteins from non-chloroplastic organisms to plant chloroplast envelopes are unknown. The goal of this study was to identify such signals, using agrobacteria-mediated transient expression and confocal microscopy to determine the sub-cellular localization of ~37 GFP-tagged chimeras. Initially, fragments of chloroplast proteins known to target soluble cargos to the stroma were tested for their ability to redirect BicA, but they proved ineffective. Next, different N-terminal regions from Arabidopsis IEM transporters were tested. We demonstrated that the N-terminus of AtHP59, AtPLGG1 or AtNTT1 (92-115 amino acids, containing a cleavable chloroplast transit peptide (cTP and a membrane protein leader (MPL, was sufficient to redirect BicA or SbtA to the chloroplast envelope. This constitutes the first evidence that nuclear-encoded transmembrane proteins from non-chloroplastic organisms can be targeted to the envelope of plant chloroplasts; a finding which represents an important advance in chloroplast engineering by opening up the door to further manipulation of the chloroplastic envelope.

  4. Impact of targeted education on managing warning and error signals by children and adolescents with type 1 diabetes using the Accu-Chek Combo Insulin Pump System

    Directory of Open Access Journals (Sweden)

    Asma Deeb

    2017-09-01

    Full Text Available Purpose Insulin pumps are widely used in diabetes. They are equipped with safety alarms to alert users. Pump manuals contain alarm codes and how to troubleshoot them. However, these manuals are lengthy and difficult to use, particularly in emergencies. We aim to assess the impact of targeted education on warnings and errors in improving competency to troubleshoot the alarms. Methods Twenty-one patients, with a median age of 13, were recruited over a 5-month period. Each patient had 2 study visits. The frequencies and types of alarms were recorded, and patients were given a summary sheet that outlined common alarms encountered and troubleshooting tips. In visit 2, the frequencies and types of alarms were compared to those of visit 1. The patients were asked to fill a questionnaire and to rate the education session given in visit 1, their level of competency in decrypting alarm codes, and their promptness in responding to alarms. Results Low cartridge (W1, low battery (W2, and bolus cancelled (W8 were the commonest warnings. The most noted errors were occlusion (E4, power interruption (E8, empty battery (E2, set not primed (E11, and cartridge empty (E1. The numbers of warning and error signals markedly decreased after targeted education (P<0.05. The ability in decrypting warning signals significantly improved (P=0.02, and the frequency of response to pump alarms significantly increased (P=0.001. Conclusions Certain warnings and errors are more common than others in insulin pumps. Targeted education is useful in improving competency and response of patients in managing pump alarms.

  5. Impact of targeted education on managing warning and error signals by children and adolescents with type 1 diabetes using the Accu-Chek Combo Insulin Pump System.

    Science.gov (United States)

    Deeb, Asma; Qahtani, Nabras Al; Ali, Amal Al; Akle, Mariette

    2017-09-01

    Insulin pumps are widely used in diabetes. They are equipped with safety alarms to alert users. Pump manuals contain alarm codes and how to troubleshoot them. However, these manuals are lengthy and difficult to use, particularly in emergencies. We aim to assess the impact of targeted education on warnings and errors in improving competency to troubleshoot the alarms. Twenty-one patients, with a median age of 13, were recruited over a 5-month period. Each patient had 2 study visits. The frequencies and types of alarms were recorded, and patients were given a summary sheet that outlined common alarms encountered and troubleshooting tips. In visit 2, the frequencies and types of alarms were compared to those of visit 1. The patients were asked to fill a questionnaire and to rate the education session given in visit 1, their level of competency in decrypting alarm codes, and their promptness in responding to alarms. Low cartridge (W1), low battery (W2), and bolus cancelled (W8) were the commonest warnings. The most noted errors were occlusion (E4), power interruption (E8), empty battery (E2), set not primed (E11), and cartridge empty (E1). The numbers of warning and error signals markedly decreased after targeted education (Pwarning signals significantly improved (P=0.02), and the frequency of response to pump alarms significantly increased (P=0.001). Certain warnings and errors are more common than others in insulin pumps. Targeted education is useful in improving competency and response of patients in managing pump alarms.

  6. TFII-I regulates target genes in the PI-3K and TGF-β signaling pathways through a novel DNA binding motif.

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    Segura-Puimedon, Maria; Borralleras, Cristina; Pérez-Jurado, Luis A; Campuzano, Victoria

    2013-09-25

    General transcription factor (TFII-I) is a multi-functional protein involved in the transcriptional regulation of critical developmental genes, encoded by the GTF2I gene located on chromosome 7q11.23. Haploinsufficiency at GTF2I has been shown to play a major role in the neurodevelopmental features of Williams-Beuren syndrome (WBS). Identification of genes regulated by TFII-I is thus critical to detect molecular determinants of WBS as well as to identify potential new targets for specific pharmacological interventions, which are currently absent. We performed a microarray screening for transcriptional targets of TFII-I in cortex and embryonic cells from Gtf2i mutant and wild-type mice. Candidate genes with altered expression were verified using real-time PCR. A novel motif shared by deregulated genes was found and chromatin immunoprecipitation assays in embryonic fibroblasts were used to document in vitro TFII-I binding to this motif in the promoter regions of deregulated genes. Interestingly, the PI3K and TGFβ signaling pathways were over-represented among TFII-I-modulated genes. In this study we have found a highly conserved DNA element, common to a set of genes regulated by TFII-I, and identified and validated novel in vivo neuronal targets of this protein affecting the PI3K and TGFβ signaling pathways. Overall, our data further contribute to unravel the complexity and variability of the different genetic programs orchestrated by TFII-I. © 2013 Elsevier B.V. All rights reserved.

  7. Human Cytomegalovirus miR-UL112-3p Targets TLR2 and Modulates the TLR2/IRAK1/NFκB Signaling Pathway.

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    Igor Landais

    2015-05-01

    Full Text Available Human Cytomegalovirus (HCMV encodes multiple microRNAs (miRNAs whose functions are just beginning to be uncovered. Using in silico approaches, we identified the Toll-Like Receptor (TLR innate immunity pathway as a possible target of HCMV miRNAs. Luciferase reporter assay screens further identified TLR2 as a target of HCMV miR-UL112-3p. TLR2 plays a major role in innate immune response by detecting both bacterial and viral ligands, including HCMV envelope proteins gB and gH. TLR2 activates a variety of signal transduction routes including the NFκB pathway. Furthermore, TLR2 plays an important role in controlling CMV infection both in humans and in mice. Immunoblot analysis of cells transfected with a miR-UL112-3p mimic revealed that endogenous TLR2 is down-regulated by miR-UL112-3p with similar efficiency as a TLR2-targeting siRNA (siTLR2. We next found that TLR2 protein level decreases at late times during HCMV infection and correlates with miR-UL112-3p accumulation in fibroblasts and monocytic THP1 cells. Confirming direct miR-UL112-3p targeting, down-regulation of endogenous TLR2 was not observed in cells infected with HCMV mutants deficient in miR-UL112-3p expression, but transfection of miR-UL112-3p in these cells restored TLR2 down-regulation. Using a NFκB reporter cell line, we found that miR-UL112-3p transfection significantly inhibited NFκB-dependent luciferase activity with similar efficiency as siTLR2. Consistent with this observation, miR-UL112-3p transfection significantly reduced the expression of multiple cytokines (IL-1β, IL-6 and IL-8 upon stimulation with a TLR2 agonist. Finally, miR-UL112-3p transfection significantly inhibited the TLR2-induced post-translational activation of IRAK1, a kinase located in the upstream section of the TLR2/NFκB signaling axis. To our knowledge, this is the first identified mechanism of TLR2 modulation by HCMV and is the first report of functional targeting of TLR2 by a viral miRNA. These

  8. Extracellular Signal-Regulated Kinase Is a Direct Target of the Anti-Inflammatory Compound Amentoflavone Derived from Torreya nucifera

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    Jueun Oh

    2013-01-01

    Full Text Available Amentoflavone is a biflavonoid compound with antioxidant, anticancer, antibacterial, antiviral, anti-inflammatory, and UV-blocking activities that can be isolated from Torreya nucifera, Biophytum sensitivum, and Selaginella tamariscina. In this study, the molecular mechanism underlying amentoflavone’s anti-inflammatory activity was investigated. Amentoflavone dose dependently suppressed the production of nitric oxide (NO and prostaglandin E2 (PGE2 in RAW264.7 cells stimulated with the TLR4 ligand lipopolysaccharide (LPS; derived from Gram-negative bacteria. Amentoflavone suppressed the nuclear translocation of c-Fos, a subunit of activator protein (AP-1, at 60 min after LPS stimulation and inhibited the activity of purified and immunoprecipitated extracellular signal-regulated kinase (ERK, which mediates c-Fos translocation. In agreement with these results, amentoflavone also suppressed the formation of a molecular complex including ERK and c-Fos. Therefore, our data strongly suggest that amentoflavone’s immunopharmacological activities are due to its direct effect on ERK.

  9. Targeting Hedgehog signaling pathway and autophagy overcomes drug resistance of BCR-ABL-positive chronic myeloid leukemia.

