Discovery and Biological Evaluation of a Series of Pyrrolo[2,3-b]pyrazines as Novel FGFR Inhibitors

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Yan Zhang

2017-04-01

Full Text Available Abnormality of fibroblast growth factor receptor (FGFR-mediated signaling pathways were frequently found in various human malignancies, making FGFRs hot targets for cancer treatment. To address the consistent need for a new chemotype of FGFR inhibitors, here, we started with a hit structure identified from our internal hepatocyte growth factor receptor (also called c-Met inhibitor project, and conducted a chemical optimization. After exploring three parts of the hit compound, we finally discovered a new series of pyrrolo[2,3-b]pyrazine FGFR inhibitors, which contain a novel scaffold and unique molecular shape. We believe that our findings can help others to further develop selective FGFR inhibitors.

Odontogenic differentiation of human dental pulp cells by calcium silicate materials stimulating via FGFR/ERK signaling pathway

International Nuclear Information System (INIS)

Liu, Chao-Hsin; Hung, Chi-Jr; Huang, Tsui-Hsien; Lin, Chi-Chang; Kao, Chia-Tze; Shie, Ming-You

2014-01-01

Bone healing needs a complex interaction of growth factors that establishes an environment for efficient bone formation. We examine how calcium silicate (CS) and tricalcium phosphate (β-TCP) cements influence the behavior of human dental pulp cells (hDPCs) through fibroblast growth factor receptor (FGFR) and active MAPK pathways, in particular ERK. The hDPCs are cultured with β-TCP and CS, after which the cells' viability and odontogenic differentiation markers are determined by using PrestoBlue® assay and western blot, respectively. The effect of small interfering RNA (siRNA) transfection targeting FGFR was also evaluated. The results showed that CS promoted cell proliferation and enhances FGFR expression. It was also found that CS increases ERK and p38 activity in hDPCs, and furthermore, raises the expression and secretion of DSP, and DMP-1. Additionally, statistically significant differences (p < 0.05) have been found in the calcium deposition in si-FGFR transfection and ERK inhibitor between CS and β-TCP; these variations indicated that ERK/MAPK signaling is involved in the silicon-induced odontogenic differentiation of hDPCs. The current study shows that CS substrates play a key role in odontoblastic differentiation of hDPCs through FGFR and modulate ERK/MAPK activation. - Highlights: • CS influences the behavior of hDPCs through fibroblast growth factor receptor. • CS increases ERK and p38 activity in hDPCs. • ERK/MAPK signaling is involved in the Si-induced odontogenic differentiation of hDPCs. • Ca staining shows that FGFR regulates hDPC differentiation on CS, but not on β-TCP

Fibroblast growth factor receptor 4 (FGFR4) and fibroblast growth factor 19 (FGF19) autocrine enhance breast cancer cells survival.

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Tiong, Kai Hung; Tan, Boon Shing; Choo, Heng Lungh; Chung, Felicia Fei-Lei; Hii, Ling-Wei; Tan, Si Hoey; Khor, Nelson Tze Woei; Wong, Shew Fung; See, Sze-Jia; Tan, Yuen-Fen; Rosli, Rozita; Cheong, Soon-Keng; Leong, Chee-Onn

2016-09-06

Basal-like breast cancer is an aggressive tumor subtype with poor prognosis. The discovery of underlying mechanisms mediating tumor cell survival, and the development of novel agents to target these pathways, is a priority for patients with basal-like breast cancer. From a functional screen to identify key drivers of basal-like breast cancer cell growth, we identified fibroblast growth factor receptor 4 (FGFR4) as a potential mediator of cell survival. We found that FGFR4 mediates cancer cell survival predominantly via activation of PI3K/AKT. Importantly, a subset of basal-like breast cancer cells also secrete fibroblast growth factor 19 (FGF19), a canonical ligand specific for FGFR4. siRNA-mediated silencing of FGF19 or neutralization of extracellular FGF19 by anti-FGF19 antibody (1A6) decreases AKT phosphorylation, suppresses cancer cell growth and enhances doxorubicin sensitivity only in the FGFR4+/FGF19+ breast cancer cells. Consistently, FGFR4/FGF19 co-expression was also observed in 82 out of 287 (28.6%) primary breast tumors, and their expression is strongly associated with AKT phosphorylation, Ki-67 staining, higher tumor stage and basal-like phenotype. In summary, our results demonstrated the presence of an FGFR4/FGF19 autocrine signaling that mediates the survival of a subset of basal-like breast cancer cells and suggest that inactivation of this autocrine loop may potentially serve as a novel therapeutic intervention for future treatment of breast cancers.

Pan-FGFR inhibition leads to blockade of FGF23 signaling, soft tissue mineralization, and cardiovascular dysfunction.

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Yanochko, Gina M; Vitsky, Allison; Heyen, Jonathan R; Hirakawa, Brad; Lam, Justine L; May, Jeff; Nichols, Tim; Sace, Frederick; Trajkovic, Dusko; Blasi, Eileen

2013-10-01

The fibroblast growth factor receptors (FGFR) play a major role in angiogenesis and are desirable targets for the development of therapeutics. Groups of Wistar Han rats were dosed orally once daily for 4 days with a small molecule pan-FGFR inhibitor (5mg/kg) or once daily for 6 days with a small molecule MEK inhibitor (3mg/kg). Serum phosphorous and FGF23 levels increased in all rats during the course of the study. Histologically, rats dosed with either drug exhibited multifocal, multiorgan soft tissue mineralization. Expression levels of the sodium phosphate transporter Npt2a and the vitamin D-metabolizing enzymes Cyp24a1 and Cyp27b1 were modulated in kidneys of animals dosed with the pan-FGFR inhibitor. Both inhibitors decreased ERK phosphorylation in the kidneys and inhibited FGF23-induced ERK phosphorylation in vitro in a dose-dependent manner. A separate cardiovascular outcome study was performed to monitor hemodynamics and cardiac structure and function of telemetered rats dosed with either the pan-FGFR inhibitor or MEK inhibitor for 3 days. Both compounds increased blood pressure (~+ 17 mmHg), decreased heart rate (~-75 bpm), and modulated echocardiography parameters. Our data suggest that inhibition of FGFR signaling following administration of either pan-FGFR inhibitor or MEK inhibitor interferes with the FGF23 pathway, predisposing animals to hyperphosphatemia and a tumoral calcinosis-like syndrome in rodents.

FGFR-1 amplification in metastatic lymph-nodal and haematogenous lobular breast carcinoma

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Brunello Eleonora

2012-12-01

Full Text Available Abstract Background Lobular breast carcinoma usually shows poor responsiveness to chemotherapies and often lacks targeted therapies. Since FGFR1 expression has been shown to play pivotal roles in primary breast cancer tumorigenesis, we sought to analyze the status of FGFR1 gene in a metastatic setting of lobular breast carcinoma, since promising FGFR1 inhibitors has been recently developed. Methods Fifteen tissue metastases from lobular breast carcinomas with matched primary infiltrative lobular breast carcinoma were recruited. Eleven cases showed loco-regional lymph-nodal and four haematogenous metastases. FGFR-1 gene (8p12 amplification was evaluated by chromogenic in situ hybridization (CISH analysis. Her-2/neu and topoisomerase-IIα gene status was assessed. E-cadherin and Hercept Test were also performed. We distinguished amplification (>6 or cluster of signals versus gains (3–6 signals of the locus specific FGFR-1 gene. Results Three (20% primary lobular breast carcinomas showed >6 or cluster of FGFR1 signals (amplification, six cases (40% had a mean of three (range 3–6 chromogenic signals (gains whereas in 6 (40% was not observed any abnormality. Three of 15 metastasis (20% were amplified, 2/15 (13,4% did not. The ten remaining cases (66,6% showed three chromogenic signals. The three cases with FGFR-1 amplification matched with those primary breast carcinomas showing FGFR-1 amplification. The six cases showing FGFR-1 gains in the primary tumour again showed FGFR-1 gains in the metastases. Four cases showed gains of FGFR-1 gene signals in the metastases and not in the primary tumours. Her-2/neu gene amplification was not observed in all cases but one (6% case. Topoisomerase-IIα was not amplified in all cases. Conclusions 1 a subset of metastatic lobular breast carcinoma harbors FGFR-1 gene amplification or gains of chromogenic signals; 2 a minor heterogeneity has been observed after matching primary and metastatic carcinomas; 3 in the

Discovery and Biological Evaluation of a Series of Pyrrolo[2,3-b]pyrazines as Novel FGFR Inhibitors

OpenAIRE

Yan Zhang; Hongchun Liu; Zhen Zhang; Ruifeng Wang; Tongchao Liu; Chaoyun Wang; Yuchi Ma; Jing Ai; Dongmei Zhao; Jingkang Shen; Bing Xiong

2017-01-01

Abnormality of fibroblast growth factor receptor (FGFR)-mediated signaling pathways were frequently found in various human malignancies, making FGFRs hot targets for cancer treatment. To address the consistent need for a new chemotype of FGFR inhibitors, here, we started with a hit structure identified from our internal hepatocyte growth factor receptor (also called c-Met) inhibitor project, and conducted a chemical optimization. After exploring three parts of the hit compound, we finally dis...

Crystal Structure of the FGFR4/LY2874455 Complex Reveals Insights into the Pan-FGFR Selectivity of LY2874455.

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Wu, Daichao; Guo, Ming; Philips, Michael A; Qu, Lingzhi; Jiang, Longying; Li, Jun; Chen, Xiaojuan; Chen, Zhuchu; Chen, Lin; Chen, Yongheng

2016-01-01

Aberrant FGFR4 signaling has been documented abundantly in various human cancers. The majority of FGFR inhibitors display significantly reduced potency toward FGFR4 compared to FGFR1-3. However, LY2874455 has similar inhibition potency for FGFR1-4 with IC50 less than 6.4 nM. To date, there is no published crystal structure of LY2874455 in complex with any kinase. To better understand the pan-FGFR selectivity of LY2874455, we have determined the crystal structure of the FGFR4 kinase domain bound to LY2874455 at a resolution of 2.35 Å. LY2874455, a type I inhibitor for FGFR4, binds to the ATP-binding pocket of FGFR4 in a DFG-in active conformation with three hydrogen bonds and a number of van der Waals contacts. After alignment of the kinase domain sequence of 4 FGFRs, and superposition of the ATP binding pocket of 4 FGFRs, our structural analyses reveal that the interactions of LY2874455 to FGFR4 are largely conserved in 4 FGFRs, explaining at least partly, the broad inhibitory activity of LY2874455 toward 4 FGFRs. Consequently, our studies reveal new insights into the pan-FGFR selectivity of LY2874455 and provide a structural basis for developing novel FGFR inhibitors that target FGFR1-4 broadly.

Common developmental genome deprogramming in schizophrenia - Role of Integrative Nuclear FGFR1 Signaling (INFS).

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Narla, S T; Lee, Y-W; Benson, C A; Sarder, P; Brennand, K J; Stachowiak, E K; Stachowiak, M K

2017-07-01

The watershed-hypothesis of schizophrenia asserts that over 200 different mutations dysregulate distinct pathways that converge on an unspecified common mechanism(s) that controls disease ontogeny. Consistent with this hypothesis, our RNA-sequencing of neuron committed cells (NCCs) differentiated from established iPSCs of 4 schizophrenia patients and 4 control subjects uncovered a dysregulated transcriptome of 1349 mRNAs common to all patients. Data reveals a global dysregulation of developmental genome, deconstruction of coordinated mRNA networks, and the formation of aberrant, new coordinated mRNA networks indicating a concerted action of the responsible factor(s). Sequencing of miRNA transcriptomes demonstrated an overexpression of 16 miRNAs and deconstruction of interactive miRNA-mRNA networks in schizophrenia NCCs. ChiPseq revealed that the nuclear (n) form of FGFR1, a pan-ontogenic regulator, is overexpressed in schizophrenia NCCs and overtargets dysregulated mRNA and miRNA genes. The nFGFR1 targeted 54% of all human gene promoters and 84.4% of schizophrenia dysregulated genes. The upregulated genes reside within major developmental pathways that control neurogenesis and neuron formation, whereas downregulated genes are involved in oligodendrogenesis. Our results indicate (i) an early (preneuronal) genomic etiology of schizophrenia, (ii) dysregulated genes and new coordinated gene networks are common to unrelated cases of schizophrenia, (iii) gene dysregulations are accompanied by increased nFGFR1-genome interactions, and (iv) modeling of increased nFGFR1 by an overexpression of a nFGFR1 lead to up or downregulation of selected genes as observed in schizophrenia NCCs. Together our results designate nFGFR1 signaling as a potential common dysregulated mechanism in investigated patients and potential therapeutic target in schizophrenia. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Polysaccharides purified from wild Cordyceps activate FGF2/FGFR1c signaling

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Zeng, Yangyang; Han, Zhangrun; Yu, Guangli; Hao, Jiejie; Zhang, Lijuan

2015-02-01

Land animals as well as all organisms in ocean synthesize sulfated polysaccharides. Fungi split from animals about 1.5 billion years ago. As fungi make the evolutionary journey from ocean to land, the biggest changes in their living environment may be a sharp decrease in salt concentration. It is established that sulfated polysaccharides interact with hundreds of signaling molecules and facilitate many signaling transduction pathways, including fibroblast growth factor (FGF) and FGF receptor signaling pathway. The disappearance of sulfated polysaccharides in fungi and plants on land might indicate that polysaccharides without sulfation might be sufficient in facilitating protein ligand/receptor interactions in low salinity land. Recently, it was reported that plants on land start to synthesize sulfated polysaccharides in high salt environment, suggesting that fungi might be able to do the same when exposed in such environment. Interestingly, Cordyceps, a fungus habituating inside caterpillar body, is the most valued traditional Chinese Medicine. One of the important pharmaceutical active ingredients in Cordyceps is polysaccharides. Therefore, we hypothesize that the salty environment inside caterpillar body might allow the fungi to synthesize sulfated polysaccharides. To test the hypothesis, we isolated polysaccharides from both lava and sporophore of wild Cordyceps and also from Cordyceps militaris cultured without or with added salts. We then measured the polysaccharide activity using a FGF2/FGFR1c signaling-dependent BaF3 cell proliferation assay and found that polysaccharides isolated from wild Cordyceps activated FGF2/FGFR signaling, indicating that the polysaccharides synthesized by wild Cordyceps are indeed different from those by the cultured mycelium.

Suramin blocks interaction between human FGF1 and FGFR2 D2 domain and reduces downstream signaling activity

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Wu, Zong-Sian, E-mail: gary810426@hotmail.com [Department of Chemistry, National Tsing Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China); Liu, Che Fu, E-mail: s9823002@m98.nthu.edu.tw [Department of Chemistry, National Tsing Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China); Fu, Brian, E-mail: brianfu9@gmail.com [Northwood High School, Irvine, CA (United States); Chou, Ruey-Hwang, E-mail: rhchou@mail.cmu.edu.tw [Graduate Institute of Cancer Biology and Center for Molecular Medicine, China Medical University, No.91, Hsueh-Shih Road, Taichung 40402, Taiwan (China); Department of Biotechnology, Asia University, Taiwan (China); Yu, Chin, E-mail: cyu.nthu@gmail.com [Department of Chemistry, National Tsing Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China)

2016-09-02

The extracellular portion of the human fibroblast growth factor receptor2 D2 domain (FGFR2 D2) interacts with human fibroblast growth factor 1 (hFGF1) to activate a downstream signaling cascade that ultimately affects mitosis and differentiation. Suramin is an antiparasiticdrug and a potent inhibitor of FGF-induced angiogenesis. Suramin has been shown to bind to hFGF1, and might block the interaction between hFGF1 and FGFR2 D2. Here, we titrated hFGF1 with FGFR2 D2 and suramin to elucidate their interactions using the detection of NMR. The docking results of both hFGF1-FGFR2 D2 domain and hFGF1-suramin complex were superimposed. The results indicate that suramin blocks the interaction between hFGF1 and FGFR2 D2. We used the PyMOL software to show the hydrophobic interaction of hFGF1-suramin. In addition, we used a Water-soluble Tetrazolium salts assay (WST1) to assess hFGF1 bioactivity. The results will be useful for the development of new antimitogenic activity drugs. - Highlights: • The interfacial residues on hFGF1-FGFR2 D2 and hFGF1-Suramin contact surface were mapped by {sup 1}H-{sup 15}N HSQC experiments. • hFGF1-FGFR2 D2 and hFGF1-Suramin complex models were generated from NMR restraints by using HADDOCK program. • We analyzed hFGF1-Suramin complex models and found the interaction between hFGF1-Suramin is hydrophobic. • The bioactivity of the hFGF1-FGFR2 D2 and hFGF1-Suramin complex was studied by using WST1 assay.