    Science.gov (United States)

    Zeng, Xian; Zhao, Hui; Li, Yubin; Fan, Jiajun; Sun, Yun; Wang, Shaofei; Wang, Ziyu; Song, Ping; Ju, Dianwen

    2015-01-01

    The frontline tyrosine kinase inhibitor (TKI) imatinib has revolutionized the treatment of patients with chronic myeloid leukemia (CML). However, drug resistance is the major clinical challenge in the treatment of CML. The Hedgehog (Hh) signaling pathway and autophagy are both related to tumorigenesis, cancer therapy, and drug resistance. This study was conducted to explore whether the Hh pathway could regulate autophagy in CML cells and whether simultaneously regulating the Hh pathway and autophagy could induce cell death of drug-sensitive or -resistant BCR-ABL(+) CML cells. Our results indicated that pharmacological or genetic inhibition of Hh pathway could markedly induce autophagy in BCR-ABL(+) CML cells. Autophagic inhibitors or ATG5 and ATG7 silencing could significantly enhance CML cell death induced by Hh pathway suppression. Based on the above findings, our study demonstrated that simultaneously inhibiting the Hh pathway and autophagy could markedly reduce cell viability and induce apoptosis of imatinib-sensitive or -resistant BCR-ABL(+) cells. Moreover, this combination had little cytotoxicity in human peripheral blood mononuclear cells (PBMCs). Furthermore, this combined strategy was related to PARP cleavage, CASP3 and CASP9 cleavage, and inhibition of the BCR-ABL oncoprotein. In conclusion, this study indicated that simultaneously inhibiting the Hh pathway and autophagy could potently kill imatinib-sensitive or -resistant BCR-ABL(+) cells, providing a novel concept that simultaneously inhibiting the Hh pathway and autophagy might be a potent new strategy to overcome CML drug resistance.

  10. SCFFbw7 Modulates the NFκB Signaling Pathway by Targeting NFκB2 for Ubiquitination and Destruction

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    Hidefumi Fukushima

    2012-05-01

    Full Text Available The NFκB/Rel family of proteins play critical roles in a variety of cellular processes. Thus, their physiological activation is tightly controlled. Recently, the NFκB2/p100 precursor has been characterized as the fourth IκB type of suppressor for NFκB. However, the molecular mechanism(s underlying regulated destruction of NFκB2 remains largely unknown. Here, we report that, unlike other IκBs, ubiquitination and destruction of NFκB2 are governed by SCFFbw7 in a GSK3-dependent manner. In Fbw7−/− cells, elevated expression of NFκB2/p100 leads to a subsequent reduction in NFκB signaling pathways and elevated sensitivity to TNFα-induced cell death. Reintroducing wild-type Fbw7, but not disease-derived mutant forms of Fbw7, rescues NFκB activity. Furthermore, T cell-specific depletion of Fbw7 also leads to reduced NFκB activity and perturbed T cell differentiation. Therefore, our work identifies Fbw7 as a physiological E3 ligase controlling NFκB2′s stability. It further implicates that Fbw7 might exert its tumor-suppressor function by regulating NFκB activity.

  11. Cardiac myosin binding protein-C as a central target of cardiac sarcomere signaling: a special mini-review series

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    de Tombe, Pieter P.

    2013-01-01

    Cardiac myosin binding protein-C (cMyBP-C) is a cardiac-specific thick filament assembly, accessory and regulatory protein. Physiologically, it is a key regulator of cardiac contractility. With more than two hundred mutations in the cMyBP-C gene directly linked to the development of cardiomyopathy and heart failure, cMyBP-C clearly plays a critical role in heart function. At baseline, cMyBP-C is highly phosphorylated, a condition required for normal cardiac function. However, the level of cMyBP-C phosphorylation is significantly decreased during heart failure, indicating that the level of cMyBP-C phosphorylation is directly linked to signaling and cardiac function. Early studies indicated that cMyBP-C interacts with myosin and titin, whereas studies now show that it also interacts with thin filament proteins. However, the exact role(s) of cMyBP-C in the heart remain(s) to be elucidated. As such, we invited experts in the field of cardiac muscle to identify and address key issues related to cMyBP-C by contributing a mini-review on such topics as structure, function, regulation, cardiomyopathy, proteolysis, and gene therapy. Starting from this issue, Pflügers Archiv European Journal of Physiology will publish two invited mini-review articles each month to discuss the most recent advances in the study of cMyBP-C. PMID:24196566

  12. Cardiac myosin binding protein-C as a central target of cardiac sarcomere signaling: a special mini review series.

    Science.gov (United States)

    Sadayappan, Sakthivel; de Tombe, Pieter P

    2014-02-01

    Cardiac myosin binding protein-C (cMyBP-C) is a cardiac-specific thick filament assembly, accessory, and regulatory protein. Physiologically, it is a key regulator of cardiac contractility. With more than 200 mutations in the cMyBP-C gene directly linked to the development of cardiomyopathy and heart failure, cMyBP-C clearly plays a critical role in heart function. At baseline, cMyBP-C is highly phosphorylated, a condition required for normal cardiac function. However, the level of cMyBP-C phosphorylation is significantly decreased during heart failure, indicating that the level of cMyBP-C phosphorylation is directly linked to signaling and cardiac function. Early studies indicated that cMyBP-C interacts with myosin and titin, whereas studies now show that it also interacts with thin filament proteins. However, the exact role(s) of cMyBP-C in the heart remain(s) to be elucidated. As such, we invited experts in the field of cardiac muscle to identify and address key issues related to cMyBP-C by contributing a mini review on such topics as structure, function, regulation, cardiomyopathy, proteolysis, and gene therapy. Starting from this issue, Pflügers Archiv European Journal of Physiology will publish two invited mini review articles each month to discuss the most recent advances in the study of cMyBP-C.

  13. Non-canonical peroxisome targeting signals: identification of novel PTS1 tripeptides and characterization of enhancer elements by computational permutation analysis

    Science.gov (United States)

    2012-01-01

    Background High-accuracy prediction tools are essential in the post-genomic era to define organellar proteomes in their full complexity. We recently applied a discriminative machine learning approach to predict plant proteins carrying peroxisome targeting signals (PTS) type 1 from genome sequences. For Arabidopsis thaliana 392 gene models were predicted to be peroxisome-targeted. The predictions were extensively tested in vivo, resulting in a high experimental verification rate of Arabidopsis proteins previously not known to be peroxisomal. Results In this study, we experimentally validated the predictions in greater depth by focusing on the most challenging Arabidopsis proteins with unknown non-canonical PTS1 tripeptides and prediction scores close to the threshold. By in vivo subcellular targeting analysis, three novel PTS1 tripeptides (QRL>, SQM>, and SDL>) and two novel tripeptide residues (Q at position −3 and D at pos. -2) were identified. To understand why, among many Arabidopsis proteins carrying the same C-terminal tripeptides, these proteins were specifically predicted as peroxisomal, the residues upstream of the PTS1 tripeptide were computationally permuted and the changes in prediction scores were analyzed. The newly identified Arabidopsis proteins were found to contain four to five amino acid residues of high predicted targeting enhancing properties at position −4 to −12 in front of the non-canonical PTS1 tripeptide. The identity of the predicted targeting enhancing residues was unexpectedly diverse, comprising besides basic residues also proline, hydroxylated (Ser, Thr), hydrophobic (Ala, Val), and even acidic residues. Conclusions Our computational and experimental analyses demonstrate that the plant PTS1 tripeptide motif is more diverse than previously thought, including an increasing number of non-canonical sequences and allowed residues. Specific targeting enhancing elements can be predicted for particular sequences of interest and are far

  14. Non-canonical peroxisome targeting signals: identification of novel PTS1 tripeptides and characterization of enhancer elements by computational permutation analysis

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    Chowdhary Gopal

    2012-08-01

    Full Text Available Abstract Background High-accuracy prediction tools are essential in the post-genomic era to define organellar proteomes in their full complexity. We recently applied a discriminative machine learning approach to predict plant proteins carrying peroxisome targeting signals (PTS type 1 from genome sequences. For Arabidopsis thaliana 392 gene models were predicted to be peroxisome-targeted. The predictions were extensively tested in vivo, resulting in a high experimental verification rate of Arabidopsis proteins previously not known to be peroxisomal. Results In this study, we experimentally validated the predictions in greater depth by focusing on the most challenging Arabidopsis proteins with unknown non-canonical PTS1 tripeptides and prediction scores close to the threshold. By in vivo subcellular targeting analysis, three novel PTS1 tripeptides (QRL>, SQM>, and SDL> and two novel tripeptide residues (Q at position −3 and D at pos. -2 were identified. To understand why, among many Arabidopsis proteins carrying the same C-terminal tripeptides, these proteins were specifically predicted as peroxisomal, the residues upstream of the PTS1 tripeptide were computationally permuted and the changes in prediction scores were analyzed. The newly identified Arabidopsis proteins were found to contain four to five amino acid residues of high predicted targeting enhancing properties at position −4 to −12 in front of the non-canonical PTS1 tripeptide. The identity of the predicted targeting enhancing residues was unexpectedly diverse, comprising besides basic residues also proline, hydroxylated (Ser, Thr, hydrophobic (Ala, Val, and even acidic residues. Conclusions Our computational and experimental analyses demonstrate that the plant PTS1 tripeptide motif is more diverse than previously thought, including an increasing number of non-canonical sequences and allowed residues. Specific targeting enhancing elements can be predicted for particular sequences

  15. miR-144 attenuates the host response to influenza virus by targeting the TRAF6-IRF7 signaling axis.

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    Carrie M Rosenberger

    2017-04-01

    Full Text Available Antiviral responses must rapidly defend against infection while minimizing inflammatory damage, but the mechanisms that regulate the magnitude of response within an infected cell are not well understood. miRNAs are small non-coding RNAs that suppress protein levels by binding target sequences on their cognate mRNA. Here, we identify miR-144 as a negative regulator of the host antiviral response. Ectopic expression of miR-144 resulted in increased replication of three RNA viruses in primary mouse lung epithelial cells: influenza virus, EMCV, and VSV. We identified the transcriptional network regulated by miR-144 and demonstrate that miR-144 post-transcriptionally suppresses TRAF6 levels. In vivo ablation of miR-144 reduced influenza virus replication in the lung and disease severity. These data suggest that miR-144 reduces the antiviral response by attenuating the TRAF6-IRF7 pathway to alter the cellular antiviral transcriptional landscape.

  16. Differentiating signals to make biological sense - A guide through databases for MS-based non-targeted metabolomics.

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    Gil de la Fuente, Alberto; Grace Armitage, Emily; Otero, Abraham; Barbas, Coral; Godzien, Joanna

    2017-09-01

    Metabolite identification is one of the most challenging steps in metabolomics studies and reflects one of the greatest bottlenecks in the entire workflow. The success of this step determines the success of the entire research, therefore the quality at which annotations are given requires special attention. A variety of tools and resources are available to aid metabolite identification or annotation, offering different and often complementary functionalities. In preparation for this article, almost 50 databases were reviewed, from which 17 were selected for discussion, chosen for their online ESI-MS functionality. The general characteristics and functions of each database is discussed in turn, considering the advantages and limitations of each along with recommendations for optimal use of each tool, as derived from experiences encountered at the Centre for Metabolomics and Bioanalysis (CEMBIO) in Madrid. These databases were evaluated considering their utility in non-targeted metabolomics, including aspects such as identifier assignment, structural assignment and interpretation of results. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Targeting NF-kappa B Signaling by Artesunate Restores Sensitivity of Castrate-Resistant Prostate Cancer Cells to Antiandrogens

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    Jessica J. Nunes

    2017-04-01

    Full Text Available Androgen deprivation therapy (ADT is the most preferred treatment for men with metastatic prostate cancer (PCa. However, the disease eventually progresses and develops resistance to ADT in majority of the patients, leading to the emergence of metastatic castration-resistant prostate cancer (mCRPC. Here, we assessed artesunate (AS, an artemisinin derivative, for its anticancer properties and ability to alleviate resistance to androgen receptor (AR antagonists. We have shown AS in combination with bicalutamide (Bic attenuates the oncogenic properties of the castrate-resistant (PC3, 22RV1 and androgen-responsive (LNCaP PCa cells. Mechanistically, AS and Bic combination inhibits nuclear factor (NF-κB signaling and decreases AR and/or AR-variant 7 expression via ubiquitin-mediated proteasomal degradation. The combination induces oxidative stress and apoptosis via survivin downregulation and caspase-3 activation, resulting in poly-ADP-ribose polymerase (PARP cleavage. Moreover, preclinical castrate-resistant PC3 xenograft studies in NOD/SCID mice (n =28, seven per group show remarkable tumor regression and significant reduction in lungs and bone metastases upon administering AS (50 mg/kg per day in two divided doses and Bic (50 mg/kg per day via oral gavage. Taken together, we for the first time provide a compelling preclinical rationale that AS could disrupt AR antagonist–mediated resistance observed in mCRPC. The current study also indicates that the therapeutic combination of Food and Drug Administration–approved AS or NF-κB inhibitors and AR antagonists may enhance the clinical efficacy in the treatment of mCRPC patients.

  18. Terpene metabolic engineering via nuclear or chloroplast genomes profoundly and globally impacts off-target pathways through metabolite signalling.

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    Pasoreck, Elise K; Su, Jin; Silverman, Ian M; Gosai, Sager J; Gregory, Brian D; Yuan, Joshua S; Daniell, Henry

    2016-09-01

    The impact of metabolic engineering on nontarget pathways and outcomes of metabolic engineering from different genomes are poorly understood questions. Therefore, squalene biosynthesis genes FARNESYL DIPHOSPHATE SYNTHASE (FPS) and SQUALENE SYNTHASE (SQS) were engineered via the Nicotiana tabacum chloroplast (C), nuclear (N) or both (CN) genomes to promote squalene biosynthesis. SQS levels were ~4300-fold higher in C and CN lines than in N, but all accumulated ~150-fold higher squalene due to substrate or storage limitations. Abnormal leaf and flower phenotypes, including lower pollen production and reduced fertility, were observed regardless of the compartment or level of transgene expression. Substantial changes in metabolomes of all lines were observed: levels of 65-120 unrelated metabolites, including the toxic alkaloid nicotine, changed by as much as 32-fold. Profound effects of transgenesis on nontarget gene expression included changes in the abundance of 19 076 transcripts by up to 2000-fold in CN; 7784 transcripts by up to 1400-fold in N; and 5224 transcripts by as much as 2200-fold in C. Transporter-related transcripts were induced, and cell cycle-associated transcripts were disproportionally repressed in all three lines. Transcriptome changes were validated by qRT-PCR. The mechanism underlying these large changes likely involves metabolite-mediated anterograde and/or retrograde signalling irrespective of the level of transgene expression or end product, due to imbalance of metabolic pools, offering new insight into both anticipated and unanticipated consequences of metabolic engineering. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  19. Preclinical evaluation of destruxin B as a novel Wnt signaling target suppressing proliferation and metastasis of colorectal cancer using non-invasive bioluminescence imaging

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    Yeh, Chi-Tai [Graduate Institute of Clinical Medicine, Taipei Medical University, Taipei, Taiwan (China); Center of Excellence for Cancer Research, Taipei Medical University, Taipei, Taiwan (China); Department of Surgery, Taipei Medical University-Shuang Ho Hospital, Taipei, Taiwan (China); Rao, Yerra Koteswara [Institute of Biochemical Sciences and Technology, Chaoyang University of Technology, Taichung, Taiwan (China); Ye, Min [Department of Natural Medicine, School of Pharmaceutical Sciences, Peking University, Beijing (China); Wu, Wen-Shi [Department of Horticulture and Biotechnology, Chinese Culture University, Taipei, Taiwan (China); Chang, Tung-Chen [Department of Surgery, Taipei Medical University-Shuang Ho Hospital, Taipei, Taiwan (China); Wang, Liang-Shun [Graduate Institute of Clinical Medicine, Taipei Medical University, Taipei, Taiwan (China); Division of Thoracic Surgery, Department of Surgery, Shuang Ho Hospital, Taipei Medical University, Taipei, Taiwan (China); Wu, Chih-Hsiung [Center of Excellence for Cancer Research, Taipei Medical University, Taipei, Taiwan (China); Department of Surgery, Taipei Medical University-Shuang Ho Hospital, Taipei, Taiwan (China); Wu, Alexander T.H., E-mail: chaw1211@tmu.edu.tw [Ph.D. Program for Translational Medicine, College of Medical Science and Technology, Taipei Medical University, Taipei, Taiwan (China); Department of Radiation Oncology, Taipei Medical University Hospital, Taipei, Taiwan (China); Tzeng, Yew-Min, E-mail: ymtzeng@cyut.edu.tw [Institute of Biochemical Sciences and Technology, Chaoyang University of Technology, Taichung, Taiwan (China)

    2012-05-15

    In continuation to our studies toward the identification of direct anti-cancer targets, here we showed that destruxin B (DB) from Metarhizium anisopliae suppressed the proliferation and induced cell cycle arrest in human colorectal cancer (CRC) HT29, SW480 and HCT116 cells. Additionally, DB induced apoptosis in HT29 cells by decreased expression level of anti-apoptotic proteins Bcl-2 and Bcl-xL while increased pro-apoptotic Bax. On the other hand, DB attenuated Wnt-signaling by downregulation of β-catenin, Tcf4 and β-catenin/Tcf4 transcriptional activity, concomitantly with decreased expression of β-catenin target genes cyclin D1, c-myc and survivin. Furthermore, DB affected the migratory and invasive ability of HT29 cells through suppressed MMPs-2 and -9 enzymatic activities. We also found that DB targeted the MAPK and/or PI3K/Akt pathway by reduced expression of Akt, IKK-α, JNK, NF-κB, c-Jun and c-Fos while increased that of IκBα. Finally, we demonstrated that DB inhibited tumorigenesis in HT29 xenograft mice using non-invasive bioluminescence technique. Consistently, tumor samples from DB-treated mice demonstrated suppressed expression of β-catenin, cyclin D1, survivin, and endothelial marker CD31 while increased caspase-3 expression. Collectively, our data supports DB as an inhibitor of Wnt/β-catenin/Tcf signaling pathway that may be beneficial in the CRC management. Highlights: ► Destruxin B (DB) inhibited colorectal cancer cells growth and induced apoptosis. ► MAPK and/or PI3K/Akt cascade cooperates in DB induced apoptosis. ► DB affected the migratory and invasive ability of HT29 cells through MMP-9. ► DB attenuated Wnt-signaling components β-catenin, Tcf4. ► DB attenuated cyclin D1, c-myc, survivin and tumorigenesis in HT29 xenograft mice.

  20. Induction of biogenic magnetization and redox control by a component of the target of rapamycin complex 1 signaling pathway.