Suramin blocks interaction between human FGF1 and FGFR2 D2 domain and reduces downstream signaling activity

International Nuclear Information System (INIS)

Wu, Zong-Sian; Liu, Che Fu; Fu, Brian; Chou, Ruey-Hwang; Yu, Chin

2016-01-01

The extracellular portion of the human fibroblast growth factor receptor2 D2 domain (FGFR2 D2) interacts with human fibroblast growth factor 1 (hFGF1) to activate a downstream signaling cascade that ultimately affects mitosis and differentiation. Suramin is an antiparasiticdrug and a potent inhibitor of FGF-induced angiogenesis. Suramin has been shown to bind to hFGF1, and might block the interaction between hFGF1 and FGFR2 D2. Here, we titrated hFGF1 with FGFR2 D2 and suramin to elucidate their interactions using the detection of NMR. The docking results of both hFGF1-FGFR2 D2 domain and hFGF1-suramin complex were superimposed. The results indicate that suramin blocks the interaction between hFGF1 and FGFR2 D2. We used the PyMOL software to show the hydrophobic interaction of hFGF1-suramin. In addition, we used a Water-soluble Tetrazolium salts assay (WST1) to assess hFGF1 bioactivity. The results will be useful for the development of new antimitogenic activity drugs. - Highlights: • The interfacial residues on hFGF1-FGFR2 D2 and hFGF1-Suramin contact surface were mapped by "1H-"1"5N HSQC experiments. • hFGF1-FGFR2 D2 and hFGF1-Suramin complex models were generated from NMR restraints by using HADDOCK program. • We analyzed hFGF1-Suramin complex models and found the interaction between hFGF1-Suramin is hydrophobic. • The bioactivity of the hFGF1-FGFR2 D2 and hFGF1-Suramin complex was studied by using WST1 assay.

New insight on FGFR3-related chondrodysplasias molecular physiopathology revealed by human chondrocyte gene expression profiling.

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Laurent Schibler

Full Text Available Endochondral ossification is the process by which the appendicular skeleton, facial bones, vertebrae and medial clavicles are formed and relies on the tight control of chondrocyte maturation. Fibroblast growth factor receptor (FGFR3 plays a role in bone development and maintenance and belongs to a family of proteins which differ in their ligand affinities and tissue distribution. Activating mutations of the FGFR3 gene lead to craniosynostosis and multiple types of skeletal dysplasia with varying degrees of severity: thanatophoric dysplasia (TD, achondroplasia and hypochondroplasia. Despite progress in the characterization of FGFR3-mediated regulation of cartilage development, many aspects remain unclear. The aim and the novelty of our study was to examine whole gene expression differences occurring in primary human chondrocytes isolated from normal cartilage or pathological cartilage from TD-affected fetuses, using Affymetrix technology. The phenotype of the primary cells was confirmed by the high expression of chondrocytic markers. Altered expression of genes associated with many cellular processes was observed, including cell growth and proliferation, cell cycle, cell adhesion, cell motility, metabolic pathways, signal transduction, cell cycle process and cell signaling. Most of the cell cycle process genes were down-regulated and consisted of genes involved in cell cycle progression, DNA biosynthesis, spindle dynamics and cytokinesis. About eight percent of all modulated genes were found to impact extracellular matrix (ECM structure and turnover, especially glycosaminoglycan (GAG and proteoglycan biosynthesis and sulfation. Altogether, the gene expression analyses provide new insight into the consequences of FGFR3 mutations in cell cycle regulation, onset of pre-hypertrophic differentiation and concomitant metabolism changes. Moreover, impaired motility and ECM properties may also provide clues about growth plate disorganization. These

Novel mutation detection of fibroblast growth factor receptor 1 (FGFR1) gene, FGFR2IIIa, FGFR2IIIb, FGFR2IIIc, FGFR3, FGFR4 gene for craniosynostosis: A prospective study in Asian Indian patient.

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Barik, Mayadhar; Bajpai, Minu; Malhotra, Arun; Samantaray, Jyotish Chandra; Dwivedi, Sadananda; Das, Sambhunath

2015-01-01

Craniosynostosis (CS) syndrome is an autosomal dominant condition classically combining craniosynostosis and non-syndromic craniosynostosis with digital anomalies of the hands and feet. The majority of cases are caused by heterozygous mutations in the third immunoglobulin-like domain (IgIII) of FGFR2, whilst a larger number of cases can be attributed to mutations outside this region of the protein. To find out the FGFR1, FGFR2, FGFR3 and FGFR4 gene in craniosynostosis syndrome. A hospital based prospective study. Prospective analysis of clinical records of patients registered in CS clinic from December 2007 to January 2015 was done in patients between 4 months to 13 years of age. We have performed genetic findings in a three generation Indian family with Craniosynostosis syndrome. We report for the first time the clinical and genetic findings in a three generation Indian family with Craniosynostosis syndrome caused by a heterozygous missense mutation, Thr 392 Thr and ser 311 try, located in the IgII domain of FGFR2. FGFR 3 and 4 gene basis syndrome was eponymously named. Genetic analysis demonstrated that 51/56 families to be unrelated. In FGFR3 gene 10/TM location of 1172 the nucleotide changes C>A, Ala 391 Glu 19/56 and Exon-19, 5q35.2 at conserved linker region the changes occurred pro 246 Arg in 25/56 families. Independent genetic origins, but phenotypic similarities in the 51 families add to the evidence supporting the theory of selfish spermatogonial selective advantage for this rare gain-of-function FGFR2 mutation.

miR-24-3p/FGFR3 Signaling as a Novel Axis Is Involved in Epithelial-Mesenchymal Transition and Regulates Lung Adenocarcinoma Progression

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Pengyu Jing

2018-01-01

Full Text Available Our previous studies showed that Fibroblast growth factor receptor 3 (FGFR3 contributed to cell growth in lung cancer. However, the correlation between FGFR3 and tumor progression, coupled with the underlying mechanisms, are not fully understood. The clinical significance of FGFR3 was determined in two cohorts of clinical samples (n=22, n=78. A panel of biochemical assays and functional experiments was utilized to elucidate the underlying mechanisms and effects of FGFR3 and miR-24-3p on lung adenocarcinoma progression. Upregulated FGFR3 expression indicated an adverse prognosis for lung adenocarcinoma individuals and promoted metastatic potential of lung adenocarcinoma cells. Owing to the direct regulation towards FGFR3, miR-24-3p could interfere with the potential of proliferation, migration, and invasion in lung adenocarcinoma, following variations of EMT-related protein expression. As a significant marker of EMT, E-cadherin was negatively correlated with FGFR3, of which ectopic overexpression could neutralize the antitumour effects of miR-24-3p and reverse its regulatory effects on EMT markers. Taken together, these findings define a novel insight into the miR-24-3p/FGFR3 signaling axis in regulating lung adenocarcinoma progression and suggest that targeting the miR-24-3p/FGFR3 axis could be an effective and efficient way to prevent tumor progression.

FGFR4 role in epithelial-mesenchymal transition and its therapeutic value in colorectal cancer.

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Alberto Peláez-García

Full Text Available Fibroblast growth factor receptor 4 (FGFR4 is vital in early development and tissue repair. FGFR4 expression levels are very restricted in adult tissues, except in several solid tumors including colorectal cancer, which showed overexpression of FGFR4. Here, FGFR4 mutation analysis discarded the presence of activating mutations, other than Arg(388, in different colorectal cancer cell lines and tumoral samples. Stable shRNA FGFR4-silencing in SW480 and SW48 cell lines resulted in a significant decrease in cell proliferation, adhesion, cell migration and invasion. This decrease in the tumorigenic and invasive capabilities of colorectal cancer cells was accompanied by a decrease of Snail, Twist and TGFβ gene expression levels and an increase of E-cadherin, causing a reversion to a more epithelial phenotype, in three different cell lines. In addition, FGFR4-signaling activated the oncogenic SRC, ERK1/2 and AKT pathways in colon cancer cells and promoted an increase in cell survival. The relevance of FGFR4 in tumor growth was supported by two different strategies. Kinase inhibitors abrogated FGFR4-related cell growth and signaling pathways at the same extent than FGFR4-silenced cells. Specific FGFR4-targeting using antibodies provoked a similar reduction in cell growth. Moreover, FGFR4 knock-down cells displayed a reduced capacity for in vivo tumor formation and angiogenesis in nude mice. Collectively, our data support a crucial role for FGFR4 in tumorigenesis, invasion and survival in colorectal cancer. In addition, FGFR4 targeting demonstrated its applicability for colorectal cancer therapy.

FGFR4 Role in Epithelial-Mesenchymal Transition and Its Therapeutic Value in Colorectal Cancer

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Torres, Sofía; Hernández-Varas, Pablo; Teixidó, Joaquín; Bonilla, Félix; de Herreros, Antonio Garcia; Casal, J. Ignacio

2013-01-01

Fibroblast growth factor receptor 4 (FGFR4) is vital in early development and tissue repair. FGFR4 expression levels are very restricted in adult tissues, except in several solid tumors including colorectal cancer, which showed overexpression of FGFR4. Here, FGFR4 mutation analysis discarded the presence of activating mutations, other than Arg388, in different colorectal cancer cell lines and tumoral samples. Stable shRNA FGFR4-silencing in SW480 and SW48 cell lines resulted in a significant decrease in cell proliferation, adhesion, cell migration and invasion. This decrease in the tumorigenic and invasive capabilities of colorectal cancer cells was accompanied by a decrease of Snail, Twist and TGFβ gene expression levels and an increase of E-cadherin, causing a reversion to a more epithelial phenotype, in three different cell lines. In addition, FGFR4-signaling activated the oncogenic SRC, ERK1/2 and AKT pathways in colon cancer cells and promoted an increase in cell survival. The relevance of FGFR4 in tumor growth was supported by two different strategies. Kinase inhibitors abrogated FGFR4-related cell growth and signaling pathways at the same extent than FGFR4-silenced cells. Specific FGFR4-targeting using antibodies provoked a similar reduction in cell growth. Moreover, FGFR4 knock-down cells displayed a reduced capacity for in vivo tumor formation and angiogenesis in nude mice. Collectively, our data support a crucial role for FGFR4 in tumorigenesis, invasion and survival in colorectal cancer. In addition, FGFR4 targeting demonstrated its applicability for colorectal cancer therapy. PMID:23696849

Expression of FGFR3 during human testis development and in germ cell-derived tumours of young adults.

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Ewen, Katherine A; Olesen, Inge A; Winge, Sofia B; Nielsen, Ana R; Nielsen, John E; Graem, Niels; Juul, Anders; Rajpert-De Meyts, Ewa

2013-01-01

Observations in patients with an activating mutation of fibroblast growth factor receptor 3 (FGFR3) suggest a role for FGFR3 signalling in promoting proliferation or survival of germ cells. In this study, we aimed to identify the FGFR3 subtype and the ontogeny of expression during human testis development and to ascertain whether FGFR3 signalling is linked to germ cell proliferation and the pathogenesis of testicular germ cell tumours (TGCTs) of young adult men. Using RT-PCR, immunohistochemistry and Western blotting, we examined 58 specimens of human testes throughout development for FGFR3 expression, and then compared expression of FGFR3 with proliferation markers (PCNA or Ki67). We also analysed for FGFR3 expression 30 TGCTs and 28 testes containing the tumour precursor cell, carcinoma in situ (CIS). Fetal and adult testes expressed exclusively the FGFR3IIIc isoform. FGFR3 protein expression was restricted to the cytoplasm/plasma membrane of spermatogonia and was most prevalent at mid-gestation, infancy and from puberty onwards. Phosphorylated (p)FGFR was detected in pre-spermatogonia at mid-gestation and in spermatogonia during puberty and in the adult testis. Throughout normal human testis development, expression of FGFR3 did not directly correlate with proliferation markers. In preinvasive CIS cells and in TGCTs, including classical seminoma and embryonal carcinoma, FGFR3IIIc was detected only in a small number of cells, with a heterogeneous expression pattern. FGFR3 is an excellent marker for human pre-/spermatogonia throughout development. Signalling through this receptor is likely associated with spermatogonial survival rather than proliferation. FGFR3 is not expressed in gonocytes and may not be essential to the aetiology of TGCTs stemming from CIS.

FGFR2 amplification is predictive of sensitivity to regorafenib in gastric and colorectal cancers in vitro.

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Cha, Yongjun; Kim, Hwang-Phill; Lim, Yoojoo; Han, Sae-Won; Song, Sang-Hyun; Kim, Tae-You

2018-03-24

Although regorafenib has demonstrated survival benefits in patients with metastatic colorectal and gastrointestinal stromal tumors, no proven biomarker has been identified for predicting sensitivity to regorafenib. Here, we investigated preclinical activity of regorafenib in gastric and colorectal cancer cells to identify genetic alterations associated with sensitivity to regorafenib. Mutation profiles and copy number assays of regorafenib target molecules indicated that amplification of FGFR2 was the only genetic alteration associated with in vitro sensitivity to regorafenib. Regorafenib effectively inhibited phosphorylation of FGFR2 and its downstream signaling molecules in a dose-dependent manner and selectively in FGFR2 amplified cells. Regorafenib induced G1 arrest (SNU-16, KATO-III) and apoptosis (NCI-H716), however, no significant changes were seen in cell lines without FGFR2 amplification. In SNU-16 mice xenografts, regorafenib significantly inhibited tumor growth, proliferation, and FGFR signaling compared to treatment with control vehicle. Regorafenib effectively abrogates activated FGFR2 signaling in FGFR2 amplified gastric and colorectal cancer and therefore, might be considered for integration into treatment in patients with FGFR2 amplified gastric and colorectal cancers. This article is protected by copyright. All rights reserved. Molecular Oncology (2018) © 2018 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.

Activation of Aurora A kinase through the FGF1/FGFR signaling axis sustains the stem cell characteristics of glioblastoma cells

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Hsu, Yi-Chao [Division of Regenerative Medicine, Institute of Cellular and System Medicine, National Health Research Institutes, Miaoli, Taiwan (China); Institute of Biomedical Sciences, Mackay Medical College, New Taipei City, Taiwan (China); Kao, Chien-Yu [Division of Regenerative Medicine, Institute of Cellular and System Medicine, National Health Research Institutes, Miaoli, Taiwan (China); Graduate Program of Biotechnology in Medicine, Institute of Biotechnology and Department of Life Science, National Tsing Hua University, Hsinchu, Taiwan (China); Chung, Yu-Fen; Lee, Don-Ching [Division of Regenerative Medicine, Institute of Cellular and System Medicine, National Health Research Institutes, Miaoli, Taiwan (China); Liu, Jen-Wei [Division of Regenerative Medicine, Institute of Cellular and System Medicine, National Health Research Institutes, Miaoli, Taiwan (China); Department of Life Sciences, National Chung Hsing University, Taichung, Taiwan (China); Chiu, Ing-Ming, E-mail: ingming@nhri.org.tw [Division of Regenerative Medicine, Institute of Cellular and System Medicine, National Health Research Institutes, Miaoli, Taiwan (China); Graduate Program of Biotechnology in Medicine, Institute of Biotechnology and Department of Life Science, National Tsing Hua University, Hsinchu, Taiwan (China); Department of Life Sciences, National Chung Hsing University, Taichung, Taiwan (China)

2016-06-10

Fibroblast growth factor 1 (FGF1) binds and activates FGF receptors, thereby regulating cell proliferation and neurogenesis. Human FGF1 gene 1B promoter (−540 to +31)-driven SV40 T antigen has been shown to result in tumorigenesis in the brains of transgenic mice. FGF1B promoter (−540 to +31)-driven green fluorescent protein (F1BGFP) has also been used in isolating neural stem cells (NSCs) with self-renewal and multipotency from developing and adult mouse brains. In this study, we provide six lines of evidence to demonstrate that FGF1/FGFR signaling is implicated in the expression of Aurora A (AurA) and the activation of its kinase domain (Thr288 phosphorylation) in the maintenance of glioblastoma (GBM) cells and NSCs. First, treatment of FGF1 increases AurA expression in human GBM cell lines. Second, using fluorescence-activated cell sorting, we observed that F1BGFP reporter facilitates the isolation of F1BGFP(+) GBM cells with higher expression levels of FGFR and AurA. Third, both FGFR inhibitor (SU5402) and AurA inhibitor (VX680) could down-regulate F1BGFP-dependent AurA activity. Fourth, inhibition of AurA activity by two different AurA inhibitors (VX680 and valproic acid) not only reduced neurosphere formation but also induced neuronal differentiation of F1BGFP(+) GBM cells. Fifth, flow cytometric analyses demonstrated that F1BGFP(+) GBM cells possessed different NSC cell surface markers. Finally, inhibition of AurA by VX680 reduced the neurosphere formation of different types of NSCs. Our results show that activation of AurA kinase through FGF1/FGFR signaling axis sustains the stem cell characteristics of GBM cells. Implications: This study identified a novel mechanism for the malignancy of GBM, which could be a potential therapeutic target for GBM. - Highlights: • We report that FGF1 treatment can stimulate AurA kinase expression in human GBM cells. • FGF1/FGFR signaling is involved in the activation of AurA kinase. • FGF1 sustains the self