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    Keiji Nishida

    Full Text Available Most organisms are simply diamagnetic, while magnetotactic bacteria and migratory animals are among organisms that exploit magnetism. Biogenic magnetization not only is of fundamental interest, but also has industrial potential. However, the key factor(s that enable biogenic magnetization in coordination with other cellular functions and metabolism remain unknown. To address the requirements for induction and the application of synthetic bio-magnetism, we explored the creation of magnetism in a simple model organism. Cell magnetization was first observed by attraction towards a magnet when normally diamagnetic yeast Saccharomyces cerevisiae were grown with ferric citrate. The magnetization was further enhanced by genetic modification of iron homeostasis and introduction of ferritin. The acquired magnetizable properties enabled the cells to be attracted to a magnet, and be trapped by a magnetic column. Superconducting quantum interference device (SQUID magnetometry confirmed and quantitatively characterized the acquired paramagnetism. Electron microscopy and energy-dispersive X-ray spectroscopy showed electron-dense iron-containing aggregates within the magnetized cells. Magnetization-based screening of gene knockouts identified Tco89p, a component of TORC1 (Target of rapamycin complex 1, as important for magnetization; loss of TCO89 and treatment with rapamycin reduced magnetization in a TCO89-dependent manner. The TCO89 expression level positively correlated with magnetization, enabling inducible magnetization. Several carbon metabolism genes were also shown to affect magnetization. Redox mediators indicated that TCO89 alters the intracellular redox to an oxidized state in a dose-dependent manner. Taken together, we demonstrated that synthetic induction of magnetization is possible and that the key factors are local redox control through carbon metabolism and iron supply.

  1. Cross-talk between freezing response and signaling for regulatory transcriptions of MIR475b and its targets by miR475b promoter in Populus suaveolens.

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    Niu, Jun; Wang, Jia; Hu, Huiwen; Chen, Yinlei; An, Jiyong; Cai, Jian; Sun, Runze; Sheng, Zhongting; Liu, Xieping; Lin, Shanzhi

    2016-02-08

    MicroRNAs (miRNAs) are small, non-coding RNAs that play important roles in post-transcriptional regulation of their target genes, yet the transcriptional regulation of plant miRNAs by promoter is poorly understood. Here, we firstly clone pri-miR475b cDNA and its native promoter from P. suaveolens, and characterize Psu-MIR475b as class-II gene transcribed by RNA polymerase II. By 5' deletion analysis of Psu-miR475b promoter in a series of promoter-GUS chimeric vectors, we functionally identify three positive regulatory regions and multiple cis-acting elements responsible for Psu-miR475b promoter activity in response to freezing stress and exogenous hormone treatment. Moreover, the Psu-miR475b promoter activity displays a tissue-specific manner, negatively regulated by freezing stress and positively by MeJA, SA or GA treatment. Importantly, we comparatively analyze the time-course transcriptional profiles of Psu-miR475b and its targets in Psu-miR475b over-expression transgenic plants controlled by Psu-miR475b-specific promoter or CaMV 35S constitutive promoter, and explore the regulatory mechanism of Psu-miR475b promoter controlling transcriptional expressions of Psu-MIR475b and its targets in response to freezing stress and exogenous hormone treatment. Our results reveal that Psu-miR475b promoter-mediated transcriptions of Psu-MIR475b and its targets in response to freezing stress may be involved in a cross-talk between freezing response and stress signaling process.

  2. miR-124 suppresses proliferation and invasion of nasopharyngeal carcinoma cells through the Wnt/β-catenin signaling pathway by targeting Capn4

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    Hu H

    2017-05-01

    Full Text Available Haili Hu,1,* Guanghui Wang,1,* Congying Li2 1Department of Otorhinolaryngology, Huaihe Hospital of Henan University, 2Department of Otorhinolaryngology, School of Medicine, Kaifeng University, Kaifeng, People’s Republic of China *These authors contributed equally to this work Background: Recent studies have demonstrated that microRNA 124 (miR-124 acts as a tumor suppressor in nasopharyngeal carcinoma (NPC; however, the exact molecular mechanism by which miR-124 exerts tumor suppression has not been well elucidated.Materials and methods: We performed quantitative real-time PCR (qRT-PCR to measure the expression of metastasis associated lung adenocarcinoma transcript 1, miR-124, and calpain small subunit 1 (Capn4 mRNAs in NPC cell lines. We also performed western blot analysis to detect the levels of Capn4. Furthermore, we performed MTT assay and transwell invasion assay to determine the proliferation and invasion ability of two NPC cell lines, namely, HONE1 and CNE2 cells, respectively. The verification of targets of miR-124 was performed using prediction softwares and luciferase reporter analysis.Results: According to our results, the expression of Capn4 was found to be elevated, whereas the expression of miR-124 was lowered in NPC cell lines compared with normal nasopharyngeal cells. When we preformed overexpression of miR-124, it suppressed the proliferation and invasion of NPC cells. Moreover, miR-124 suppressed the expression of Capn4 by targeting Capn4 in HONE1 and CNE2 cells. When we preformed overexpression of Capn4, it reversed the inhibitory effect of miR-124 on the proliferation and invasion of NPC cells. Furthermore, miR-124–Capn4 axis decreased the levels of β-catenin, cyclin D1, and c-Myc, the components of the Wnt/β-catenin signaling pathway.Conclusion: The suppression of proliferation and invasion of NPC cells by miR-124 were achieved by the regulation of Wnt/β-catenin signaling pathway by targeting Capn4. The results of

  3. The signal transducers Stat1 and Stat3 and their novel target Jmjd3 drive the expression of inflammatory genes in microglia.

    Science.gov (United States)

    Przanowski, Piotr; Dabrowski, Michal; Ellert-Miklaszewska, Aleksandra; Kloss, Michal; Mieczkowski, Jakub; Kaza, Beata; Ronowicz, Anna; Hu, Feng; Piotrowski, Arkadiusz; Kettenmann, Helmut; Komorowski, Jan; Kaminska, Bozena

    2014-03-01

    Most neurological diseases are associated with chronic inflammation initiated by the activation of microglia, which produce cytotoxic and inflammatory factors. Signal transducers and activators of transcription (STATs) are potent regulators of gene expression but contribution of particular STAT to inflammatory gene expression and STAT-dependent transcriptional networks underlying brain inflammation need to be identified. In the present study, we investigated the genomic distribution of Stat binding sites and the role of Stats in the gene expression in lipopolysaccharide (LPS)-activated primary microglial cultures. Integration of chromatin immunoprecipitation-promoter microarray data and transcriptome data revealed novel Stat-target genes including Jmjd3, Ccl5, Ezr, Ifih1, Irf7, Uba7, and Pim1. While knockdown of individual Stat had little effect on the expression of tested genes, knockdown of both Stat1 and Stat3 inhibited the expression of Jmjd3 and inflammatory genes. Transcriptional regulation of Jmjd3 by Stat1 and Stat3 is a novel mechanism crucial for launching inflammatory responses in microglia. The effects of Jmjd3 on inflammatory gene expression were independent of its H3K27me3 demethylase activity. Forced expression of constitutively activated Stat1 and Stat3 induced the expression of Jmjd3, inflammation-related genes, and the production of pro-inflammatory cytokines as potently as lipopolysacharide. Gene set enrichment and gene function analysis revealed categories linked to the inflammatory response in LPS and Stat1C + Stat3C groups. We defined upstream pathways that activate STATs in response to LPS and demonstrated contribution of Tlr4 and Il-6 and interferon-γ signaling. Our findings define novel direct transcriptional targets of Stat1 and Stat3 and highlight their contribution to inflammatory gene expression. Combined analysis of genomic Stat occupancy and transcriptome revealed novel Stat target genes in LPS-induced microglia. Jmjd3 transcription

  4. Cellular MicroRNA Let-7a Suppresses KSHV Replication through Targeting MAP4K4 Signaling Pathways.

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    Xiaohua Tan

    Full Text Available Kaposi's sarcoma (KS-associated herpesvirus (KSHV is the etiologic agent of KS, the most common AIDS-related malignancy. The majority of KS tumor cells harbor latent KSHV virus but only a small percentage undergoes spontaneous lytic replication. Viral reactivation from latency is crucial for the pathogenesis and development of KS, but the cellular mechanisms underlying the switch between viral latency and replication are not well understood.The level of let-7 miRNAs and MAP4K4 in KSHV infected 293T cells were quantified by real-time PCRs. Let-7 expression was silenced by the miRNA sponge technique. In let-7a transfected 293T cells, the expression of MAP4K4 was measured by real-time PCR and western blot. Luciferease expression was employed to examine the effect of let-7a on the 3'-untranslated region (UTR of the MAP4K4 gene in 293T cells. Real-time PCR was used to quantify the KSHV copy numbers in BC-3 cells in which the expression of let-7a and/or MAP4K4 were altered. Finally, ERK, JNK and p38 protein production and their phosphorylation status were detected by western blots in let-7a or MAP4K4 transfected BCBL-1 cells.The expression of microRNA let-7 was dramatically decreased in KSHV infected 293T cells, but that of MAP4K4 was increased significantly. Let-7a is physically associated with and targets the MAP4K4 3'UTR, and inhibits MAP4K4 expression at both mRNA and protein levels. MAP4K4 stimulates KSHV reactivation from latency, whereas let-7a inhibits the function of MAP4K4 by reversing the function of MAP4K4 on JNK, phospho-JNK and phospho-ERK1/2 levels.Our results establish that let-7a specifically suppresses MAP4K4 expression, and further inhibits KSHV reactivation by interfering with the function of MAP4K4 on the MAPK pathway, highlighting let-7a as a potential treatment for KS.

  5. The E3 ubiquitin ligase Triad3A negatively regulates the RIG-I/MAVS signaling pathway by targeting TRAF3 for degradation.