Activation of Aurora A kinase through the FGF1/FGFR signaling axis sustains the stem cell characteristics of glioblastoma cells

International Nuclear Information System (INIS)

Hsu, Yi-Chao; Kao, Chien-Yu; Chung, Yu-Fen; Lee, Don-Ching; Liu, Jen-Wei; Chiu, Ing-Ming

2016-01-01

Fibroblast growth factor 1 (FGF1) binds and activates FGF receptors, thereby regulating cell proliferation and neurogenesis. Human FGF1 gene 1B promoter (−540 to +31)-driven SV40 T antigen has been shown to result in tumorigenesis in the brains of transgenic mice. FGF1B promoter (−540 to +31)-driven green fluorescent protein (F1BGFP) has also been used in isolating neural stem cells (NSCs) with self-renewal and multipotency from developing and adult mouse brains. In this study, we provide six lines of evidence to demonstrate that FGF1/FGFR signaling is implicated in the expression of Aurora A (AurA) and the activation of its kinase domain (Thr288 phosphorylation) in the maintenance of glioblastoma (GBM) cells and NSCs. First, treatment of FGF1 increases AurA expression in human GBM cell lines. Second, using fluorescence-activated cell sorting, we observed that F1BGFP reporter facilitates the isolation of F1BGFP(+) GBM cells with higher expression levels of FGFR and AurA. Third, both FGFR inhibitor (SU5402) and AurA inhibitor (VX680) could down-regulate F1BGFP-dependent AurA activity. Fourth, inhibition of AurA activity by two different AurA inhibitors (VX680 and valproic acid) not only reduced neurosphere formation but also induced neuronal differentiation of F1BGFP(+) GBM cells. Fifth, flow cytometric analyses demonstrated that F1BGFP(+) GBM cells possessed different NSC cell surface markers. Finally, inhibition of AurA by VX680 reduced the neurosphere formation of different types of NSCs. Our results show that activation of AurA kinase through FGF1/FGFR signaling axis sustains the stem cell characteristics of GBM cells. Implications: This study identified a novel mechanism for the malignancy of GBM, which could be a potential therapeutic target for GBM. - Highlights: • We report that FGF1 treatment can stimulate AurA kinase expression in human GBM cells. • FGF1/FGFR signaling is involved in the activation of AurA kinase. • FGF1 sustains the self

FGFR a promising druggable target in cancer: Molecular biology and new drugs.

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Porta, Rut; Borea, Roberto; Coelho, Andreia; Khan, Shahanavaj; Araújo, António; Reclusa, Pablo; Franchina, Tindara; Van Der Steen, Nele; Van Dam, Peter; Ferri, Jose; Sirera, Rafael; Naing, Aung; Hong, David; Rolfo, Christian

2017-05-01

The Fibroblast Growth Factor Receptor (FGFR) family consists of Tyrosine Kinase Receptors (TKR) involved in several biological functions. Recently, alterations of FGFR have been reported to be important for progression and development of several cancers. In this setting, different studies are trying to evaluate the efficacy of different therapies targeting FGFR. This review summarizes the current status of treatments targeting FGFR, focusing on the trials that are evaluating the FGFR profile as inclusion criteria: Multi-Target, Pan-FGFR Inhibitors and anti-FGF (Fibroblast Growth Factor)/FGFR Monoclonal Antibodies. Most of the TKR share intracellular signaling pathways; therefore, cancer cells tend to overcome the inhibition of one tyrosine kinase receptor by activating another. The future of TKI (Tyrosine Kinase Inhibitor) therapy will potentially come from multi-targeted TKIs that target different TKR simultaneously. It is crucial to understand the interaction of the FGF-FGFR axis with other known driver TKRs. Based on this, it is possible to develop therapeutic strategies targeting multiple connected TKRs at once. One correct step in this direction is the reassessment of multi target inhibitors considering the FGFR status of the tumor. Another opportunity arises from assessing the use of FGFR TKI on patients harboring FGFR alterations. Copyright © 2017 Elsevier B.V. All rights reserved.

Liver-Enriched Gene 1, a Glycosylated Secretory Protein, Binds to FGFR and Mediates an Anti-stress Pathway to Protect Liver Development in Zebrafish.

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Minjie Hu

2016-02-01

Full Text Available Unlike mammals and birds, teleost fish undergo external embryogenesis, and therefore their embryos are constantly challenged by stresses from their living environment. These stresses, when becoming too harsh, will cause arrest of cell proliferation, abnormal cell death or senescence. Such organisms have to evolve a sophisticated anti-stress mechanism to protect the process of embryogenesis/organogenesis. However, very few signaling molecule(s mediating such activity have been identified. liver-enriched gene 1 (leg1 is an uncharacterized gene that encodes a novel secretory protein containing a single domain DUF781 (domain of unknown function 781 that is well conserved in vertebrates. In the zebrafish genome, there are two copies of leg1, namely leg1a and leg1b. leg1a and leg1b are closely linked on chromosome 20 and share high homology, but are differentially expressed. In this report, we generated two leg1a mutant alleles using the TALEN technique, then characterized liver development in the mutants. We show that a leg1a mutant exhibits a stress-dependent small liver phenotype that can be prevented by chemicals blocking the production of reactive oxygen species. Further studies reveal that Leg1a binds to FGFR3 and mediates a novel anti-stress pathway to protect liver development through enhancing Erk activity. More importantly, we show that the binding of Leg1a to FGFR relies on the glycosylation at the 70th asparagine (Asn(70 or N(70, and mutating the Asn(70 to Ala(70 compromised Leg1's function in liver development. Therefore, Leg1 plays a unique role in protecting liver development under different stress conditions by serving as a secreted signaling molecule/modulator.

The Adhesion Molecule KAL-1/anosmin-1 Regulates Neurite Branching through a SAX-7/L1CAM–EGL-15/FGFR Receptor Complex

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Carlos A. Díaz-Balzac

2015-06-01

Full Text Available Neurite branching is essential for correct assembly of neural circuits, yet it remains a poorly understood process. For example, the neural cell adhesion molecule KAL-1/anosmin-1, which is mutated in Kallmann syndrome, regulates neurite branching through mechanisms largely unknown. Here, we show that KAL-1/anosmin-1 mediates neurite branching as an autocrine co-factor with EGL-17/FGF through a receptor complex consisting of the conserved cell adhesion molecule SAX-7/L1CAM and the fibroblast growth factor receptor EGL-15/FGFR. This protein complex, which appears conserved in humans, requires the immunoglobulin (Ig domains of SAX-7/L1CAM and the FN(III domains of KAL-1/anosmin-1 for formation in vitro as well as function in vivo. The kinase domain of the EGL-15/FGFR is required for branching, and genetic evidence suggests that ras-mediated signaling downstream of EGL-15/FGFR is necessary to effect branching. Our studies establish a molecular pathway that regulates neurite branching during development of the nervous system.

Mutational analysis of EGFR and related signaling pathway genes in lung adenocarcinomas identifies a novel somatic kinase domain mutation in FGFR4.

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Jenifer L Marks

2007-05-01

Full Text Available Fifty percent of lung adenocarcinomas harbor somatic mutations in six genes that encode proteins in the EGFR signaling pathway, i.e., EGFR, HER2/ERBB2, HER4/ERBB4, PIK3CA, BRAF, and KRAS. We performed mutational profiling of a large cohort of lung adenocarcinomas to uncover other potential somatic mutations in genes of this signaling pathway that could contribute to lung tumorigenesis.We analyzed genomic DNA from a total of 261 resected, clinically annotated non-small cell lung cancer (NSCLC specimens. The coding sequences of 39 genes were screened for somatic mutations via high-throughput dideoxynucleotide sequencing of PCR-amplified gene products. Mutations were considered to be somatic only if they were found in an independent tumor-derived PCR product but not in matched normal tissue. Sequencing of 9MB of tumor sequence identified 239 putative genetic variants. We further examined 22 variants found in RAS family genes and 135 variants localized to exons encoding the kinase domain of respective proteins. We identified a total of 37 non-synonymous somatic mutations; 36 were found collectively in EGFR, KRAS, BRAF, and PIK3CA. One somatic mutation was a previously unreported mutation in the kinase domain (exon 16 of FGFR4 (Glu681Lys, identified in 1 of 158 tumors. The FGFR4 mutation is analogous to a reported tumor-specific somatic mutation in ERBB2 and is located in the same exon as a previously reported kinase domain mutation in FGFR4 (Pro712Thr in a lung adenocarcinoma cell line.This study is one of the first comprehensive mutational analyses of major genes in a specific signaling pathway in a sizeable cohort of lung adenocarcinomas. Our results suggest the majority of gain-of-function mutations within kinase genes in the EGFR signaling pathway have already been identified. Our findings also implicate FGFR4 in the pathogenesis of a subset of lung adenocarcinomas.

A variant of fibroblast growth factor receptor 2 (Fgfr2 regulates left-right asymmetry in zebrafish.

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Da-Wei Liu

Full Text Available Many organs in vertebrates are left-right asymmetrical located. For example, liver is at the right side and stomach is at the left side in human. Fibroblast growth factor (Fgf signaling is important for left-right asymmetry. To investigate the roles of Fgfr2 signaling in zebrafish left-right asymmetry, we used splicing blocking morpholinos to specifically block the splicing of fgfr2b and fgfr2c variants, respectively. We found that the relative position of the liver and the pancreas were disrupted in fgfr2c morphants. Furthermore, the left-right asymmetry of the heart became random. Expression pattern of the laterality controlling genes, spaw and pitx2c, also became random in the morphants. Furthermore, lefty1 was not expressed in the posterior notochord, indicating that the molecular midline barrier had been disrupted. It was also not expressed in the brain diencephalon. Kupffer's vesicle (KV size became smaller in fgfr2c morphants. Furthermore, KV cilia were shorter in fgfr2c morphants. We conclude that the fgfr2c isoform plays an important role in the left-right asymmetry during zebrafish development.

A variant of fibroblast growth factor receptor 2 (Fgfr2) regulates left-right asymmetry in zebrafish.

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Liu, Da-Wei; Hsu, Chia-Hao; Tsai, Su-Mei; Hsiao, Chung-Der; Wang, Wen-Pin

2011-01-01

Many organs in vertebrates are left-right asymmetrical located. For example, liver is at the right side and stomach is at the left side in human. Fibroblast growth factor (Fgf) signaling is important for left-right asymmetry. To investigate the roles of Fgfr2 signaling in zebrafish left-right asymmetry, we used splicing blocking morpholinos to specifically block the splicing of fgfr2b and fgfr2c variants, respectively. We found that the relative position of the liver and the pancreas were disrupted in fgfr2c morphants. Furthermore, the left-right asymmetry of the heart became random. Expression pattern of the laterality controlling genes, spaw and pitx2c, also became random in the morphants. Furthermore, lefty1 was not expressed in the posterior notochord, indicating that the molecular midline barrier had been disrupted. It was also not expressed in the brain diencephalon. Kupffer's vesicle (KV) size became smaller in fgfr2c morphants. Furthermore, KV cilia were shorter in fgfr2c morphants. We conclude that the fgfr2c isoform plays an important role in the left-right asymmetry during zebrafish development.

Activation of p38 MAPK pathway in the skull abnormalities of Apert syndrome Fgfr2+P253R mice

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Hill Cheryl A

2010-02-01

Full Text Available Abstract Background Apert syndrome is characterized by craniosynostosis and limb abnormalities and is primarily caused by FGFR2 +/P253R and +/S252W mutations. The former mutation is present in approximately one third whereas the latter mutation is present in two-thirds of the patients with this condition. We previously reported an inbred transgenic mouse model with the Fgfr2 +/S252W mutation on the C57BL/6J background for Apert syndrome. Here we present a mouse model for the Fgfr2+/P253R mutation. Results We generated inbred Fgfr2+/P253R mice on the same C56BL/6J genetic background and analyzed their skeletal abnormalities. 3D micro-CT scans of the skulls of the Fgfr2+/P253R mice revealed that the skull length was shortened with the length of the anterior cranial base significantly shorter than that of the Fgfr2+/S252W mice at P0. The Fgfr2+/P253R mice presented with synostosis of the coronal suture and proximate fronts with disorganized cellularity in sagittal and lambdoid sutures. Abnormal osteogenesis and proliferation were observed at the developing coronal suture and long bones of the Fgfr2+/P253R mice as in the Fgfr2+/S252W mice. Activation of mitogen-activated protein kinases (MAPK was observed in the Fgfr2+/P253R neurocranium with an increase in phosphorylated p38 as well as ERK1/2, whereas phosphorylated AKT and PKCα were not obviously changed as compared to those of wild-type controls. There were localized phenotypic and molecular variations among individual embryos with different mutations and among those with the same mutation. Conclusions Our in vivo studies demonstrated that the Fgfr2 +/P253R mutation resulted in mice with cranial features that resemble those of the Fgfr2+/S252W mice and human Apert syndrome. Activated p38 in addition to the ERK1/2 signaling pathways may mediate the mutant neurocranial phenotype. Though Apert syndrome is traditionally thought to be a consistent phenotype, our results suggest localized and regional

Constitutive activation of MEK1 in chondrocytes causes Stat1-independent achondroplasia-like dwarfism and rescues the Fgfr3-deficient mouse phenotype

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Murakami, Shunichi; Balmes, Gener; McKinney, Sandra; Zhang, Zhaoping; Givol, David; de Crombrugghe, Benoit

2004-01-01

We generated transgenic mice that express a constitutively active mutant of MEK1 in chondrocytes. These mice showed a dwarf phenotype similar to achondroplasia, the most common human dwarfism, caused by activating mutations in FGFR3. These mice displayed incomplete hypertrophy of chondrocytes in the growth plates and a general delay in endochondral ossification, whereas chondrocyte proliferation was unaffected. Immunohistochemical analysis of the cranial base in transgenic embryos showed reduced staining for collagen type X and persistent expression of Sox9 in chondrocytes. These observations indicate that the MAPK pathway inhibits hypertrophic differentiation of chondrocytes and negatively regulates bone growth without inhibiting chondrocyte proliferation. Expression of a constitutively active mutant of MEK1 in chondrocytes of Fgfr3-deficient mice inhibited skeletal overgrowth, strongly suggesting that regulation of bone growth by FGFR3 is mediated at least in part by the MAPK pathway. Although loss of Stat1 restored the reduced chondrocyte proliferation in mice expressing an achondroplasia mutant of Fgfr3, it did not rescue the reduced hypertrophic zone, the delay in formation of secondary ossification centers, and the achondroplasia-like phenotype. These observations suggest a model in which Fgfr3 signaling inhibits bone growth by inhibiting chondrocyte differentiation through the MAPK pathway and by inhibiting chondrocyte proliferation through Stat1. PMID:14871928

Polysaccharides and their depolymerized fragments from Costaria costata: Molecular weight and sulfation-dependent anticoagulant and FGF/FGFR signal activating activities.

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Hou, Ningning; Zhang, Meng; Xu, Yingjie; Sun, Zhongmin; Wang, Jing; Zhang, Lijuan; Zhang, Quanbin

2017-12-01

Crude polysaccharides from Costaria costata were extracted by hot water and further fractionated by anion exchange chromatography into three polysaccharide fractions. Three low molecular weight fragments were then prepared by degradation of the polysaccharides with hydrogen peroxide and ascorbic acid. The structural features of the polysaccharides and their low molecular weight fragments were elucidated for the first time based on the HGPC, FT-IR, NMR, MS, monosaccharide composition, and other chemical analyses. Their anticoagulant and FGF-1, -2, -7, -8, -9, -10/FGFR1c signaling activation activities in BaF3 cells were also examined. Our studies showed that the polysaccharides were sulfated at different positions of galactose and fucose residues. The APTT-, PT- and TT-based anticoagulant assay results indicated that a high molecular weight and a higher degree of sulfation were essential for their anticoagulant activities. In contrast, not only the polysaccharides but also the depolymerized fragments showed significant FGF/FGFR signal activating activities in a FGF-, molecular weight-, and sulfation-dependent manner. The results presented in current study demonstrated the potential use of the polysaccharides and their fragments as anticoagulants and FGF signal regulators. Copyright © 2017 Elsevier B.V. All rights reserved.

Effect of the achondroplasia mutation on FGFR3 dimerization and FGFR3 structural response to fgf1 and fgf2: A quantitative FRET study in osmotically derived plasma membrane vesicles.