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    Peyman Nakhaei

    2009-11-01

    Full Text Available The primary role of the innate immune response is to limit the spread of infectious pathogens, with activation of Toll-like receptor (TLR and RIG-like receptor (RLR pathways resulting in a pro-inflammatory response required to combat infection. Limiting the activation of these signaling pathways is likewise essential to prevent tissue injury in the host. Triad3A is an E3 ubiquitin ligase that interacts with several components of TLR signaling and modulates TLR activity. In the present study, we demonstrate that Triad3A negatively regulates the RIG-I RNA sensing pathway through Lys48-linked, ubiquitin-mediated degradation of the tumor necrosis factor receptor-associated factor 3 (TRAF3 adapter. Triad3A was induced following dsRNA exposure or virus infection and decreased TRAF3 levels in a dose-dependent manner; moreover, Triad3A expression blocked IRF-3 activation by Ser-396 phosphorylation and inhibited the expression of type 1 interferon and antiviral genes. Lys48-linked ubiquitination of TRAF3 by Triad3A increased TRAF3 turnover, whereas reduction of Triad3A expression by stable shRNA expression correlated with an increase in TRAF3 protein expression and enhancement of the antiviral response following VSV or Sendai virus infection. Triad3A and TRAF3 physically interacted together, and TRAF3 residues Y440 and Q442--previously shown to be important for association with the MAVS adapter--were also critical for Triad3A. Point mutation of the TRAF-Interacting-Motif (TIM of Triad3A abrogated its ability to interact with TRAF3 and modulate RIG-I signaling. TRAF3 appears to undergo sequential ubiquitin "immuno-editing" following virus infection that is crucial for regulation of RIG-I-dependent signaling to the antiviral response. Thus, Triad3A represents a versatile E3 ubiquitin ligase that negatively regulates RIG-like receptor signaling by targeting TRAF3 for degradation following RNA virus infection.

  6. Targeting beta-catenin signaling to induce apoptosis in human breast cancer cells by z-guggulsterone and Gugulipid extract of Ayurvedic medicine plant Commiphora mukul.

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    Jiang, Guoqin; Xiao, Xiao; Zeng, Yan; Nagabhushanam, Kalyanam; Majeed, Muhammed; Xiao, Dong

    2013-08-03

    z-Guggulsterone (z-Gug) and Gugulipid (GL) have been used to treat a variety of ailments. We now report their anti-cancer effect and mechanism against human breast cancer. Using the human estrogen receptor-positive (MCF-7) and triple-negative (MDA-MB-231) breast cancer cells as well as the normal human mammary epithelial cell line (HMEC), we evaluated the anti-breast-cancer efficacy and apoptosis inducing activity of GL. We determined the cellular and molecular mechanism of GL-inhibited breast cancer cell growth. GL significantly inhibited growth of MCF-7 and MDA-MB-231 cells with an IC50~2 μM at pharmacologically relevant concentrations standardized to its major active constituent z-Gug. The GL-induced growth inhibition correlated with apoptosis induction as evidenced by an increase in cytoplasmic histone-associated DNA fragmentation and caspase 3 activity. The GL-induced apoptosis was associated with down-regulation of the β-Catenin signaling pathway. The decreased expression of Wnt/β-Catenin targeting genes, such as cyclin D1, C-myc and survivin, and the inhibition of the activity of the transcription factor (T-cell factor 4, TCF-4) were observed in GL-treated breast cancer cells. The GL treatment resulted in a significant reduction of β-Catenin /TCF-4 complex in both of the cancer cells. The GL-induced apoptotic cell death was significantly enhanced by RNA Interference of β-Catenin and TCF-4. On the other hand, the normal human mammary epithelial cell HMEC, compared with the human breast cancer cells, is significantly more resistant to growth inhibition and apoptosis induction by GL. The present study indicates that the β-Catenin signaling pathway is the target for GL-induced growth inhibition and apoptosis in human breast cancer.

  7. MiR-26a and miR-26b mediate osteoarthritis progression by targeting FUT4 via NF-κB signaling pathway.

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    Hu, Jialei; Wang, Zi; Pan, Yue; Ma, Jia; Miao, Xiaoyan; Qi, Xia; Zhou, Huimin; Jia, Li

    2018-01-01

    Osteoarthritis (OA) is the most common joint disease, characterized by articular cartilage degradation and changes in all other joint tissues. MicroRNAs (miRNAs) play an important role in mediating the main risk factors for OA. This study aimed to investigate the effect of miR-26a/26b on the proliferation and apoptosis of human chondrocytes by targeting fucosyltransferase 4 (FUT4) through NF-κB signaling pathway. We revealed the differential expression profiles of FUT4 and miR-26a/26b in the articular cartilage tissues of OA patients and normal people. The ability of miR-26a/26b to specifically interact with the 3'UTR of FUT4 was demonstrated via a luciferase reporter assay in chondrocytes. Further results showed altered levels of miR-26a/26b and FUT4 could regulate the process of IL-1β-induced extracellular matrix degradation in chondrocytes. Forced miR-26a/26b expression was able to affect chondrocytes proliferation and apoptosis, while altered expression of FUT4 in chondrocytes modulated progression upon transfection with miR-26a/26b mimic or inhibitor. In OA mice, the overexpression of miR-26a/26b by intra-articular injection significantly attenuated OA progression. In addition, regulating FUT4 expression markedly modulated the activity of NF-κB signaling pathway, and this effect could be reversed by miR-26a/26b. In short, miR-26a/-26b/FUT4/NF-κB axis may serve as a predictive biomarker and a potential therapeutic target in OA treatment. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. MicroRNA-199b Modulates Vascular Cell Fate During iPS Cell Differentiation by Targeting the Notch Ligand Jagged1 and Enhancing VEGF Signaling.

    Science.gov (United States)

    Chen, Ting; Margariti, Andriana; Kelaini, Sophia; Cochrane, Amy; Guha, Shaunta T; Hu, Yanhua; Stitt, Alan W; Zhang, Li; Xu, Qingbo

    2015-05-01

    Recent ability to derive endothelial cells (ECs) from induced pluripotent stem (iPS) cells holds a great therapeutic potential for personalized medicine and stem cell therapy. We aimed that better understanding of the complex molecular signals that are evoked during iPS cell differentiation toward ECs may allow specific targeting of their activities to enhance cell differentiation and promote tissue regeneration. In this study, we have generated mouse iPS cells from fibroblasts using established protocol. When iPS cells were cultivated on type IV mouse collagen-coated dishes in differentiation medium, cell differentiation toward vascular lineages were observed. To study the molecular mechanisms of iPS cell differentiation, we found that miR-199b is involved in EC differentiation. A step-wise increase in expression of miR-199 was detected during EC differentiation. Notably, miR-199b targeted the Notch ligand JAG1, resulting in vascular endothelial growth factor (VEGF) transcriptional activation and secretion through the transcription factor STAT3. Upon shRNA-mediated knockdown of the Notch ligand JAG1, the regulatory effect of miR-199b was ablated and there was robust induction of STAT3 and VEGF during EC differentiation. Knockdown of JAG1 also inhibited miR-199b-mediated inhibition of iPS cell differentiation toward smooth muscle markers. Using the in vitro tube formation assay and implanted Matrigel plugs, in vivo, miR-199b also regulated VEGF expression and angiogenesis. This study indicates a novel role for miR-199b as a regulator of the phenotypic switch during vascular cell differentiation derived from iPS cells by regulating critical signaling angiogenic responses. Stem Cells 2015;33:1405-1418. © 2015 AlphaMed Press.

  9. Adrenoceptors promote glucose uptake into adipocytes and muscle by an insulin-independent signaling pathway involving mechanistic target of rapamycin complex 2.

    Science.gov (United States)

    Mukaida, Saori; Evans, Bronwyn A; Bengtsson, Tore; Hutchinson, Dana S; Sato, Masaaki

    2017-02-01

    Uptake of glucose into skeletal muscle and adipose tissue plays a vital role in metabolism and energy balance. Insulin released from β-islet cells of the pancreas promotes glucose uptake in these target tissues by stimulating translocation of GLUT4 transporters to the cell surface. This process is complex, involving signaling proteins including the mechanistic (or mammalian) target of rapamycin (mTOR) and Akt that intersect with multiple pathways controlling cell survival, growth and proliferation. mTOR exists in two forms, mTOR complex 1 (mTORC1), and mTOR complex 2 (mTORC2). mTORC1 has been intensively studied, acting as a key regulator of protein and lipid synthesis that integrates cellular nutrient availability and energy balance. Studies on mTORC2 have focused largely on its capacity to activate Akt by phosphorylation at Ser473, however recent findings demonstrate a novel role for mTORC2 in cellular glucose uptake. For example, agonists acting at β2-adrenoceptors (ARs) in skeletal muscle or β3-ARs in brown adipose tissue increase glucose uptake in vitro and in vivo via mechanisms dependent on mTORC2 but not Akt. In this review, we will focus on the signaling pathways downstream of β-ARs that promote glucose uptake in skeletal muscle and brown adipocytes, and will highlight how the insulin and adrenergic pathways converge and interact in these cells. The identification of insulin-independent mechanisms that promote glucose uptake should facilitate novel treatment strategies for metabolic disease. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Folic acid orchestrates root development linking cell elongation with auxin response and acts independently of the TARGET OF RAPAMYCIN signaling in Arabidopsis thaliana.

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    Ayala-Rodríguez, Juan Ángel; Barrera-Ortiz, Salvador; Ruiz-Herrera, León Francisco; López-Bucio, José

    2017-11-01

    Folic acid is a precursor of tetrahydrofolate (vitamin B9), which is an essential cofactor in most organisms, acting as a carrier for one-carbon units in enzymatic reactions. In this work, we employed pharmacological, genetic and confocal imaging strategies to unravel the signaling mechanism by which folic acid modulates root growth and development. Folic acid supplementation inhibits primary root elongation and induces lateral root formation in a concentration-dependent manner. An analysis of the expression of cell cycle genes pCycD6;1:GFP and CycB1:uidA, and cell expansion Exp7:uidA showed that folic acid promotes cell division but prevented cell elongation, and this correlated with altered expression of auxin-responsive DR5:GFP gene, and PIN1:PIN1:GFP, PIN3:PIN3:GFP, and PIN7:PIN7:GFP auxin transporters at the columella and vasculature of primary roots, whereas mutants defective in auxin signaling (tir1/afb1/afb2 [receptors], slr1 [repressor] and arf7/arf19 [transcription factors]) were less sensitive to folic acid induced primary root shortening and lateral root proliferation. Comparison of growth of WT and TARGET OF RAPAMYCIN (TOR) antisense lines indicates that folic acid acts by an alternative mechanism to this central regulator. Thus, folic acid modulation of root architecture involves auxin and acts independently of the TOR kinase to influence basic cellular programs. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Programmed Death-1 Inhibition of Phosphatidylinositol 3-Kinase/AKT/Mechanistic Target of Rapamycin Signaling Impairs Sarcoidosis CD4+ T Cell Proliferation.

    Science.gov (United States)

    Celada, Lindsay J; Rotsinger, Joseph E; Young, Anjuli; Shaginurova, Guzel; Shelton, Debresha; Hawkins, Charlene; Drake, Wonder P

    2017-01-01

    Patients with progressive sarcoidosis exhibit increased expression of programmed death-1 (PD-1) receptor on their CD4+ T cells. Up-regulation of this marker of T cell exhaustion is associated with a reduction in the proliferative response to T cell receptor (TCR) stimulation, a defect that is reversed by PD-1 pathway blockade. Genome-wide association studies and microarray analyses have correlated signaling downstream from the TCR with sarcoidosis disease severity, but the mechanism is not yet known. Reduced phosphatidylinositol 3-kinase (PI3K)/AKT expression inhibits proliferation by inhibiting cell cycle progression. To test the hypothesis that PD-1 expression attenuates TCR-dependent activation of PI3K/AKT activity in progressive systemic sarcoidosis, we analyzed PI3K/AKT/mechanistic target of rapamycin (mTOR) expression at baseline and after PD-1 pathway blockade in CD4+ T cells isolated from patients with sarcoidosis and healthy control subjects. We confirmed an increased percentage of PD-1+ CD4+ T cells and reduced proliferative capacity in patients with sarcoidosis compared with healthy control subjects (P sarcoidosis CD4+ T cell proliferative capacity is secondary to altered expression of key mediators of cell cycle progression, including the PI3K/AKT/mTOR pathway, via PD-1 up-regulation. This supports the concept that PD-1 up-regulation drives the immunologic deficits associated with sarcoidosis severity by inducing signaling aberrancies in key mediators of cell cycle progression.

  12. Silibinin Treatment Inhibits the Growth of Hedgehog Inhibitor-Resistant Basal Cell Carcinoma Cells via Targeting EGFR-MAPK-Akt and Hedgehog Signaling.

    Science.gov (United States)

    Dheeraj, Arpit; Rigby, Cynthia M; O'Bryant, Cindy L; Agarwal, Chapla; Singh, Rana P; Deep, Gagan; Agarwal, Rajesh

    2017-07-01

    Basal cell carcinoma (BCC) is the most common skin malignancy. Deregulated hedgehog signaling plays a central role in BCC development; therefore, hedgehog inhibitors have been approved to treat locally advanced or metastatic BCC. However, the development of resistance to hedgehog inhibitors is the major challenge in effective treatment of this disease. Herein, we evaluated the efficacy of a natural agent silibinin to overcome resistance with hedgehog inhibitors (Sant-1 and GDC-0449) in BCC cells. Silibinin (25-100 μm) treatment for 48 h strongly inhibited growth and induced death in ASZ001, Sant-1-resistant (ASZ001-Sant-1) and GDC-0449-resistant (ASZ001-GDC-0449) BCC cells. Furthermore, colony-forming ability of ASZ001, ASZ001-Sant-1 and ASZ001-GDC-0449 cells was completely inhibited by silibinin treatment. Molecular analysis showed that silibinin treatment decreased the level of phosphorylated EGFR (Tyrosine 1173) and total EGFR in ASZ001-Sant-1 cells, key signaling molecules responsible for BCC resistance toward hedgehog inhibitors. Further, silibinin treatment decreased the phosphorylated Akt (Serine 473), phosphorylated ERK1/2 (Threonine 202/Tyrosine 204), cyclin D1 and Gli-1 level but increased the SUFU expression in ASZ001-Sant-1-resistant cells. Silibinin treatment of ASZ001-Sant-1-resistant cells also decreased bcl-2 but increased cleaved caspase 3 and PARP cleavage, suggesting induction of apoptosis. Together, these results support silibinin use to target hedgehog inhibitor-resistant BCC cells. © 2017 The American Society of Photobiology.

  13. PRAME-induced inhibition of retinoic acid receptor signaling-mediated differentiation--a possible target for ATRA response in AML without t(15;17).

    Science.gov (United States)

    Bullinger, Lars; Schlenk, Richard F; Götz, Marlies; Botzenhardt, Ursula; Hofmann, Susanne; Russ, Annika C; Babiak, Anna; Zhang, Lu; Schneider, Vanessa; Döhner, Konstanze; Schmitt, Michael; Döhner, Hartmut; Greiner, Jochen

    2013-05-01

    In acute myeloid leukemia (AML) without retinoic acid receptor (RAR) rearrangement, the effect of all-trans-retinoic acid (ATRA) is still poorly understood despite an association of NPM1 mutation and ATRA response. Recently, preferentially expressed antigen in melanoma (PRAME) has been shown to be a dominant repressor of RAR signaling. Thus, we further investigated ATRA response mechanisms, especially the impact of PRAME expression on ATRA responsiveness. We profiled gene expression in diagnostic samples derived from our AML HD98B trial, in which ATRA was administered in addition to intensive chemotherapy. Our data revealed a PRAME expression-associated gene pattern to be significantly enriched for genes involved in the retinoic acid metabolic process. In leukemia cell line models, we could show that retinoic acid-regulated cell proliferation and differentiation are impacted by PRAME expression. In patients with primary AML, repressor activity of high-PRAME levels might be overcome by the addition of ATRA as indicated by better outcome in 2 independent studies (P = 0.029). PRAME seems to impair differentiation and to increase proliferation likely via blocking RAR signaling, which might be reversed by ATRA. PRAME therefore represents a promising target for both ATRA treatment and possibly future immunotherapeutic approaches in AML. ©2013 AACR.

  14. Ethylene signalling and ethylene-targeted transcription factors are required to balance beneficial and nonbeneficial traits in the symbiosis between the endophytic fungus Piriformospora indica and Arabidopsis thaliana.

    Science.gov (United States)

    Camehl, Iris; Sherameti, Irena; Venus, Yvonne; Bethke, Gerit; Varma, Ajit; Lee, Justin; Oelmüller, Ralf

    2010-03-01

    *The endophytic fungus Piriformospora indica colonizes the roots of the model plant Arabidopsis thaliana and promotes its growth and seed production. The fungus can be cultivated in axenic culture without a host, and therefore this is an excellent system to investigate plant-fungus symbiosis. *The growth of etr1, ein2 and ein3/eil1 mutant plants was not promoted or even inhibited by the fungus; the plants produced less seeds and the roots were more colonized compared with the wild-type. This correlates with a mild activation of defence responses. The overexpression of ETHYLENE RESPONSE FACTOR1 constitutively activated defence responses, strongly reduced root colonization and abolished the benefits for the plants. *Piriformospora indica-mediated stimulation of growth and seed yield was not affected by jasmonic acid, and jasmonic acid-responsive promoter beta-glucuronidase gene constructs did not respond to the fungus in Arabidopsis roots. *We propose that ethylene signalling components and ethylene-targeted transcription factors are required to balance beneficial and nonbeneficial traits in the symbiosis. The results show that the restriction of fungal growth by ethylene signalling components is required for the beneficial interaction between the two symbionts.

  15. miR-15a/16 Enhances Radiation Sensitivity of Non-Small Cell Lung Cancer Cells by Targeting the TLR1/NF-κB Signaling Pathway

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    Lan, Fengming [Radiation Oncology Department, PLA Airforce General Hospital, Beijing (China); Radiation Oncology Department, Tianjin Hospital, Tianjin (China); Yue, Xiao [Tianjin Huanhu Hospital, Tianjin Neurosurgery Institute, Tianjin (China); Ren, Gang; Li, Hongqi; Ping, Li; Wang, Yingjie [Radiation Oncology Department, PLA Airforce General Hospital, Beijing (China); Xia, Tingyi, E-mail: xiatingyi1959@163.com [Radiation Oncology Department, PLA Airforce General Hospital, Beijing (China)

    2015-01-01

    Purpose: Many miRNAs have been identified as essential issues and core determining factors in tumor radiation. Recent reports have demonstrated that miRNAs and Toll-like receptors could exert reciprocal effects to control cancer development in various ways. However, a novel role of miR-15a/16 in enhancing radiation sensitivity by directly targeting TLR1 has not been reported, to our knowledge. Methods and Materials: Bioinformatic analyses, luciferase reporter assay, biochemical assays, and subcutaneous tumor establishment were used to characterize the signaling pathways of miRNA-15a/16 in response to radiation treatment. Results: First, an inverse correlation between the expression of miR-15a/16 and TLR1 protein was revealed in non-small cell lung cancer (NSCLC) and normal lung tissues. Next, we corroborated that miR-15a/16 specifically bound to TLR1 3′UTR and inhibited the expression of TLR1 in H358 and A549 cells. Furthermore, miR-15a/16 downregulated the activity of the NF-κB signaling pathway through TLR1. In addition, overexpression of miR-15a/16 inhibited survival capability and increased radiation-induced apoptosis, resulting in enhancement of radiosensitivity in H358 and A549 cells. Finally, subcutaneous tumor bearing NSCLC cells in a nude mice model was established, and the results showed that combined groups (miR-15a/16 + radiation) inhibited tumor growth more significantly than did radiation alone. Conclusions: We mainly elucidate that miRNA-15a/16 can enhance radiation sensitivity by regulating the TLR1/NF-κB signaling pathway and act as a potential therapeutic approach to overcome radioresistance for lung cancer treatment.