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Sarabipour, Sarvenaz; Hristova, Kalina

2016-07-01

The G380R mutation in the transmembrane domain of FGFR3 is a germline mutation responsible for most cases of Achondroplasia, a common form of human dwarfism. Here we use quantitative Fӧster Resonance Energy Transfer (FRET) and osmotically derived plasma membrane vesicles to study the effect of the achondroplasia mutation on the early stages of FGFR3 signaling in response to the ligands fgf1 and fgf2. Using a methodology that allows us to capture structural changes on the cytoplasmic side of the membrane in response to ligand binding to the extracellular domain of FGFR3, we observe no measurable effects of the G380R mutation on FGFR3 ligand-bound dimer configurations. Instead, the most notable effect of the achondroplasia mutation is increased propensity for FGFR3 dimerization in the absence of ligand. This work reveals new information about the molecular events that underlie the achondroplasia phenotype, and highlights differences in FGFR3 activation due to different single amino-acid pathogenic mutations. Copyright © 2016 Elsevier B.V. All rights reserved.

Postnatal soluble FGFR3 therapy rescues achondroplasia symptoms and restores bone growth in mice.

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Garcia, Stéphanie; Dirat, Béatrice; Tognacci, Thomas; Rochet, Nathalie; Mouska, Xavier; Bonnafous, Stéphanie; Patouraux, Stéphanie; Tran, Albert; Gual, Philippe; Le Marchand-Brustel, Yannick; Gennero, Isabelle; Gouze, Elvire

2013-09-18

Achondroplasia is a rare genetic disease characterized by abnormal bone development, resulting in short stature. It is caused by a single point mutation in the gene coding for fibroblast growth factor receptor 3 (FGFR3), which leads to prolonged activation upon ligand binding. To prevent excessive intracellular signaling and rescue the symptoms of achondroplasia, we have developed a recombinant protein therapeutic approach using a soluble form of human FGFR3 (sFGFR3), which acts as a decoy receptor and prevents FGF from binding to mutant FGFR3. sFGFR3 was injected subcutaneously to newborn Fgfr3(ach/+) mice-the mouse model of achondroplasia-twice per week throughout the growth period during 3 weeks. Effective maturation of growth plate chondrocytes was restored in bones of treated mice, with a dose-dependent enhancement of skeletal growth in Fgfr3(ach/+) mice. This resulted in normal stature and a significant decrease in mortality and associated complications, without any evidence of toxicity. These results describe a new approach for restoring bone growth and suggest that sFGFR3 could be a potential therapy for children with achondroplasia and related disorders.

FGF19 regulates cell proliferation, glucose and bile acid metabolism via FGFR4-dependent and independent pathways.

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Ai-Luen Wu

Full Text Available Fibroblast growth factor 19 (FGF19 is a hormone-like protein that regulates carbohydrate, lipid and bile acid metabolism. At supra-physiological doses, FGF19 also increases hepatocyte proliferation and induces hepatocellular carcinogenesis in mice. Much of FGF19 activity is attributed to the activation of the liver enriched FGF Receptor 4 (FGFR4, although FGF19 can activate other FGFRs in vitro in the presence of the coreceptor βKlotho (KLB. In this report, we investigate the role of FGFR4 in mediating FGF19 activity by using Fgfr4 deficient mice as well as a variant of FGF19 protein (FGF19v which is specifically impaired in activating FGFR4. Our results demonstrate that FGFR4 activation mediates the induction of hepatocyte proliferation and the suppression of bile acid biosynthesis by FGF19, but is not essential for FGF19 to improve glucose and lipid metabolism in high fat diet fed mice as well as in leptin-deficient ob/ob mice. Thus, FGF19 acts through multiple receptor pathways to elicit pleiotropic effects in regulating nutrient metabolism and cell proliferation.

Expression of FGFR3 during human testis development and in germ cell-derived tumours of young adults

DEFF Research Database (Denmark)

Ewen, Katherine A; Olesen, Inge A; Winge, Sofia B

2013-01-01

development and to ascertain whether FGFR3 signalling is linked to germ cell proliferation and the pathogenesis of testicular germ cell tumours (TGCTs) of young adult men. Using RT-PCR, immunohistochemistry and Western blotting, we examined 58 specimens of human testes throughout development for FGFR3...... expression, and then compared expression of FGFR3 with proliferation markers (PCNA or Ki67). We also analysed for FGFR3 expression 30 TGCTs and 28 testes containing the tumour precursor cell, carcinoma in situ (CIS). Fetal and adult testes expressed exclusively the FGFR3IIIc isoform. FGFR3 protein expression...... was restricted to the cytoplasm/plasma membrane of spermatogonia and was most prevalent at mid-gestation, infancy and from puberty onwards. Phosphorylated (p)FGFR was detected in pre-spermatogonia at mid-gestation and in spermatogonia during puberty and in the adult testis. Throughout normal human testis...

The selectivity of receptor tyrosine kinase signaling is controlled by a secondary SH2 domain binding site.

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Bae, Jae Hyun; Lew, Erin Denise; Yuzawa, Satoru; Tomé, Francisco; Lax, Irit; Schlessinger, Joseph

2009-08-07

SH2 domain-mediated interactions represent a crucial step in transmembrane signaling by receptor tyrosine kinases. SH2 domains recognize phosphotyrosine (pY) in the context of particular sequence motifs in receptor phosphorylation sites. However, the modest binding affinity of SH2 domains to pY containing peptides may not account for and likely represents an oversimplified mechanism for regulation of selectivity of signaling pathways in living cells. Here we describe the crystal structure of the activated tyrosine kinase domain of FGFR1 in complex with a phospholipase Cgamma fragment. The structural and biochemical data and experiments with cultured cells show that the selectivity of phospholipase Cgamma binding and signaling via activated FGFR1 are determined by interactions between a secondary binding site on an SH2 domain and a region in FGFR1 kinase domain in a phosphorylation independent manner. These experiments reveal a mechanism for how SH2 domain selectivity is regulated in vivo to mediate a specific cellular process.

ARQ 087 inhibits FGFR signaling and rescues aberrant cell proliferation and differentiation in experimental models of craniosynostoses and chondrodysplasias caused by activating mutations in FGFR1, FGFR2 and FGFR3

Czech Academy of Sciences Publication Activity Database

Bálek, L.; Gudernová, I.; Veselá, Iva; Hampl, Marek; Oralová, Veronika; Kunová Bosáková, M.; Vařecha, M.; Němec, P.; Hall, T.C.; Abbadessa, G.; Hatch, N.; Buchtová, Marcela; Krejčí, P.

2017-01-01

Roč. 105, č. 1 (2017), s. 57-66 ISSN 8756-3282 R&D Projects: GA ČR(CZ) GA17-09525S; GA ČR(CZ) GA14-31540S Institutional support: RVO:67985904 Keywords : ARQ087 * fibroblast growth factor receptor * FGFR Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Developmental biology Impact factor: 4.140, year: 2016

Fibroblast growth factor receptor mediates fibroblast-dependent growth in EMMPRIN-depleted head and neck cancer tumor cells.

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Liu, Zhiyong; Hartman, Yolanda E; Warram, Jason M; Knowles, Joseph A; Sweeny, Larissa; Zhou, Tong; Rosenthal, Eben L

2011-08-01

Head and neck squamous cell carcinoma tumors (HNSCC) contain a dense fibrous stroma which is known to promote tumor growth, although the mechanism of stroma-mediated growth remains unclear. As dysplastic mucosal epithelium progresses to cancer, there is incremental overexpression of extracellular matrix metalloprotease inducer (EMMPRIN) which is associated with tumor growth and metastasis. Here, we present evidence that gain of EMMPRIN expression allows tumor growth to be less dependent on fibroblasts by modulating fibroblast growth factor receptor-2 (FGFR2) signaling. We show that silencing EMMPRIN in FaDu and SCC-5 HNSCC cell lines inhibits cell growth, but when EMMPRIN-silenced tumor cells were cocultured with fibroblasts or inoculated with fibroblasts into severe combined immunodeficient mice, the growth inhibition by silencing EMMPRIN was blunted by the presence of fibroblasts. Coculture experiments showed fibroblast-dependent tumor cell growth occurred via a paracrine signaling. Analysis of tumor gene expression revealed expression of FGFR2 was inversely related to EMMPRIN expression. To determine the role of FGFR2 signaling in EMMPRIN-silenced tumor cells, ligands and inhibitors of FGFR2 were assessed. Both FGF1 and FGF2 enhanced tumor growth in EMMPRIN-silenced cells compared with control vector-transfected cells, whereas inhibition of FGFR2 with blocking antibody or with a synthetic inhibitor (PD173074) inhibited tumor cell growth in fibroblast coculture, suggesting the importance of FGFR2 signaling in fibroblast-mediated tumor growth. Analysis of xenografted tumors revealed that EMMPRIN-silenced tumors had a larger stromal compartment compared with control. Taken together, these results suggest that EMMPRIN acquired during tumor progression promotes fibroblast-independent tumor growth.

Fibroblast growth factor receptor mediates fibroblast-dependent growth in EMMPRIN depleted head and neck cancer tumor cells

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Liu, Zhiyong; Hartman, Yolanda E.; Warram, Jason M.; Knowles, Joseph A.; Sweeny, Larrisa; Zhou, Tong; Rosenthal, Eben L.

2011-01-01

Head and neck squamous cell carcinoma tumors (HNSCC) contain a dense fibrous stroma which is known to promote tumor growth, although the mechanism of stroma mediated growth remains unclear. As dysplastic mucosal epithelium progresses to cancer there is incremental overexpression of extracellular matrix metalloprotease inducer (EMMPRIN) which is associated with tumor growth and metastasis. Here we present evidence that gain of EMMPRIN expression allows tumor growth to be less dependent on fibroblasts by modulating fibroblast growth factor receptor-2 (FGFR2) signaling. We show that silencing EMMPRIN in FaDu and SCC-5 HNSCC cell lines inhibits cell growth, but when EMMPRIN-silenced tumor cells were co-cultured with fibroblasts or inoculated with fibroblasts into SCID mice, the growth inhibition by silencing EMMPRIN was blunted by the presence of fibroblasts. Co-culture experiments demonstrated fibroblast-dependent tumor cell growth occurred via a paracrine signaling. Analysis of tumor gene expression revealed expression of FGFR2 was inversely related to EMMPRIN expression. To determine the role of FGFR2 signaling in EMMPRIN silenced tumor cells, ligands and inhibitors of FGFR2 were assessed. Both FGF1 and FGF2 enhanced tumor growth in EMMPRIN silenced cells compared to control vector transfected cells, while inhibition of FGFR2 with blocking antibody or with a synthetic inhibitor (PD173074) inhibited tumor cell growth in fibroblast co-culture, suggesting the importance of FGFR2 signaling in fibroblast mediated tumor growth. Analysis of xenografted tumors revealed EMMPRIN silenced tumors had a larger stromal compartment compared to control. Taken together, these results suggest that EMMPRIN acquired during tumor progression promotes fibroblast independent tumor growth. PMID:21665938

Fibroblast Growth Factor (FGF-2) and Its Receptors FGFR-2 and FGFR-3 May Be Putative Biomarkers of Malignant Transformation of Potentially Malignant Oral Lesions into Oral Squamous Cell Carcinoma.

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Nayak, Seema; Goel, Madhu Mati; Makker, Annu; Bhatia, Vikram; Chandra, Saumya; Kumar, Sandeep; Agarwal, S P

2015-01-01

There are several factors like angiogenesis, lymphangiogenesis, genetic alterations, mutational factors that are involved in malignant transformation of potentially malignant oral lesions (PMOLs) to oral squamous cell carcinoma (OSCC). Fibroblast growth factor-2 (FGF-2) is one of the prototypes of the large family of growth factors that bind heparin. FGF-2 induces angiogenesis and its receptors may play a role in synthesis of collagen. FGFs are involved in transmission of signals between the epithelium and connective tissue, and influence growth and differentiation of a wide variety of tissue including epithelia. The present study was undertaken to analyze expression of FGF-2 and its receptors FGFR-2 and FGFR-3 in 72 PMOLs, 108 OSCC and 52 healthy controls, and their role in risk assessment for malignant transformation of Leukoplakia (LKP) and Oral submucous fibrosis (OSMF) to OSCC. Immunohistochemistry was performed using antibodies against FGF-2, FGFR-2 and FGFR-3. IHC results were validated by Real Time PCR. Expression of FGF-2, FGFR-2 and FGFR-3 was upregulated from PMOLs to OSCC. While 90% (9/10) of PMOLs which showed malignant transformation (transformed) expressed FGF-2, only 24.19% cases (15/62) of PMOLs which were not transformed (untransformed) to OSCC expressed FGF-2. Similarly, FGFR-2 expression was seen in 16/62 (25.81%) of untransformed PMOLs and 8/10 (80%) cases of transformed PMOLs. FGFR-3 expression was observed in 23/62 (37.10%) cases of untransformed PMOLs and 6/10 (60%) cases of transformed PMOLs. A significant association of FGF-2 and FGFR-2 expression with malignant transformation from PMOLs to OSCC was observed both at phenotypic and molecular level. The results suggest that FGF-2 and FGFR-2 may be useful as biomarkers of malignant transformation in patients with OSMF and LKP.

The binding of NCAM to FGFR1 induces a specific cellular response mediated by receptor trafficking

DEFF Research Database (Denmark)

Francavilla, Chiara; Cattaneo, Paola; Berezin, Vladimir

2009-01-01

different from that elicited by FGF-2. In contrast to FGF-induced degradation of endocytic FGFR1, NCAM promotes the stabilization of the receptor, which is recycled to the cell surface in a Rab11- and Src-dependent manner. In turn, FGFR1 recycling is required for NCAM-induced sustained activation of various...

Genetic variants in FGFR2 and FGFR4 genes and skin cancer risk in the Nurses' Health Study

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Nan, Hongmei; Qureshi, Abrar A; Hunter, David J; Han, Jiali

2009-01-01

The human fibroblast growth factor (FGF) and its receptor (FGFR) play an important role in tumorigenesis. Deregulation of the FGFR2 gene has been identified in a number of cancer sites. Overexpression of the FGFR4 protein has been linked to cutaneous melanoma progression. Previous studies reported associations between genetic variants in the FGFR2 and FGFR4 genes and development of various cancers. We evaluated the associations of four genetic variants in the FGFR2 gene highly related to breast cancer risk and the three common tag-SNPs in the FGFR4 gene with skin cancer risk in a nested case-control study of Caucasians within the Nurses' Health Study (NHS) among 218 melanoma cases, 285 squamous cell carcinoma (SCC) cases, 300 basal cell carcinoma (BCC) cases, and 870 controls. We found no evidence for associations between these seven genetic variants and the risks of melanoma and nonmelanocytic skin cancer. Given the power of this study, we did not detect any contribution of genetic variants in the FGFR2 or FGFR4 genes to inherited predisposition to skin cancer among Caucasian women

Tyrosine kinase inhibitor NVP-BGJ398 functionally improves FGFR3-related dwarfism in mouse model.

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Komla-Ebri, Davide; Dambroise, Emilie; Kramer, Ina; Benoist-Lasselin, Catherine; Kaci, Nabil; Le Gall, Cindy; Martin, Ludovic; Busca, Patricia; Barbault, Florent; Graus-Porta, Diana; Munnich, Arnold; Kneissel, Michaela; Di Rocco, Federico; Biosse-Duplan, Martin; Legeai-Mallet, Laurence

2016-05-02

Achondroplasia (ACH) is the most frequent form of dwarfism and is caused by gain-of-function mutations in the fibroblast growth factor receptor 3-encoding (FGFR3-encoding) gene. Although potential therapeutic strategies for ACH, which aim to reduce excessive FGFR3 activation, have emerged over many years, the use of tyrosine kinase inhibitor (TKI) to counteract FGFR3 hyperactivity has yet to be evaluated. Here, we have reported that the pan-FGFR TKI, NVP-BGJ398, reduces FGFR3 phosphorylation and corrects the abnormal femoral growth plate and calvaria in organ cultures from embryos of the Fgfr3Y367C/+ mouse model of ACH. Moreover, we demonstrated that a low dose of NVP-BGJ398, injected subcutaneously, was able to penetrate into the growth plate of Fgfr3Y367C/+ mice and modify its organization. Improvements to the axial and appendicular skeletons were noticeable after 10 days of treatment and were more extensive after 15 days of treatment that started from postnatal day 1. Low-dose NVP-BGJ398 treatment reduced intervertebral disc defects of lumbar vertebrae, loss of synchondroses, and foramen-magnum shape anomalies. NVP-BGJ398 inhibited FGFR3 downstream signaling pathways, including MAPK, SOX9, STAT1, and PLCγ, in the growth plates of Fgfr3Y367C/+ mice and in cultured chondrocyte models of ACH. Together, our data demonstrate that NVP-BGJ398 corrects pathological hallmarks of ACH and support TKIs as a potential therapeutic approach for ACH.

Myeloproliferative disorder FOP-FGFR1 fusion kinase recruits phosphoinositide-3 kinase and phospholipase Cγ at the centrosome

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Tassin Anne-Marie

2008-04-01

Full Text Available Abstract Background The t(6;8 translocation found in rare and agressive myeloproliferative disorders results in a chimeric gene encoding the FOP-FGFR1 fusion protein. This protein comprises the N-terminal region of the centrosomal protein FOP and the tyrosine kinase of the FGFR1 receptor. FOP-FGFR1 is localized at the centrosome where it exerts a constitutive kinase activity. Results We show that FOP-FGFR1 interacts with the large centrosomal protein CAP350 and that CAP350 is necessary for FOP-FGFR1 localisation at centrosome. FOP-FGFR1 activates the phosphoinositide-3 kinase (PI3K pathway. We show that p85 interacts with tyrosine 475 of FOP-FGFR1, which is located in a YXXM consensus binding sequence for an SH2 domain of p85. This interaction is in part responsible for PI3K activation. Ba/F3 cells that express FOP-FGFR1 mutated at tyrosine 475 have reduced proliferative ability. Treatment with PI3K pathway inhibitors induces death of FOP-FGFR1 expressing cells. FOP-FGFR1 also recruits phospholipase Cγ1 (PLCγ1 at the centrosome. We show that this enzyme is recruited by FOP-FGFR1 at the centrosome during interphase. Conclusion These results delineate a particular type of oncogenic mechanism by which an ectopic kinase recruits its substrates at the centrosome whence unappropriate signaling induces continuous cell growth and MPD.

Fibroblast growth factor 10-fibroblast growth factor receptor 2b mediated signaling is not required for adult glandular stomach homeostasis.

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Allison L Speer

Full Text Available The signaling pathways that are essential for gastric organogenesis have been studied in some detail; however, those that regulate the maintenance of the gastric epithelium during adult homeostasis remain unclear. In this study, we investigated the role of Fibroblast growth factor 10 (FGF10 and its main receptor, Fibroblast growth factor receptor 2b (FGFR2b, in adult glandular stomach homeostasis. We first showed that mouse adult glandular stomach expressed Fgf10, its receptors, Fgfr1b and Fgfr2b, and most of the other FGFR2b ligands (Fgf1, Fgf7, Fgf22 except for Fgf3 and Fgf20. Fgf10 expression was mesenchymal whereas FGFR1 and FGFR2 expression were mostly epithelial. Studying double transgenic mice that allow inducible overexpression of Fgf10 in adult mice, we showed that Fgf10 overexpression in normal adult glandular stomach increased epithelial proliferation, drove mucous neck cell differentiation, and reduced parietal and chief cell differentiation. Although a similar phenotype can be associated with the development of metaplasia, we found that Fgf10 overexpression for a short duration does not cause metaplasia. Finally, investigating double transgenic mice that allow the expression of a soluble form of Fgfr2b, FGF10's main receptor, which acts as a dominant negative, we found no significant changes in gastric epithelial proliferation or differentiation in the mutants. Our work provides evidence, for the first time, that the FGF10-FGFR2b signaling pathway is not required for epithelial proliferation and differentiation during adult glandular stomach homeostasis.

Fibroblast growth factor receptor signaling is essential for normal mammary gland development and stem cell function.

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Pond, Adam C; Bin, Xue; Batts, Torey; Roarty, Kevin; Hilsenbeck, Susan; Rosen, Jeffrey M

2013-01-01

Fibroblast growth factor (FGF) signaling plays an important role in embryonic stem cells and adult tissue homeostasis, but the function of FGFs in mammary gland stem cells is less well defined. Both FGFR1 and FGFR2 are expressed in basal and luminal mammary epithelial cells (MECs), suggesting that together they might play a role in mammary gland development and stem cell dynamics. Previous studies have demonstrated that the deletion of FGFR2 resulted only in transient developmental defects in branching morphogenesis. Using a conditional deletion strategy, we investigated the consequences of FGFR1 deletion alone and then the simultaneous deletion of both FGFR1 and FGFR2 in the mammary epithelium. FGFR1 deletion using a keratin 14 promoter-driven Cre-recombinase resulted in an early, yet transient delay in development. However, no reduction in functional outgrowth potential was observed following limiting dilution transplantation analysis. In contrast, a significant reduction in outgrowth potential was observed upon the deletion of both FGFR1 and FGFR2 in MECs using adenovirus-Cre. Additionally, using a fluorescent reporter mouse model to monitor Cre-mediated recombination, we observed a competitive disadvantage following transplantation of both FGFR1/R2-null MECs, most prominently in the basal epithelial cells. This correlated with the complete loss of the mammary stem cell repopulating population in the FGFR1/R2-attenuated epithelium. FGFR1/R2-null MECs were partially rescued in chimeric outgrowths containing wild-type MECs, suggesting the potential importance of paracrine mechanisms involved in the maintenance of the basal epithelial stem cells. These studies document the requirement for functional FGFR signaling in mammary stem cells during development. Copyright © 2012 AlphaMed Press.

Targeting fibroblast growth factor receptor signaling inhibits prostate cancer progression.

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Feng, Shu; Shao, Longjiang; Yu, Wendong; Gavine, Paul; Ittmann, Michael

2012-07-15

Extensive correlative studies in human prostate cancer as well as studies in vitro and in mouse models indicate that fibroblast growth factor receptor (FGFR) signaling plays an important role in prostate cancer progression. In this study, we used a probe compound for an FGFR inhibitor, which potently inhibits FGFR-1-3 and significantly inhibits FGFR-4. The purpose of this study is to determine whether targeting FGFR signaling from all four FGFRs will have in vitro activities consistent with inhibition of tumor progression and will inhibit tumor progression in vivo. Effects of AZ8010 on FGFR signaling and invasion were analyzed using immortalized normal prostate epithelial (PNT1a) cells and PNT1a overexpressing FGFR-1 or FGFR-4. The effect of AZ8010 on invasion and proliferation in vitro was also evaluated in prostate cancer cell lines. Finally, the impact of AZ8010 on tumor progression in vivo was evaluated using a VCaP xenograft model. AZ8010 completely inhibits FGFR-1 and significantly inhibits FGFR-4 signaling at 100 nmol/L, which is an achievable in vivo concentration. This results in marked inhibition of extracellular signal-regulated kinase (ERK) phosphorylation and invasion in PNT1a cells expressing FGFR-1 and FGFR-4 and all prostate cancer cell lines tested. Treatment in vivo completely inhibited VCaP tumor growth and significantly inhibited angiogenesis and proliferation and increased cell death in treated tumors. This was associated with marked inhibition of ERK phosphorylation in treated tumors. Targeting FGFR signaling is a promising new approach to treating aggressive prostate cancer.

Combinatorial Pharmacophore-Based 3D-QSAR Analysis and Virtual Screening of FGFR1 Inhibitors

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Nannan Zhou

2015-06-01

Full Text Available The fibroblast growth factor/fibroblast growth factor receptor (FGF/FGFR signaling pathway plays crucial roles in cell proliferation, angiogenesis, migration, and survival. Aberration in FGFRs correlates with several malignancies and disorders. FGFRs have proved to be attractive targets for therapeutic intervention in cancer, and it is of high interest to find FGFR inhibitors with novel scaffolds. In this study, a combinatorial three-dimensional quantitative structure-activity relationship (3D-QSAR model was developed based on previously reported FGFR1 inhibitors with diverse structural skeletons. This model was evaluated for its prediction performance on a diverse test set containing 232 FGFR inhibitors, and it yielded a SD value of 0.75 pIC50 units from measured inhibition affinities and a Pearson’s correlation coefficient R2 of 0.53. This result suggests that the combinatorial 3D-QSAR model could be used to search for new FGFR1 hit structures and predict their potential activity. To further evaluate the performance of the model, a decoy set validation was used to measure the efficiency of the model by calculating EF (enrichment factor. Based on the combinatorial pharmacophore model, a virtual screening against SPECS database was performed. Nineteen novel active compounds were successfully identified, which provide new chemical starting points for further structural optimization of FGFR1 inhibitors.

PD173074, a selective FGFR inhibitor, reverses MRP7 (ABCC10-mediated MDR

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Nagaraju Anreddy

2014-06-01

Full Text Available Multidrug resistance protein 7 (MRP7, ABCC10 is a recently identified member of the ATP-binding cassette (ABC transporter family, which adequately confers resistance to a diverse group of antineoplastic agents, including taxanes, vinca alkaloids and nucleoside analogs among others. Clinical studies indicate an increased MRP7 expression in non-small cell lung carcinomas (NSCLC compared to a normal healthy lung tissue. Recent studies revealed increased paclitaxel sensitivity in the Mrp7−/− mouse model compared to their wild-type counterparts. This demonstrates that MRP7 is a key contributor in developing drug resistance. Recently our group reported that PD173074, a specific fibroblast growth factor receptor (FGFR inhibitor, could significantly reverse P-glycoprotein-mediated MDR. However, whether PD173074 can interact with and inhibit other MRP members is unknown. In the present study, we investigated the ability of PD173074 to reverse MRP7-mediated MDR. We found that PD173074, at non-toxic concentration, could significantly increase the cellular sensitivity to MRP7 substrates. Mechanistic studies indicated that PD173074 (1 μmol/L significantly increased the intracellular accumulation and in-turn decreased the efflux of paclitaxel by inhibiting the transport activity without altering expression levels of the MRP7 protein, thereby representing a promising therapeutic agent in the clinical treatment of chemoresistant cancer patients.

FGFR3 mutation causes abnormal membranous ossification in achondroplasia.

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Di Rocco, Federico; Biosse Duplan, Martin; Heuzé, Yann; Kaci, Nabil; Komla-Ebri, Davide; Munnich, Arnold; Mugniery, Emilie; Benoist-Lasselin, Catherine; Legeai-Mallet, Laurence

2014-06-01

FGFR3 gain-of-function mutations lead to both chondrodysplasias and craniosynostoses. Achondroplasia (ACH), the most frequent dwarfism, is due to an FGFR3-activating mutation which results in impaired endochondral ossification. The effects of the mutation on membranous ossification are unknown. Fgfr3(Y367C/+) mice mimicking ACH and craniofacial analysis of patients with ACH and FGFR3-related craniosynostoses provide an opportunity to address this issue. Studying the calvaria and skull base, we observed abnormal cartilage and premature fusion of the synchondroses leading to modifications of foramen magnum shape and size in Fgfr3(Y367C/+) mice, ACH and FGFR3-related craniosynostoses patients. Partial premature fusion of the coronal sutures and non-ossified gaps in frontal bones were also present in Fgfr3(Y367C/+) mice and ACH patients. Our data provide strong support that not only endochondral ossification but also membranous ossification is severely affected in ACH. Demonstration of the impact of FGFR3 mutations on craniofacial development should initiate novel pharmacological and surgical therapeutic approaches.

Tumor heterogeneity of fibroblast growth factor receptor 3 (FGFR3) mutations in invasive bladder cancer: implications for perioperative anti-FGFR3 treatment.

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Pouessel, D; Neuzillet, Y; Mertens, L S; van der Heijden, M S; de Jong, J; Sanders, J; Peters, D; Leroy, K; Manceau, A; Maille, P; Soyeux, P; Moktefi, A; Semprez, F; Vordos, D; de la Taille, A; Hurst, C D; Tomlinson, D C; Harnden, P; Bostrom, P J; Mirtti, T; Horenblas, S; Loriot, Y; Houédé, N; Chevreau, C; Beuzeboc, P; Shariat, S F; Sagalowsky, A I; Ashfaq, R; Burger, M; Jewett, M A S; Zlotta, A R; Broeks, A; Bapat, B; Knowles, M A; Lotan, Y; van der Kwast, T H; Culine, S; Allory, Y; van Rhijn, B W G

2016-07-01

Fibroblast growth factor receptor 3 (FGFR3) is an actionable target in bladder cancer. Preclinical studies show that anti-FGFR3 treatment slows down tumor growth, suggesting that this tyrosine kinase receptor is a candidate for personalized bladder cancer treatment, particularly in patients with mutated FGFR3. We addressed tumor heterogeneity in a large multicenter, multi-laboratory study, as this may have significant impact on therapeutic response. We evaluated possible FGFR3 heterogeneity by the PCR-SNaPshot method in the superficial and deep compartments of tumors obtained by transurethral resection (TUR, n = 61) and in radical cystectomy (RC, n = 614) specimens and corresponding cancer-positive lymph nodes (LN+, n = 201). We found FGFR3 mutations in 13/34 (38%) T1 and 8/27 (30%) ≥T2-TUR samples, with 100% concordance between superficial and deeper parts in T1-TUR samples. Of eight FGFR3 mutant ≥T2-TUR samples, only 4 (50%) displayed the mutation in the deeper part. We found 67/614 (11%) FGFR3 mutations in RC specimens. FGFR3 mutation was associated with pN0 (P < 0.001) at RC. In 10/201 (5%) LN+, an FGFR3 mutation was found, all concordant with the corresponding RC specimen. In the remaining 191 cases, RC and LN+ were both wild type. FGFR3 mutation status seems promising to guide decision-making on adjuvant anti-FGFR3 therapy as it appeared homogeneous in RC and LN+. Based on the results of TUR, the deep part of the tumor needs to be assessed if neoadjuvant anti-FGFR3 treatment is considered. We conclude that studies on the heterogeneity of actionable molecular targets should precede clinical trials with these drugs in the perioperative setting. © The Author 2016. Published by Oxford University Press on behalf of the European Society for Medical Oncology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Conserved intron positions in FGFR genes reflect the modular structure of FGFR and reveal stepwise addition of domains to an already complex ancestral FGFR.

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Rebscher, Nicole; Deichmann, Christina; Sudhop, Stefanie; Fritzenwanker, Jens Holger; Green, Stephen; Hassel, Monika

2009-10-01

We have analyzed the evolution of fibroblast growth factor receptor (FGFR) tyrosine kinase genes throughout a wide range of animal phyla. No evidence for an FGFR gene was found in Porifera, but we tentatively identified an FGFR gene in the placozoan Trichoplax adhaerens. The gene encodes a protein with three immunoglobulin-like domains, a single-pass transmembrane, and a split tyrosine kinase domain. By superimposing intron positions of 20 FGFR genes from Placozoa, Cnidaria, Protostomia, and Deuterostomia over the respective protein domain structure, we identified ten ancestral introns and three conserved intron groups. Our analysis shows (1) that the position of ancestral introns correlates to the modular structure of FGFRs, (2) that the acidic domain very likely evolved in the last common ancestor of triploblasts, (3) that splicing of IgIII was enabled by a triploblast-specific insertion, and (4) that IgI is subject to substantial loss or duplication particularly in quickly evolving genomes. Moreover, intron positions in the catalytic domain of FGFRs map to the borders of protein subdomains highly conserved in other serine/threonine kinases. Nevertheless, these introns were introduced in metazoan receptor tyrosine kinases exclusively. Our data support the view that protein evolution dating back to the Cambrian explosion took place in such a short time window that only subtle changes in the domain structure are detectable in extant representatives of animal phyla. We propose that the first multidomain FGFR originated in the last common ancestor of Placozoa, Cnidaria, and Bilateria. Additional domains were introduced mainly in the ancestor of triploblasts and in the Ecdysozoa.

Development of RNA-FISH Assay for Detection of Oncogenic FGFR3-TACC3 Fusion Genes in FFPE Samples.

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Masahiro Kurobe

Full Text Available Oncogenic FGFR3-TACC3 fusions and FGFR3 mutations are target candidates for small molecule inhibitors in bladder cancer (BC. Because FGFR3 and TACC3 genes are located very closely on chromosome 4p16.3, detection of the fusion by DNA-FISH (fluorescent in situ hybridization is not a feasible option. In this study, we developed a novel RNA-FISH assay using branched DNA probe to detect FGFR3-TACC3 fusions in formaldehyde-fixed paraffin-embedded (FFPE human BC samples.The RNA-FISH assay was developed and validated using a mouse xenograft model with human BC cell lines. Next, we assessed the consistency of the RNA-FISH assay using 104 human BC samples. In this study, primary BC tissues were stored as frozen and FFPE tissues. FGFR3-TACC3 fusions were independently detected in FFPE sections by the RNA-FISH assay and in frozen tissues by RT-PCR. We also analyzed the presence of FGFR3 mutations by targeted sequencing of genomic DNA extracted from deparaffinized FFPE sections.FGFR3-TACC3 fusion transcripts were identified by RNA-FISH and RT-PCR in mouse xenograft FFPE tissues using the human BC cell lines RT112 and RT4. These cell lines have been reported to be fusion-positive. Signals for FGFR3-TACC3 fusions by RNA-FISH were positive in 2/60 (3% of non-muscle-invasive BC (NMIBC and 2/44 (5% muscle-invasive BC (MIBC patients. The results of RT-PCR of all 104 patients were identical to those of RNA-FISH. FGFR3 mutations were detected in 27/60 (45% NMIBC and 8/44 (18% MIBC patients. Except for one NMIBC patient, FGFR3 mutation and FGFR3-TACC3 fusion were mutually exclusive.We developed an RNA-FISH assay for detection of the FGFR3-TACC3 fusion in FFPE samples of human BC tissues. Screening for not only FGFR3 mutations, but also for FGFR3-TACC3 fusion transcripts has the potential to identify additional patients that can be treated with FGFR inhibitors.