  16. Ras oncogene-independent activation of RALB signaling is a targetable mechanism of escape from NRAS(V12) oncogene addiction in acute myeloid leukemia.

    Science.gov (United States)

    Pomeroy, E J; Lee, L A; Lee, R D W; Schirm, D K; Temiz, N A; Ma, J; Gruber, T A; Diaz-Flores, E; Moriarity, B S; Downing, J R; Shannon, K M; Largaespada, D A; Eckfeldt, C E

    2017-06-08

    Somatic mutations that lead to constitutive activation of NRAS and KRAS proto-oncogenes are among the most common in human cancer and frequently occur in acute myeloid leukemia (AML). An inducible NRAS(V12)-driven AML mouse model has established a critical role for continued NRAS(V12) expression in leukemia maintenance. In this model genetic suppression of NRAS(V12) expression results in rapid leukemia remission, but some mice undergo spontaneous relapse with NRAS(V12)-independent (NRI) AMLs providing an opportunity to identify mechanisms that bypass the requirement for Ras oncogene activity and drive leukemia relapse. We found that relapsed NRI AMLs are devoid of NRAS(V12) expression and signaling through the major oncogenic Ras effector pathways, phosphatidylinositol-3-kinase and mitogen-activated protein kinase, but express higher levels of an alternate Ras effector, Ralb, and exhibit NRI phosphorylation of the RALB effector TBK1, implicating RALB signaling in AML relapse. Functional studies confirmed that inhibiting CDK5-mediated RALB activation with a clinically relevant experimental drug, dinaciclib, led to potent RALB-dependent antileukemic effects in human AML cell lines, induced apoptosis in patient-derived AML samples in vitro and led to a 2-log reduction in the leukemic burden in patient-derived xenograft mice. Furthermore, dinaciclib potently suppressed the clonogenic potential of relapsed NRI AMLs in vitro and prevented the development of relapsed AML in vivo. Our findings demonstrate that Ras oncogene-independent activation of RALB signaling is a therapeutically targetable mechanism of escape from NRAS oncogene addiction in AML.

  17. Effects of limiting energy availability via diet and physical activity on mammalian target of rapamycin-related signaling in rat mammary carcinomas.

    Science.gov (United States)

    Jiang, Weiqin; Zhu, Zongjian; Thompson, Henry J

    2013-02-01

    This study evaluated how different approaches to limiting energy availability (LEA) by 15% affected mammalian target of rapamycin (mTOR)-related signaling in mammary carcinomas. Female Sprague Dawley rats, injected with 50mg 1-methyl-1-nitrosourea per kilogram body weight, were randomized to a control or three LEA interventions: (i) sedentary and restricted rats fed to 85% of energy available to the control or motorized wheel running (37 m/min) for an average of (ii) 1621 ± 55 (WRL) or (iii) 3094 ± 126 (WRH) meters/day with food intake adjusted to provide the same net amount of available energy across LEA interventions. Under these conditions, LEA reduced overall cancer burden by 28% (P = 0.04) and down-regulated mTOR-related signaling (Hotelling multivariate, P = 0.002). Among the regulatory nodes assessed, reduced levels of activated protein kinase B (pAkt) and induction of sirtuin 1 (SIRT1) were the most influential factors in distinguishing between sham control and LEA carcinomas. P-Akt was predictive of observed changes in levels of proteins involved in cell cycle control (r = 0.698, P < 0.0001) and induction of apoptosis (r = -0.429, P = 0.014). Plasma insulin and leptin were strongly associated with carcinoma pAkt levels. Consistent with downregulation of mTOR-related signaling by LEA, evidence of decreased lipid synthesis in carcinomas was observed (Hotelling multivariate, P < 0.001) and was negatively correlated with SIRT1 induction. Despite large differences between control and LEA, effects on mTOR regulation were insufficient to distinguish among LEA intervention groups. Given the modest effects observed on the LKB1/AMP-activated protein kinase regulatory node, NADH and NADPH rather than ATP may be more limiting for tumor growth when LEA is 15%.

  18. Targeted inhibition of heat shock protein 90 disrupts multiple oncogenic signaling pathways, thus inducing cell cycle arrest and programmed cell death in human urinary bladder cancer cell lines

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    Karkoulis Panagiotis K

    2013-02-01

    Full Text Available Abstract Background Geldanamycin (GA can be considered a relatively new component with a promising mode of action against human malignancies. It specifically targets heat shock protein 90 (Hsp90 and interferes with its function as a molecular chaperone. Methods In this study, we have investigated the effects of geldanamycin on the regulation of Hsp90-dependent oncogenic signaling pathways directly implicated in cell cycle progression, survival and motility of human urinary bladder cancer cells. In order to assess the biological outcome of Hsp90 inhibition on RT4 (grade I and T24 (grade III human urinary bladder cancer cell lines, we applied MTT assay, FACS analysis, Western blotting, semi-quantitative (sq RT-PCR, electrophoretic mobility shift assay (EMSA, immunofluorescence and scratch-wound assay. Results We have herein demonstrated that, upon geldanamycin treatment, bladder cancer cells are prominently arrested in the G1 phase of cell cycle and eventually undergo programmed cell death via combined activation of apoptosis and autophagy. Furthermore, geldanamycin administration proved to induce prominent downregulation of several Hsp90 protein clients and downstream effectors, such as membrane receptors (IGF-IR and c-Met, protein kinases (Akt, IKKα, IKKβ and Erk1/2 and transcription factors (FOXOs and NF-κΒ, therefore resulting in the impairment of proliferative -oncogenic- signaling and reduction of cell motility. Conclusions In toto, we have evinced the dose-dependent and cell line-specific actions of geldanamycin on cell cycle progression, survival and motility of human bladder cancer cells, due to downregulation of critical Hsp90 clients and subsequent disruption of signaling -oncogenic- integrity.

  19. Targeted inhibition of heat shock protein 90 disrupts multiple oncogenic signaling pathways, thus inducing cell cycle arrest and programmed cell death in human urinary bladder cancer cell lines

    Science.gov (United States)

    2013-01-01

    Background Geldanamycin (GA) can be considered a relatively new component with a promising mode of action against human malignancies. It specifically targets heat shock protein 90 (Hsp90) and interferes with its function as a molecular chaperone. Methods In this study, we have investigated the effects of geldanamycin on the regulation of Hsp90-dependent oncogenic signaling pathways directly implicated in cell cycle progression, survival and motility of human urinary bladder cancer cells. In order to assess the biological outcome of Hsp90 inhibition on RT4 (grade I) and T24 (grade III) human urinary bladder cancer cell lines, we applied MTT assay, FACS analysis, Western blotting, semi-quantitative (sq) RT-PCR, electrophoretic mobility shift assay (EMSA), immunofluorescence and scratch-wound assay. Results We have herein demonstrated that, upon geldanamycin treatment, bladder cancer cells are prominently arrested in the G1 phase of cell cycle and eventually undergo programmed cell death via combined activation of apoptosis and autophagy. Furthermore, geldanamycin administration proved to induce prominent downregulation of several Hsp90 protein clients and downstream effectors, such as membrane receptors (IGF-IR and c-Met), protein kinases (Akt, IKKα, IKKβ and Erk1/2) and transcription factors (FOXOs and NF-κΒ), therefore resulting in the impairment of proliferative -oncogenic- signaling and reduction of cell motility. Conclusions In toto, we have evinced the dose-dependent and cell line-specific actions of geldanamycin on cell cycle progression, survival and motility of human bladder cancer cells, due to downregulation of critical Hsp90 clients and subsequent disruption of signaling -oncogenic- integrity. PMID:23394616

  20. Quercetin inhibits angiogenesis mediated human prostate tumor growth by targeting VEGFR- 2 regulated AKT/mTOR/P70S6K signaling pathways.

    Directory of Open Access Journals (Sweden)

    Poyil Pratheeshkumar

    Full Text Available Angiogenesis is a crucial step in the growth and metastasis of cancers, since it enables the growing tumor to receive oxygen and nutrients. Cancer prevention using natural products has become an integral part of cancer control. We studied the antiangiogenic activity of quercetin using ex vivo, in vivo and in vitro models. Rat aortic ring assay showed that quercetin at non-toxic concentrations significantly inhibited microvessel sprouting and exhibited a significant inhibition in the proliferation, migration, invasion and tube formation of endothelial cells, which are key events in the process of angiogenesis. Most importantly, quercetin treatment inhibited ex vivo angiogenesis as revealed by chicken egg chorioallantoic membrane assay (CAM and matrigel plug assay. Western blot analysis showed that quercetin suppressed VEGF induced phosphorylation of VEGF receptor 2 and their downstream protein kinases AKT, mTOR, and ribosomal protein S6 kinase in HUVECs. Quercetin (20 mg/kg/d significantly reduced the volume and the weight of solid tumors in prostate xenograft mouse model, indicating that quercetin inhibited tumorigenesis by targeting angiogenesis. Furthermore, quercetin reduced the cell viability and induced apoptosis in prostate cancer cells, which were correlated with the downregulation of AKT, mTOR and P70S6K expressions. Collectively the findings in the present study suggest that quercetin inhibits tumor growth and angiogenesis by targeting VEGF-R2 regulated AKT/mTOR/P70S6K signaling pathway, and could be used as a potential drug candidate for cancer therapy.