Protein arginine methyltransferase 5 promotes lung cancer metastasis via the epigenetic regulation of miR-99 family/FGFR3 signaling.

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Jing, Pengyu; Zhao, Nan; Ye, Mingxiang; Zhang, Yong; Zhang, Zhipei; Sun, Jianyong; Wang, Zhengxin; Zhang, Jian; Gu, Zhongping

2018-07-28

Protein arginine methyltransferase 5 (PRMT5) functions as a tumor initiator to regulate several cancer progressions, such as proliferation and apoptosis, by catalyzing the symmetrical dimethylation (me2s) of arginine residues within targeted molecules. However, the exact role of PRMT5-mediated metastasis in lung cancer is not fully understood. Here, we illustrated its potential effects in lung cancer metastasis in vivo and vitro. PRMT5 was frequently overexpressed in lung tumors, and its expression was positively related to tumor stages, lymphatic metastasis and poor outcome. In this model, PRMT5 repressed the transcription of the miR-99 family by symmetrical dimethylation of histone H4R3, which increased FGFR3 expression and in turn activated Erk1/2 and Akt, leading to cell growth and metastasis in lung cancer. Furthermore, loss of PRMT5 exerted anti-metastasis effects on lung cancer progression by blocking histone-modification of miR-99 family. Overall, this study provides new insights into the PRMT5/miR-99 family/FGFR3 axis in regulating lung cancer progression and identifies PRMT5 as a promising prognostic biomarker and therapeutic target. Copyright © 2018 The Author(s). Published by Elsevier B.V. All rights reserved.

A novel mouse Fgfr2 mutant, hobbyhorse (hob, exhibits complete XY gonadal sex reversal.

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Pam Siggers

Full Text Available The secreted molecule fibroblast growth factor 9 (FGF9 plays a critical role in testis determination in the mouse. In embryonic gonadal somatic cells it is required for maintenance of SOX9 expression, a key determinant of Sertoli cell fate. Conditional gene targeting studies have identified FGFR2 as the main gonadal receptor for FGF9 during sex determination. However, such studies can be complicated by inefficient and variable deletion of floxed alleles, depending on the choice of Cre deleter strain. Here, we report a novel, constitutive allele of Fgfr2, hobbyhorse (hob, which was identified in an ENU-based forward genetic screen for novel testis-determining loci. Fgr2hob is caused by a C to T mutation in the invariant exon 7, resulting in a polypeptide with a mis-sense mutation at position 263 (Pro263Ser in the third extracellular immunoglobulin-like domain of FGFR2. Mutant homozygous embryos show severe limb and lung defects and, when on the sensitised C57BL/6J (B6 genetic background, undergo complete XY gonadal sex reversal associated with failure to maintain expression of Sox9. Genetic crosses employing a null mutant of Fgfr2 suggest that Fgr2hob is a hypomorphic allele, affecting both the FGFR2b and FGFR2c splice isoforms of the receptor. We exploited the consistent phenotype of this constitutive mutant by analysing MAPK signalling at the sex-determining stage of gonad development, but no significant abnormalities in mutant embryos were detected.

Clinical and biological characteristics of cervical neoplasias with FGFR3 mutation

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Thiery Jean

2005-05-01

Full Text Available Abstract Background We have previously reported activating mutations of the gene coding for the fibroblast growth factor receptor 3 (FGFR3 in invasive cervical carcinoma. To further analyze the role of FGFR3 in cervical tumor progression, we extended our study to screen a total of 75 invasive tumors and 80 cervical intraepithelial neoplasias (40 low-grade and 40 high-grade lesions. Results Using single strand conformation polymorphism (SSCP followed by DNA sequencing, we found FGFR3 mutation (S249C in all cases in 5% of invasive cervical carcinomas and no mutation in intraepithelial lesions. These results suggest that, unlike in bladder carcinoma, FGFR3 mutation does not or rarely occur in non invasive lesions. Compared to patients with wildtype FGFR3 tumor, patients with S249C FGFR3 mutated tumors were older (mean age 64 vs. 49.4 years, P = 0.02, and were more likely to be associated with a non-16/18 HPV type in their tumor. Gene expression analysis demonstrated that FGFR3 mutated tumors were associated with higher FGFR3b mRNA expression levels compared to wildtype FGFR3 tumors. Supervised analysis of Affymetrix expression data identified a significant number of genes specifically differentially expressed in tumors with respect to FGFR3 mutation status. Conclusion This study suggest that tumors with FGFR3 mutation appear to have distinctive clinical and biological characteristics that may help in defining a population of patients for FGFR3 mutation screening.

RACK1 forms a complex with FGFR1 and PKM2, and stimulates the growth and migration of squamous lung cancer cells.

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Zhou, Chengzhi; Chen, Tao; Xie, Zhanhong; Qin, Yinyin; Ou, Yangming; Zhang, Jiexia; Li, Shiyue; Chen, Rongchang; Zhong, Nanshan

2017-11-01

Phosphorylation of Pyruvate Kinase M2 (PKM2) on Tyr105 by fibroblast growth factor receptor 1 (FGFR1) has been shown to promote its nuclear localization as well as cell growth in lung cancer. Better understanding the regulation of this process would benefit the clinical treatment for lung cancer. Here, it has been found that the adaptor protein receptor for activated PKC kinase (RACK1) formed a complex with FGFR1 and PKM2, and activated the FGFR1/PKM2 signaling. Knocking down the expression of RACK1 impaired the phosphorylation on Tyr105 of PKM2 and inhibited the growth and migration of lung cancer cells, while over-expression of RACK1 in lung cancer cells led to the resistance to Erdafitinib. Moreover, knocking down the expression of RACK1 impaired the tumorigenesis of lung cancer driven by LKB loss and mutated Ras (KrasG12D). Taken together, our study demonstrated the pivotal roles of RACK1 in FGFR1/PKM2 signaling, suggesting FGFR1/RACK1/PKM2 might be a therapeutic target for lung cancer treatment. © 2017 Wiley Periodicals, Inc.

Cellular Internalization of Fibroblast Growth Factor-12 Exerts Radioprotective Effects on Intestinal Radiation Damage Independently of FGFR Signaling

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Nakayama, Fumiaki, E-mail: f_naka@nirs.go.jp [Advanced Radiation Biology Research Program, Research Center for Charged Particle Therapy, Chiba (Japan); Umeda, Sachiko [Advanced Radiation Biology Research Program, Research Center for Charged Particle Therapy, Chiba (Japan); Yasuda, Takeshi [Radiation Emergency Medicine Research Program, Research Center for Radiation Emergency Medicine, National Institute of Radiological Sciences, Chiba (Japan); Fujita, Mayumi [Advanced Radiation Biology Research Program, Research Center for Charged Particle Therapy, Chiba (Japan); Asada, Masahiro [Signaling Molecules Research Group, Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology, Tsukuba (Japan); Meineke, Viktor [Bundeswehr Institute of Radiobiology affiliated to the University of Ulm, Munich (Germany); Imamura, Toru [Signaling Molecules Research Group, Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology, Tsukuba (Japan); Imai, Takashi [Advanced Radiation Biology Research Program, Research Center for Charged Particle Therapy, Chiba (Japan)

2014-02-01

Purpose: Several fibroblast growth factors (FGFs) were shown to inhibit radiation-induced tissue damage through FGF receptor (FGFR) signaling; however, this signaling was also found to be involved in the pathogenesis of several malignant tumors. In contrast, FGF12 cannot activate any FGFRs. Instead, FGF12 can be internalized readily into cells using 2 cell-penetrating peptide domains (CPP-M, CPP-C). Therefore, this study focused on clarifying the role of FGF12 internalization in protection against radiation-induced intestinal injury. Methods and Materials: Each FGF or peptide was administered intraperitoneally to BALB/c mice in the absence of heparin 24 hours before or after total body irradiation with γ rays at 9 to 12 Gy. Several radioprotective effects were examined in the jejunum. Results: Administration of FGF12 after radiation exposure was as effective as pretreatment in significantly promoting intestinal regeneration, proliferation of crypt cells, and epithelial differentiation. Two domains, comprising amino acid residues 80 to 109 and 140 to 169 of FGF12B, were identified as being responsible for the radioprotective activity, so that deletion of both domains from FGF12B resulted in a reduction in activity. Interestingly, these regions included the CPP-M and CPP-C domains, respectively; however, CPP-C by itself did not show an antiapoptotic effect. In addition, FGF1, prototypic FGF, possesses a domain corresponding to CPP-M, whereas it lacks CPP-C, so the fusion of FGF1 with CPP-C (FGF1/CPP-C) enhanced cellular internalization and increased radioprotective activity. However, FGF1/CPP-C reduced in vitro mitogenic activity through FGFRs compared with FGF1, implying that FGFR signaling might not be essential for promoting the radioprotective effect of FGF1/CPP-C. In addition, internalized FGF12 suppressed the activation of p38α after irradiation, resulting in reduced radiation-induced apoptosis. Conclusions: These findings indicate that FGF12 can protect the

Peripheral reduction of FGFR4 with antisense oligonucleotides increases metabolic rate and lowers adiposity in diet-induced obese mice.

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Xing Xian Yu

Full Text Available Obesity is a primary risk factor for multiple metabolic disorders. Many drugs for the treatment of obesity, which mainly act through CNS as appetite suppressants, have failed during development or been removed from the market due to unacceptable adverse effects. Thus, there are very few efficacious drugs available and remains a great unmet medical need for anti-obesity drugs that increase energy expenditure by acting on peripheral tissues without severe side effects. Here, we report a novel approach involving antisense inhibition of fibroblast growth factor receptor 4 (FGFR4 in peripheral tissues. Treatment of diet-induce obese (DIO mice with FGFR4 antisense oligonucleotides (ASO specifically reduced liver FGFR4 expression that not only resulted in decrease in body weight (BW and adiposity in free-feeding conditions, but also lowered BW and adiposity under caloric restriction. In addition, combination treatment with FGFR4 ASO and rimonabant showed additive reduction in BW and adiposity. FGFR4 ASO treatment increased basal metabolic rate during free-feeding conditions and, more importantly, prevented adaptive decreases of metabolic rate induced by caloric restriction. The treatment increased fatty acid oxidation while decreased lipogenesis in both liver and fat. Mechanistic studies indicated that anti-obesity effect of FGFR4 ASO was mediated at least in part through an induction of plasma FGF15 level resulted from reduction of hepatic FGFR4 expression. The anti-obesity effect was accompanied by improvement in plasma glycemia, whole body insulin sensitivity, plasma lipid levels and liver steatosis. Therefore, FGFR4 could be a potential novel target and antisense reduction of hepatic FGFR4 expression could be an efficacious therapy as an adjunct to diet restriction or to an appetite suppressant for the treatment of obesity and related metabolic disorders.

Peripheral reduction of FGFR4 with antisense oligonucleotides increases metabolic rate and lowers adiposity in diet-induced obese mice.

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Yu, Xing Xian; Watts, Lynnetta M; Manchem, Vara Prasad; Chakravarty, Kaushik; Monia, Brett P; McCaleb, Michael L; Bhanot, Sanjay

2013-01-01

Obesity is a primary risk factor for multiple metabolic disorders. Many drugs for the treatment of obesity, which mainly act through CNS as appetite suppressants, have failed during development or been removed from the market due to unacceptable adverse effects. Thus, there are very few efficacious drugs available and remains a great unmet medical need for anti-obesity drugs that increase energy expenditure by acting on peripheral tissues without severe side effects. Here, we report a novel approach involving antisense inhibition of fibroblast growth factor receptor 4 (FGFR4) in peripheral tissues. Treatment of diet-induce obese (DIO) mice with FGFR4 antisense oligonucleotides (ASO) specifically reduced liver FGFR4 expression that not only resulted in decrease in body weight (BW) and adiposity in free-feeding conditions, but also lowered BW and adiposity under caloric restriction. In addition, combination treatment with FGFR4 ASO and rimonabant showed additive reduction in BW and adiposity. FGFR4 ASO treatment increased basal metabolic rate during free-feeding conditions and, more importantly, prevented adaptive decreases of metabolic rate induced by caloric restriction. The treatment increased fatty acid oxidation while decreased lipogenesis in both liver and fat. Mechanistic studies indicated that anti-obesity effect of FGFR4 ASO was mediated at least in part through an induction of plasma FGF15 level resulted from reduction of hepatic FGFR4 expression. The anti-obesity effect was accompanied by improvement in plasma glycemia, whole body insulin sensitivity, plasma lipid levels and liver steatosis. Therefore, FGFR4 could be a potential novel target and antisense reduction of hepatic FGFR4 expression could be an efficacious therapy as an adjunct to diet restriction or to an appetite suppressant for the treatment of obesity and related metabolic disorders.

Meclozine promotes longitudinal skeletal growth in transgenic mice with achondroplasia carrying a gain-of-function mutation in the FGFR3 gene.

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Matsushita, Masaki; Hasegawa, Satoru; Kitoh, Hiroshi; Mori, Kensaku; Ohkawara, Bisei; Yasoda, Akihiro; Masuda, Akio; Ishiguro, Naoki; Ohno, Kinji

2015-02-01

Achondroplasia (ACH) is one of the most common skeletal dysplasias causing short stature owing to a gain-of-function mutation in the FGFR3 gene, which encodes the fibroblast growth factor receptor 3. We found that meclozine, an over-the-counter drug for motion sickness, inhibited elevated FGFR3 signaling in chondrocytic cells. To examine the feasibility of meclozine administration in clinical settings, we investigated the effects of meclozine on ACH model mice carrying the heterozygous Fgfr3(ach) transgene. We quantified the effect of meclozine in bone explant cultures employing limb rudiments isolated from developing embryonic tibiae from Fgfr3(ach) mice. We found that meclozine significantly increased the full-length and cartilaginous primordia of embryonic tibiae isolated from Fgfr3(ach) mice. We next analyzed the skeletal phenotypes of growing Fgfr3(ach) mice and wild-type mice with or without meclozine treatment. In Fgfr3(ach) mice, meclozine significantly increased the body length after 2 weeks of administration. At skeletal maturity, the bone lengths including the cranium, radius, ulna, femur, tibia, and vertebrae were significantly longer in meclozine-treated Fgfr3(ach) mice than in untreated Fgfr3(ach) mice. Interestingly, meclozine also increased bone growth in wild-type mice. The plasma concentration of meclozine during treatment was within the range that has been used in clinical settings for motion sickness. Increased longitudinal bone growth in Fgfr3(ach) mice by oral administration of meclozine in a growth period suggests potential clinical feasibility of meclozine for the improvement of short stature in ACH.

K-Ras and β-catenin mutations cooperate with Fgfr3 mutations in mice to promote tumorigenesis in the skin and lung, but not in the bladder

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Imran Ahmad

2011-07-01

The human fibroblast growth factor receptor 3 (FGFR3 gene is frequently mutated in superficial urothelial cell carcinoma (UCC. To test the functional significance of FGFR3 activating mutations as a ‘driver’ of UCC, we targeted the expression of mutated Fgfr3 to the murine urothelium using Cre-loxP recombination driven by the uroplakin II promoter. The introduction of the Fgfr3 mutations resulted in no obvious effect on tumorigenesis up to 18 months of age. Furthermore, even when the Fgfr3 mutations were introduced together with K-Ras or β-catenin (Ctnnb1 activating mutations, no urothelial dysplasia or UCC was observed. Interestingly, however, owing to a sporadic ectopic Cre recombinase expression in the skin and lung of these mice, Fgfr3 mutation caused papilloma and promoted lung tumorigenesis in cooperation with K-Ras and β-catenin activation, respectively. These results indicate that activation of FGFR3 can cooperate with other mutations to drive tumorigenesis in a context-dependent manner, and support the hypothesis that activation of FGFR3 signaling contributes to human cancer.

Generation of high-affinity, internalizing anti-FGFR2 single-chain variable antibody fragment fused with Fc for targeting gastrointestinal cancers.

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Borek, Aleksandra; Sokolowska-Wedzina, Aleksandra; Chodaczek, Grzegorz; Otlewski, Jacek

2018-01-01

Fibroblast growth factor receptors (FGFRs) are promising targets for antibody-based cancer therapies, as their substantial overexpression has been found in various tumor cells. Aberrant activation of FGF receptor 2 (FGFR2) signaling through overexpression of FGFR2 and/or its ligands, mutations, or receptor amplification has been reported in multiple cancer types, including gastric, colorectal, endometrial, ovarian, breast and lung cancer. In this paper, we describe application of the phage display technology to produce a panel of high affinity single chain variable antibody fragments (scFvs) against the extracellular ligand-binding domain of FGFR2 (ECD_FGFR2). The binders were selected from the human single chain variable fragment scFv phage display libraries Tomlinson I + J and showed high specificity and binding affinity towards human FGFR2 with nanomolar KD values. To improve the affinity of the best binder selected, scFvF7, we reformatted it to a bivalent diabody format, or fused it with the Fc region (scFvF7-Fc). The scFvF7-Fc antibody construct presented the highest affinity for FGFR2, with a KD of 0.76 nM, and was selectively internalized into cancer cells overexpressing FGFR2, Snu-16 and NCI-H716. Finally, we prepared a conjugate of scFvF7-Fc with the cytotoxic drug monomethyl-auristatin E (MMAE) and evaluated its cytotoxicity. The conjugate delivered MMAE selectively to FGFR2-positive tumor cells. These results indicate that scFvF7-Fc-vcMMAE is a highly potent molecule for the treatment of cancers with FGFR2 overexpression.

Switching addictions between HER2 and FGFR2 in HER2-positive breast tumor cells: FGFR2 as a potential target for salvage after lapatinib failure

International Nuclear Information System (INIS)

Azuma, Koichi; Tsurutani, Junji; Sakai, Kazuko; Kaneda, Hiroyasu; Fujisaka, Yasuhito; Takeda, Masayuki; Watatani, Masahiro; Arao, Tokuzo; Satoh, Taroh; Okamoto, Isamu; Kurata, Takayasu; Nishio, Kazuto; Nakagawa, Kazuhiko

2011-01-01

Highlights: → A lapatinib-resistant breast cancer cell line, UACC812 (UACC812/LR), was found to harbor amplification of the FGFR2 gene. → Inhibition of the molecule by a specific inhibitor of FGFR dramatically induced growth inhibition accompanied by cell death. → Immunohistochemical analysis of patients with HER2-positive breast cancer demonstrated an association between FGFR2 expression and poor outcome for lapatinib-containing chemotherapy. -- Abstract: Agents that target HER2 have improved the prognosis of patients with HER2-amplified breast cancers. However, patients who initially respond to such targeted therapy eventually develop resistance to the treatment. We have established a line of lapatinib-resistant breast cancer cells (UACC812/LR) by chronic exposure of HER2-amplified and lapatinib-sensitive UACC812 cells to the drug. The mechanism by which UACC812/LR acquired resistance to lapatinib was explored using comprehensive gene hybridization. The FGFR2 gene in UACC812/LR was highly amplified, accompanied by overexpression of FGFR2 and reduced expression of HER2, and a cell proliferation assay showed that the IC 50 of PD173074, a small-molecule inhibitor of FGFR tyrosine kinase, was 10,000 times lower in UACC812/LR than in the parent cells. PD173074 decreased the phosphorylation of FGFR2 and substantially induced apoptosis in UACC812/LR, but not in the parent cells. FGFR2 appeared to be a pivotal molecule for the survival of UACC812/LR as they became independent of the HER2 pathway, suggesting that a switch of addiction from the HER2 to the FGFR2 pathway enabled cancer cells to become resistant to HER2-targeted therapy. The present study is the first to implicate FGFR in the development of resistance to lapatinib in cancer, and suggests that FGFR-targeted therapy might become a promising salvage strategy after lapatinib failure in patients with HER2-positive breast cancer.

Switching addictions between HER2 and FGFR2 in HER2-positive breast tumor cells: FGFR2 as a potential target for salvage after lapatinib failure

Energy Technology Data Exchange (ETDEWEB)

Azuma, Koichi [Department of Medical Oncology, Kinki University Faculty of Medicine, 377-2 Ohnohigashi, Osakasayama, Osaka 589-8511 (Japan); Tsurutani, Junji, E-mail: tsurutani_j@dotd.med.kindai.ac.jp [Department of Medical Oncology, Kinki University Faculty of Medicine, 377-2 Ohnohigashi, Osakasayama, Osaka 589-8511 (Japan); Sakai, Kazuko; Kaneda, Hiroyasu [Department of Genome Biology, Kinki University Faculty of Medicine, 377-2 Ohnohigashi, Osakasayama, Osaka 589-8511 (Japan); Fujisaka, Yasuhito; Takeda, Masayuki [Department of Medical Oncology, Kinki University Faculty of Medicine, 377-2 Ohnohigashi, Osakasayama, Osaka 589-8511 (Japan); Watatani, Masahiro [Department of Surgery, Kinki University Faculty of Medicine, 377-2 Ohnohigashi, Osakasayama, Osaka 589-8511 (Japan); Arao, Tokuzo [Department of Genome Biology, Kinki University Faculty of Medicine, 377-2 Ohnohigashi, Osakasayama, Osaka 589-8511 (Japan); Satoh, Taroh; Okamoto, Isamu; Kurata, Takayasu [Department of Medical Oncology, Kinki University Faculty of Medicine, 377-2 Ohnohigashi, Osakasayama, Osaka 589-8511 (Japan); Nishio, Kazuto [Department of Genome Biology, Kinki University Faculty of Medicine, 377-2 Ohnohigashi, Osakasayama, Osaka 589-8511 (Japan); Nakagawa, Kazuhiko [Department of Medical Oncology, Kinki University Faculty of Medicine, 377-2 Ohnohigashi, Osakasayama, Osaka 589-8511 (Japan)

2011-04-01

Highlights: {yields} A lapatinib-resistant breast cancer cell line, UACC812 (UACC812/LR), was found to harbor amplification of the FGFR2 gene. {yields} Inhibition of the molecule by a specific inhibitor of FGFR dramatically induced growth inhibition accompanied by cell death. {yields} Immunohistochemical analysis of patients with HER2-positive breast cancer demonstrated an association between FGFR2 expression and poor outcome for lapatinib-containing chemotherapy. -- Abstract: Agents that target HER2 have improved the prognosis of patients with HER2-amplified breast cancers. However, patients who initially respond to such targeted therapy eventually develop resistance to the treatment. We have established a line of lapatinib-resistant breast cancer cells (UACC812/LR) by chronic exposure of HER2-amplified and lapatinib-sensitive UACC812 cells to the drug. The mechanism by which UACC812/LR acquired resistance to lapatinib was explored using comprehensive gene hybridization. The FGFR2 gene in UACC812/LR was highly amplified, accompanied by overexpression of FGFR2 and reduced expression of HER2, and a cell proliferation assay showed that the IC{sub 50} of PD173074, a small-molecule inhibitor of FGFR tyrosine kinase, was 10,000 times lower in UACC812/LR than in the parent cells. PD173074 decreased the phosphorylation of FGFR2 and substantially induced apoptosis in UACC812/LR, but not in the parent cells. FGFR2 appeared to be a pivotal molecule for the survival of UACC812/LR as they became independent of the HER2 pathway, suggesting that a switch of addiction from the HER2 to the FGFR2 pathway enabled cancer cells to become resistant to HER2-targeted therapy. The present study is the first to implicate FGFR in the development of resistance to lapatinib in cancer, and suggests that FGFR-targeted therapy might become a promising salvage strategy after lapatinib failure in patients with HER2-positive breast cancer.

Diverse FGF receptor signaling controls astrocyte specification and proliferation

Energy Technology Data Exchange (ETDEWEB)

Kang, Kyungjun [School of Life Sciences, Gwangju Institute of Science and Technology, Oryong-dong, Buk-gu, Gwangju 500-712 (Korea, Republic of); Song, Mi-Ryoung, E-mail: msong@gist.ac.kr [School of Life Sciences, Gwangju Institute of Science and Technology, Oryong-dong, Buk-gu, Gwangju 500-712 (Korea, Republic of); Bioimaging Research Center and Cell Dynamics Research Center, Gwangju Institute of Science and Technology, Oryong-dong, Buk-gu, Gwangju 500-712 (Korea, Republic of)

2010-05-07

During CNS development, pluripotency neuronal progenitor cells give rise in succession to neurons and glia. Fibroblast growth factor-2 (FGF-2), a major signal that maintains neural progenitors in the undifferentiated state, is also thought to influence the transition from neurogenesis to gliogenesis. Here we present evidence that FGF receptors and underlying signaling pathways transmit the FGF-2 signals that regulate astrocyte specification aside from its mitogenic activity. Application of FGF-2 to cortical progenitors suppressed neurogenesis whereas treatment with an FGFR antagonist in vitro promoted neurogenesis. Introduction of chimeric FGFRs with mutated tyrosine residues into cortical progenitors and drug treatments to specifically block individual downstream signaling pathways revealed that the overall activity of FGFR rather than individual autophosphorylation sites is important for delivering signals for glial specification. In contrast, a signal for cell proliferation by FGFR was mainly delivered by MAPK pathway. Together our findings indicate that FGFR activity promotes astrocyte specification in the developing CNS.

Diverse FGF receptor signaling controls astrocyte specification and proliferation

International Nuclear Information System (INIS)

Kang, Kyungjun; Song, Mi-Ryoung

2010-01-01

During CNS development, pluripotency neuronal progenitor cells give rise in succession to neurons and glia. Fibroblast growth factor-2 (FGF-2), a major signal that maintains neural progenitors in the undifferentiated state, is also thought to influence the transition from neurogenesis to gliogenesis. Here we present evidence that FGF receptors and underlying signaling pathways transmit the FGF-2 signals that regulate astrocyte specification aside from its mitogenic activity. Application of FGF-2 to cortical progenitors suppressed neurogenesis whereas treatment with an FGFR antagonist in vitro promoted neurogenesis. Introduction of chimeric FGFRs with mutated tyrosine residues into cortical progenitors and drug treatments to specifically block individual downstream signaling pathways revealed that the overall activity of FGFR rather than individual autophosphorylation sites is important for delivering signals for glial specification. In contrast, a signal for cell proliferation by FGFR was mainly delivered by MAPK pathway. Together our findings indicate that FGFR activity promotes astrocyte specification in the developing CNS.

FGFR3 expression in primary and metastatic urothelial carcinoma of the bladder

International Nuclear Information System (INIS)

Guancial, Elizabeth A; Werner, Lillian; Bellmunt, Joaquim; Bamias, Aristotle; Choueiri, Toni K; Ross, Robert; Schutz, Fabio A; Park, Rachel S; O'Brien, Robert J; Hirsch, Michelle S; Barletta, Justine A; Berman, David M; Lis, Rosina; Loda, Massimo; Stack, Edward C; Garraway, Levi A; Riester, Markus; Michor, Franziska; Kantoff, Philip W; Rosenberg, Jonathan E

2014-01-01

While fibroblast growth factor receptor 3 (FGFR3) is frequently mutated or overexpressed in nonmuscle-invasive urothelial carcinoma (UC), the prevalence of FGFR3 protein expression and mutation remains unknown in muscle-invasive disease. FGFR3 protein and mRNA expression, mutational status, and copy number variation were retrospectively analyzed in 231 patients with formalin-fixed paraffin-embedded primary UCs, 33 metastases, and 14 paired primary and metastatic tumors using the following methods: immunohistochemistry, NanoString nCounterTM, OncoMap or Affymetrix OncoScanTM array, and Gain and Loss of Analysis of DNA and Genomic Identification of Significant Targets in Cancer software. FGFR3 immunohistochemistry staining was present in 29% of primary UCs and 49% of metastases and did not impact overall survival (P = 0.89, primary tumors; P = 0.78, metastases). FGFR3 mutations were observed in 2% of primary tumors and 9% of metastases. Mutant tumors expressed higher levels of FGFR3 mRNA than wild-type tumors (P < 0.001). FGFR3 copy number gain and loss were rare events in primary and metastatic tumors (0.8% each; 3.0% and 12.3%, respectively). FGFR3 immunohistochemistry staining is present in one third of primary muscle-invasive UCs and half of metastases, while FGFR3 mutations and copy number changes are relatively uncommon

Fibroblast Growth Factor signaling regulates the expansion of A6-expressing hepatocytes in association with AKT-dependent β-catenin activation

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Utley, Sarah; James, David; Mavila, Nirmala; Nguyen, Marie V.; Vendryes, Christopher; Salisbury, S. Michael; Phan, Jennifer; Wang, Kasper S.

2014-01-01

Background & Aims Fibroblast Growth Factors (FGFs) promote the proliferation and survival of hepatic progenitor cells (HPCs) via AKT-dependent β-catenin activation. Moreover, the emergence of hepatocytes expressing the HPC marker A6 during 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC)-induced liver injury is mediated partly by FGF and β-catenin signaling. Herein, we investigate the role of FGF signaling and AKT-mediated β-catenin activation in acute DDC liver injury. Methods Transgenic mice were fed DDC chow for 14 days concurrent with either Fgf10 over-expression or inhibition of FGF signaling via expression of soluble dominant-negative FGF Receptor (R)-2IIIb. Results After 14 days of DDC treatment, there was an increase in periportal cells expressing FGFR1, FGFR2, and AKT-activated phospho-Serine 552 (pSer552) β-CATENIN in association with up-regulation of genes encoding FGFR2IIIb ligands, Fgf7, Fgf10, and Fgf22. In response to Fgf10 over-expression, there was an increase in the number of pSer552-β-CATENIN(positive)+ive periportal cells as well as cells co-positive for A6 and hepatocyte marker, Hepatocyte Nuclear Factor-4α (HNF4α). A similar expansion of A6+ive cells was observed after Fgf10 over-expression with regular chow and after partial hepatectomy during ethanol toxicity. Inhibition of FGF signaling increased the periportal A6+iveHNF4α+ive cell population while reducing centrolobular A6+ive HNF4α+ive cells. AKT inhibition with Wortmannin attenuated FGF10-mediated A6+iveHNF4α+ive cell expansion. In vitro analyses using FGF10 treated HepG2 cells demonstrated AKT-mediated β-CATENIN activation but not enhanced cell migration. Conclusion During acute DDC treatment, FGF signaling promotes the expansion of A6-expressing liver cells partly via AKT-dependent activation of β-CATENIN expansion of A6+ive periportal cells and possibly by reprogramming of centrolobular hepatocytes. PMID:24365171

FGFR2 promotes breast tumorigenicity through maintenance of breast tumor-initiating cells.

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Sungeun Kim

Full Text Available Emerging evidence suggests that some cancers contain a population of stem-like TICs (tumor-initiating cells and eliminating TICs may offer a new strategy to develop successful anti-cancer therapies. As molecular mechanisms underlying the maintenance of the TIC pool are poorly understood, the development of TIC-specific therapeutics remains a major challenge. We first identified and characterized TICs and non-TICs isolated from a mouse breast cancer model. TICs displayed increased tumorigenic potential, self-renewal, heterogeneous differentiation, and bipotency. Gene expression analysis and immunostaining of TICs and non-TICs revealed that FGFR2 was preferentially expressed in TICs. Loss of FGFR2 impaired self-renewal of TICs, thus resulting in marked decreases in the TIC population and tumorigenic potential. Restoration of FGFR2 rescued the defects in TIC pool maintenance, bipotency, and breast tumor growth driven by FGFR2 knockdown. In addition, pharmacological inhibition of FGFR2 kinase activity led to a decrease in the TIC population which resulted in suppression of breast tumor growth. Moreover, human breast TICs isolated from patient tumor samples were found enriched in a FGFR2+ population that was sufficient to initiate tumor growth. Our data suggest that FGFR2 is essential in sustaining the breast TIC pool through promotion of self-renewal and maintenance of bipotent TICs, and raise the possibility of FGFR2 inhibition as a strategy for anti-cancer therapy by eradicating breast TICs.

FGFR4 Downregulation of Cell Adhesion in Prostate Cancer

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2008-09-01

in Figure 1, all constructs were stably incorporated into 293-RXR cells and were inducible upon treatment with Ponasterone A. Though we had created...through the transmembrane domain, similar to the FGFR3 Gly380Arg mutation responsible for human dwarfism , or achondroplasia. In this model, the FGFR4

A novel non-invasive detection method for the FGFR3 gene mutation in maternal plasma for a fetal achondroplasia diagnosis based on signal amplification by hemin-MOFs/PtNPs.