  1. LC-0882 targets PAK4 and inhibits PAK4-related signaling pathways to suppress the proliferation and invasion of gastric cancer cells.

    Science.gov (United States)

    Zhang, Hong-Yan; Zhang, Jian; Hao, Chen-Zhou; Zhou, Ying; Wang, Jian; Cheng, Mao-Sheng; Zhao, Dong-Mei; Li, Feng

    2017-01-01

    Gastric cancer is the most common malignant tumor and globally the third leading cause of cancer-related deaths. Therefore, there exists an urgent need to identify new effective gastric cancer treatments. Given the important roles in tumorigenesis and progression, p21-activated kinase 4 (PAK4) has been regarded as an attractive high-value druggable target. In this study, we examined the effects and molecular mechanisms of action of the small molecular compound LC-0882 on gastric cancer cells in vitro. LC-0882 was found to significantly inhibit the proliferation of human gastric cancer cells by repressing phospho-PAK4/cyclin D1 and CDK4/6 expression. In addition, LC-0882 was found to attenuate cell invasion by blocking the PAK4/LIMK1/cofilin signaling pathway. Finally, analysis of immunofluorescence revealed that LC-0882 exposure decreased filopodia formation and induced cell elongation in BGC823 and SGC7901 gastric cancer cells. These findings suggest that targeting PAK4 with the novel compound LC-0882 may provide a new chemotherapeutic approach in gastric cancer treatment.

  2. The promotion of breast cancer metastasis caused by inhibition of CSF-1R/CSF-1 signaling is blocked by targeting the G-CSF receptor.

    Science.gov (United States)

    Swierczak, Agnieszka; Cook, Andrew D; Lenzo, Jason C; Restall, Christina M; Doherty, Judy P; Anderson, Robin L; Hamilton, John A

    2014-08-01

    Treatment options are limited for patients with breast cancer presenting with metastatic disease. Targeting of tumor-associated macrophages through the inhibition of colony-stimulating factor-1 receptor (CSF-1R), a key macrophage signaling pathway, has been reported to reduce tumor growth and metastasis, and these treatments are now in clinical trials. Here, we report that, surprisingly, treatment with neutralizing anti-CSF-1R and anti-CSF-1 antibodies, or with two different small-molecule inhibitors of CSF-1R, could actually increase spontaneous metastasis without altering primary tumor growth in mice bearing two independently derived mammary tumors. The blockade of CSF-1R or CSF-1 led to increased levels of serum G-CSF, increased frequency of neutrophils in the primary tumor and in the metastasis-associated lung, as well as increased numbers of neutrophils and Ly6C(hi) monocytes in the peripheral blood. Neutralizing antibody against the G-CSF receptor, which regulates neutrophil development and function, reduced the enhanced metastasis and neutrophil numbers that resulted from CSF-1R blockade. These results indicate that the role of the CSF-1R/CSF-1 system in breast cancer is far more complex than originally proposed, and requires further investigation as a therapeutic target. ©2014 American Association for Cancer Research.

  3. Bcl-2 regulation of the inositol 1,4,5-trisphosphate receptor and calcium signaling in normal and malignant lymphocytes: potential new target for cancer treatment.

    Science.gov (United States)

    Greenberg, Edward F; Lavik, Andrew R; Distelhorst, Clark W

    2014-10-01

    The anti-apoptotic protein Bcl-2 is a versatile regulator of cell survival. Its interactions with its own pro-apoptotic family members are widely recognized for their role in promoting the survival of cancer cells. These interactions are thus being targeted for cancer treatment. Less widely recognized is the interaction of Bcl-2 with the inositol 1,4,5-trisphosphate receptor (InsP3R), an InsP3-gated Ca(2+) channel located on the endoplasmic reticulum. The nature of this interaction, the mechanism by which it controls Ca(2+) release from the ER, its role in T-cell development and survival, and the possibility of targeting it as a novel cancer treatment strategy are summarized in this review. This article is part of a Special Issue entitled: Calcium signaling in health and disease. Guest Editors: Geert Bultynck, Jacques Haiech, Claus W. Heizmann, Joachim Krebs, and Marc Moreau. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Optimizing Molecular-Targeted Therapies in Ovarian Cancer: The Renewed Surge of Interest in Ovarian Cancer Biomarkers and Cell Signaling Pathways

    Directory of Open Access Journals (Sweden)

    Donavon Hiss

    2012-01-01

    Full Text Available The hallmarks of ovarian cancer encompass the development of resistance, disease recurrence and poor prognosis. Ovarian cancer cells express gene signatures which pose significant challenges for cancer drug development, therapeutics, prevention and management. Despite enhancements in contemporary tumor debulking surgery, tentative combination regimens and abdominal radiation which can achieve beneficial response rates, the majority of ovarian cancer patients not only experience adverse effects, but also eventually relapse. Therefore, additional therapeutic possibilities need to be explored to minimize adverse events and prolong progression-free and overall response rates in ovarian cancer patients. Currently, a revival in cancer drug discovery is devoted to identifying diagnostic and prognostic ovarian cancer biomarkers. However, the sensitivity and reliability of such biomarkers may be complicated by mutations in the BRCA1 or BRCA2 genes, diverse genetic risk factors, unidentified initiation and progression elements, molecular tumor heterogeneity and disease staging. There is thus a dire need to expand existing ovarian cancer therapies with broad-spectrum and individualized molecular targeted approaches. The aim of this review is to profile recent developments in our understanding of the interrelationships among selected ovarian tumor biomarkers, heterogeneous expression signatures and related molecular signal transduction pathways, and their translation into more efficacious targeted treatment rationales.

  5. Fisetin inhibits human melanoma cell invasion through promotion of mesenchymal to epithelial transition and by targeting MAPK and NFκB signaling pathways.

    Directory of Open Access Journals (Sweden)

    Harish Chandra Pal

    Full Text Available Malignant melanoma is responsible for approximately 75% of skin cancer-related deaths. BRAF plays an important role in regulating the mitogen-activated protein kinase (MAPK signaling cascade in melanoma with activating mutations in the serine/threonine kinase BRAF occurring in 60-70% of malignant melanomas. The BRAF-MEK-ERK (MAPK pathway is a key regulator of melanoma cell invasion. In addition, activation of NFκB via the MAPK pathway is regulated through MEK-induced activation of IKK. These pathways are potential targets for prevention and treatment of melanoma. In this study, we investigated the effect of fisetin, a phytochemical present in fruits and vegetables, on melanoma cell invasion and epithelial-mesenchymal transition, and delineated the underlying molecular mechanism. Treatment of multiple human malignant melanoma cell lines with fisetin (5-20 µM resulted in inhibition of cell invasion. BRAF mutated melanoma cells were more sensitive to fisetin treatment, and this was associated with a decrease in the phosphorylation of MEK1/2 and ERK1/2. In addition, fisetin inhibited the activation of IKK leading to a reduction in the activation of the NFκB signaling pathway. Treatment of cells with an inhibitor of MEK1/2 (PD98059 or of NFκB (caffeic acid phenethyl ester also reduced melanoma cell invasion. Furthermore, treatment of fisetin promoted mesenchymal to epithelial transition in melanoma cells, which was associated with a decrease in mesenchymal markers (N-cadherin, vimentin, snail and fibronectin and an increase in epithelial markers (E-cadherin and desmoglein. Employing three dimensional skin equivalents consisting of A375 cells admixed with normal human keratinocytes embedded onto a collagen-constricted fibroblast matrix, we found that treatment of fisetin reduced the invasive potential of melanoma cells into the dermis and increased the expression of E-cadherin with a concomitant decrease in vimentin. These results indicate that

  6. Targeting PI3K, mTOR, ERK, and Bcl-2 signaling network shows superior antileukemic activity against AML ex vivo.

    Science.gov (United States)

    Su, Yongwei; Li, Xinyu; Ma, Jun; Zhao, Jianyun; Liu, Shuang; Wang, Guan; Edwards, Holly; Taub, Jeffrey W; Lin, Hai; Ge, Yubin

    2017-12-05

    Acute myeloid leukemia (AML) remains challenging to treat and needs more effective treatments. The PI3K/mTOR pathway is involved in cell survival and has been shown to be constitutively active in 50-80% of AML patients. However, targeting the PI3K/mTOR pathway results in activation of the ERK pathway, which also plays an important role in cell survival. In addition, AML cells often overexpress antiapoptotic Bcl-2 family proteins (e.g., Bcl-2), preventing cell death. Thus, our strategy here is to target the PI3K, mTOR (by VS-5584, a PI3K and mTOR dual inhibitor), ERK (by SCH772984, an ERK-selective inhibitor), and Bcl-2 (by ABT-199, a Bcl-2-selective inhibitor) signaling network to kill AML cells. In this study, we show that while inhibition of PI3K, mTOR, and ERK showed superior induction of cell death compared to inhibition of PI3K and mTOR, the levels of cell death were modest in some AML cell lines and primary patient samples tested. Although simultaneous inhibition of PI3K, mTOR, and ERK caused downregulation of Mcl-1 and upregulation of Bim, immunoprecipitation of Bcl-2 revealed increased binding of Bim to Bcl-2, which was abolished by the addition of ABT-199, suggesting that