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Chen, Jun; Yu, Chao; Zhao, Yilin; Niu, Yazhen; Zhang, Lei; Yu, Yujie; Wu, Jing; He, Junlin

2017-05-15

The small amount of cell-free fetal DNA (cffDNA) can be a useful biomarker for early non-invasive prenatal diagnosis (NIPD) of achondroplasia. In this study, a novel non-invasive electrochemical DNA sensor for ultrasensitive detecting FGFR3 mutation gene, a pathogenic gene of achondroplasia, based on biocatalytic signal materials and the biotin-streptavidin system are presented. Notably encapsulation of hemin in metal-organic frameworks-based materials (hemin-MOFs) and platinum nanoparticles (PtNPs) were used to prepare hemin-MOFs/PtNPs composites via a one-beaker-one-step reduction. We utilized hemin-MOFs/PtNPs for signal amplification because the promising hemin-MOFs/PtNPs nanomaterial has remarkable ability of catalyze H 2 O 2 as well as excellent conductivity. To further amplify the electrochemical signal, reduced graphene oxide-tetraethylene pentamine (rGO-TEPA), gold nanoparticles and streptavidin were selected for modification of the electrode to enhance the conductivity and immobilize more biotin-modified capture probe (Bio-CP) through the high specificity and superior affinity between streptavidin and biotin. The electrochemical signal was primarily derived from the synergistic catalysis of H 2 O 2 by hemin and PtNPs and recorded by Chronoamperometry. Under the optimal conditions, this newly designed biosensor exhibited sensitive detection of FGFR3 from 0.1fM to 1nM with a low detection limit of 0.033fM (S/N=3). We proposed that this ultrasensitive biosensor is useful for the early non-invasive prenatal diagnosis of achondroplasia. Copyright © 2016 Elsevier B.V. All rights reserved.

Fibroblast growth factor receptor (FGFR) alterations in squamous differentiated bladder cancer: a putative therapeutic target for a small subgroup.

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Baldia, Philipp H; Maurer, Angela; Heide, Timon; Rose, Michael; Stoehr, Robert; Hartmann, Arndt; Williams, Sarah V; Knowles, Margaret A; Knuechel, Ruth; Gaisa, Nadine T

2016-11-01

Although drugable fibroblast growth factor receptor (FGFR) alterations in squamous cell carcinomas (SCC) of various entities are well known, little is known about FGFR modifications in squamous differentiated bladder cancer. Therefore, our study evaluated FGFR1-3 alterations as a putative therapeutic target in this subgroup. We analyzed 73 squamous differentiated bladder cancers (n = 10 pT2, n = 55 pT3, n = 8 pT4) for FGFR1-3 protein expression, FGFR1-3 copy number variations, FGFR3 chromosomal rearrangements (fluorescence in situ hybridization (FISH)) and FGFR3 mutations (SNapShot analysis). Only single cases displayed enhanced protein expression, most frequently FGFR3 overexpression (9.4% (6/64)). FISH showed no amplifications of FGFR1, 2 or 3. Break apart events were only slightly above the cut off in 12.1% (8/66) of cases and no FGFR3-TACC3 rearrangements could be proven by qPCR. FGFR3 mutations (p.S249C) were found in 8.5% (6/71) of tumors and were significantly associated with FGFR3 protein overexpression (p bladder cancer (n = 85), which revealed reduced overall expression of FGFR1 and FGFR2 in tumors compared to normal tissue, while expression of FGFR3 remained high. In the TCGA "squamous-like" subtype FGFR3 mutations were found in 4.9% and correlated with high FGFR3 RNA expression. Mutations of FGFR1 and FGFR2 were less frequent (2.4% and 1.2%). Hence, our comprehensive study provides novel insights into a subgroup of squamous differentiated bladder tumors that hold clues for novel therapeutic regimens and may benefit from FGFR3-targeted therapies.

Insight into resistance mechanisms of AZD4547 and E3810 to FGFR1 gatekeeper mutation via theoretical study

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Liang D

2017-02-01

Full Text Available Donglou Liang,1,* Qiaowan Chen,2,* Yujin Guo,1 Ting Zhang,3 Wentao Guo4 1Pharmacy Department, Jining First People’s Hospital, 2Department of Obstetrics, Affiliated Hospital of Jining Medical University, Jining, Shandong, 3Department of Rheumatology, The First Affiliated Hospital of Wenzhou Medical University, 4School of Pharmacy, Wenzhou Medical University, Wenzhou, Zhejiang, People’s Republic of China *These authors contributed equally to this work Abstract: Inhibitors targeting the amplification of the fibroblast growth factor receptor 1 (FGFR1 have found success in the treatment of FGFR1-positive squamous cell lung and breast cancers. A secondary mutation of gatekeeper residue (V561M in the binding site has been linked to the acquired resistance. Recently, two well-known small molecule inhibitors of FGFR1, AZD4547 and E3810, reported that the V561M mutation confers significant resistance to E3810, while retaining affinity for AZD4547. FGFR1 is widely investigated as potential therapeutic target, while there are few computational studies made to understand the resistance mechanisms about FGFR1 V561M gatekeeper mutation. In this study, molecular docking, classical molecular dynamics simulations, molecular mechanics/generalized born surface area (MM/GBSA free energy calculations, and umbrella sampling (US simulations were carried out to make clear the principle of the binding preference of AZD4547 and E3810 toward FGFR1 V561M gatekeeper mutation. The results provided by MM/GBSA reveal that AZD4547 has similar binding affinity to both FGFR1WT and FGFR1V561M, whereas E3810 has much higher binding affinity to FGFR1WT than to FGFR1V561M. Comparison of individual energy terms indicates that the major variation of E3810 between FGFR1WT and FGFR1V561M are van der Waals interactions. In addition, US simulations prove that the potential of mean force (PMF profile of AZD4547 toward FGFR1WT and FGFR1V561M has similar PMF depth. However, the PMF profile

Recombinant expression in E. coli of human FGFR2 with its transmembrane and extracellular domains

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Adam Bajinting

2017-06-01

Full Text Available Fibroblast growth factor receptors (FGFRs are a family of receptor tyrosine kinases containing three domains: an extracellular receptor domain, a single transmembrane helix, and an intracellular tyrosine kinase domain. FGFRs are activated by fibroblast growth factors (FGFs as part of complex signal transduction cascades regulating angiogenesis, skeletal formation, cell differentiation, proliferation, cell survival, and cancer. We have developed the first recombinant expression system in E. coli to produce a construct of human FGFR2 containing its transmembrane and extracellular receptor domains. We demonstrate that the expressed construct is functional in binding heparin and dimerizing. Size exclusion chromatography demonstrates that the purified FGFR2 does not form a complex with FGF1 or adopts an inactive dimer conformation. Progress towards the successful recombinant production of intact FGFRs will facilitate further biochemical experiments and structure determination that will provide insight into how extracellular FGF binding activates intracellular kinase activity.

FGFR3, PIK3CA and RAS mutations in benign lichenoid keratosis.

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Groesser, L; Herschberger, E; Landthaler, M; Hafner, C

2012-04-01

Benign lichenoid keratoses (BLKs) are solitary skin lesions which have been proposed to represent a regressive form of pre-existent epidermal tumours such as solar lentigo or seborrhoeic keratosis. However, the genetic basis of BLK is unknown. FGFR3, PIK3CA and RAS mutations have been shown to be involved in the pathogenesis of seborrhoeic keratosis and solar lentigo. We thus investigated whether these mutations are also present in BLK. After manual microdissection and DNA isolation, 52 BLKs were screened for FGFR3, PIK3CA and RAS hotspot mutations using SNaPshot(®) multiplex assays. We identified 6/52 (12%) FGFR3 mutations, 10/52 (19%) PIK3CA mutations, 6/52 (12%) HRAS mutations and 2/52 (4%) KRAS mutations. FGFR3 and RAS mutations were mutually exclusive. One BLK showed a simultaneous PIK3CA and HRAS mutation. In nine BLKs with a mutation, nonlesional control tissue from the epidermal margin and the dermal lymphocytic infiltrate were wild-type, indicating that these mutations are somatic. To demonstrate that these findings are specific, 10 samples of lichen planus were analysed without evidence for FGFR3, PIK3CA or RAS mutations. Our results indicate that FGFR3, PIK3CA and RAS mutations are present in approximately 50% of BLKs. These findings support the concept on the molecular genetic level that at least a proportion of BLKs represents regressive variants resulting from former benign epidermal tumours such as seborrhoeic keratosis and solar lentigo. © 2011 The Authors. BJD © 2011 British Association of Dermatologists 2011.

Fibroblast growth factor and canonical WNT/beta-catenin signaling cooperate in suppression of chondrocyte differentiation in experimental models of FGFR signaling in cartilage

Czech Academy of Sciences Publication Activity Database

Buchtová, Marcela; Oralová, Veronika; Aklian, A.; Mašek, J.; Veselá, I.; Ouyang, Z.; Obadalová, T.; Konečná, Ž.; Spoustová, T.; Pospíšilová, T.; Matula, P.; Vařecha, M.; Balek, L.; Gudernová, I.; Jelínková, I.; Ďuran, I.; Červenková, I.; Murakami, S.; Kozubík, Alois; Dvořák, P.; Bryja, Vítězslav; Krejčí, P.

2015-01-01

Roč. 1852, č. 5 (2015), s. 839-850 ISSN 0925-4439 R&D Projects: GA ČR GCP302/12/J059; GA ČR GBP302/12/G157; GA ČR(CZ) GA14-31540S Institutional support: RVO:67985904 ; RVO:68081707 Keywords : fibroblast growth factor receptor * FGFR3 * WNT Subject RIV: EA - Cell Biology Impact factor: 5.158, year: 2015

Expression of FGFR3 Protein and Gene Amplification in Urinary Bladder Lesions in Relation to Schistosomiasis

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Olfat Hammam

2017-04-01

CONCLUSIONS: FGFR3 overexpression in malignant cases was significantly higher than in chronic cystitis. FGFR3 gene amplification was reported mainly in low grade and NNMBIC tumours. FGFR3 may be further studied as a subject for target therapy of bladder cancer.

Effect of the G375C and G346E achondroplasia mutations on FGFR3 activation.

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Lijuan He

Full Text Available Two mutations in FGFR3, G380R and G375C are known to cause achondroplasia, the most common form of human dwarfism. The G380R mutation accounts for 98% of the achondroplasia cases, and thus has been studied extensively. Here we study the effect of the G375C mutation on the phosphorylation and the cross-linking propensity of full-length FGFR3 in HEK 293 cells, and we compare the results to previously published results for the G380R mutant. We observe identical behavior of the two achondroplasia mutants in these experiments, a finding which supports a direct link between the severity of dwarfism phenotypes and the level and mechanism of FGFR3 over-activation. The mutations do not increase the cross-linking propensity of FGFR3, contrary to previous expectations that the achondroplasia mutations stabilize the FGFR3 dimers. Instead, the phosphorylation efficiency within un-liganded FGFR3 dimers is increased, and this increase is likely the underlying cause for pathogenesis in achondroplasia. We further investigate the G346E mutation, which has been reported to cause achondroplasia in one case. We find that this mutation does not increase FGFR3 phosphorylation and decreases FGFR3 cross-linking propensity, a finding which raises questions whether this mutation is indeed a genetic cause for human dwarfism.

Effect of the G375C and G346E achondroplasia mutations on FGFR3 activation.

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He, Lijuan; Serrano, Christopher; Niphadkar, Nitish; Shobnam, Nadia; Hristova, Kalina

2012-01-01

Two mutations in FGFR3, G380R and G375C are known to cause achondroplasia, the most common form of human dwarfism. The G380R mutation accounts for 98% of the achondroplasia cases, and thus has been studied extensively. Here we study the effect of the G375C mutation on the phosphorylation and the cross-linking propensity of full-length FGFR3 in HEK 293 cells, and we compare the results to previously published results for the G380R mutant. We observe identical behavior of the two achondroplasia mutants in these experiments, a finding which supports a direct link between the severity of dwarfism phenotypes and the level and mechanism of FGFR3 over-activation. The mutations do not increase the cross-linking propensity of FGFR3, contrary to previous expectations that the achondroplasia mutations stabilize the FGFR3 dimers. Instead, the phosphorylation efficiency within un-liganded FGFR3 dimers is increased, and this increase is likely the underlying cause for pathogenesis in achondroplasia. We further investigate the G346E mutation, which has been reported to cause achondroplasia in one case. We find that this mutation does not increase FGFR3 phosphorylation and decreases FGFR3 cross-linking propensity, a finding which raises questions whether this mutation is indeed a genetic cause for human dwarfism.

Global Developmental Gene Programing Involves a Nuclear Form of Fibroblast Growth Factor Receptor-1 (FGFR1.

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Christopher Terranova

Full Text Available Genetic studies have placed the Fgfr1 gene at the top of major ontogenic pathways that enable gastrulation, tissue development and organogenesis. Using genome-wide sequencing and loss and gain of function experiments the present investigation reveals a mechanism that underlies global and direct gene regulation by the nuclear form of FGFR1, ensuring that pluripotent Embryonic Stem Cells differentiate into Neuronal Cells in response to Retinoic Acid. Nuclear FGFR1, both alone and with its partner nuclear receptors RXR and Nur77, targets thousands of active genes and controls the expression of pluripotency, homeobox, neuronal and mesodermal genes. Nuclear FGFR1 targets genes in developmental pathways represented by Wnt/β-catenin, CREB, BMP, the cell cycle and cancer-related TP53 pathway, neuroectodermal and mesodermal programing networks, axonal growth and synaptic plasticity pathways. Nuclear FGFR1 targets the consensus sequences of transcription factors known to engage CREB-binding protein, a common coregulator of transcription and established binding partner of nuclear FGFR1. This investigation reveals the role of nuclear FGFR1 as a global genomic programmer of cell, neural and muscle development.

Prognostic value of FGFR gene amplification in patients with different types of cancer: a systematic review and meta-analysis.

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Jinjia Chang

Full Text Available BACKGROUND: Fibroblast growth factor receptor (FGFR gene amplification has been reported in different types of cancer. We performed an up-to-date meta-analysis to further characterize the prognostic value of FGFR gene amplification in patients with cancer. METHODS: A search of several databases, including MEDLINE (PubMed, EMBASE, Web of Science, and China National Knowledge Infrastructure, was conducted to identify studies examining the association between FGFR gene amplification and cancer. A total of 24 studies met the inclusion criteria, and overall incidence rates, hazard risk (HR, overall survival, disease-free survival, and 95% confidence intervals (CIs were calculated employing fixed- or random-effects models depending on the heterogeneity of the included studies. RESULTS: In the meta-analysis of 24 studies, the prevalence of FGFR gene amplification was FGFR1: 0.11 (95% CI: 0.08-0.13 and FGFR2: 0.04 (95% CI: 0.02-0.06. Overall survival was significantly worse among patients with FGFR gene amplification: FGFR1 [HR 1.57 (95% CI: 1.23-1.99; p = 0.0002] and FGFR2 [HR 2.27 (95% CI: 1.73-3.00; p<0.00001]. CONCLUSIONS: Current evidence supports the conclusion that the outcomes of patients with FGFR gene amplified cancers is worse than for those with non-FGFR gene amplified cancers.

FGFR3 and P53 characterize alternative genetic pathways in the pathogenesis of urothelial cell carcinoma

NARCIS (Netherlands)

B.W. van Rhijn (Bas); Th.H. van der Kwast (Theo); A.N. Vis (André); W.J. Kirkels (Wim); E.R. Boeve; A.C. Jobsis; E.C. Zwarthoff (Ellen)

2004-01-01

textabstractFibroblast growth factor receptor 3 (FGFR3) and P53 mutations are frequently observed in bladder cancer. We here describe the distribution of FGFR3 mutations and P53 overexpression in 260 primary urothelial cell carcinomas. FGFR3 mutations were observed in 59% and P53

Germline variant FGFR4  p.G388R exposes a membrane-proximal STAT3 binding site.

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Ulaganathan, Vijay K; Sperl, Bianca; Rapp, Ulf R; Ullrich, Axel

2015-12-24

Variant rs351855-G/A is a commonly occurring single-nucleotide polymorphism of coding regions in exon 9 of the fibroblast growth factor receptor FGFR4 (CD334) gene (c.1162G>A). It results in an amino-acid change at codon 388 from glycine to arginine (p.Gly388Arg) in the transmembrane domain of the receptor. Despite compelling genetic evidence for the association of this common variant with cancers of the bone, breast, colon, prostate, skin, lung, head and neck, as well as soft-tissue sarcomas and non-Hodgkin lymphoma, the underlying biological mechanism has remained elusive. Here we show that substitution of the conserved glycine 388 residue to a charged arginine residue alters the transmembrane spanning segment and exposes a membrane-proximal cytoplasmic signal transducer and activator of transcription 3 (STAT3) binding site Y(390)-(P)XXQ(393). We demonstrate that such membrane-proximal STAT3 binding motifs in the germline of type I membrane receptors enhance STAT3 tyrosine phosphorylation by recruiting STAT3 proteins to the inner cell membrane. Remarkably, such germline variants frequently co-localize with somatic mutations in the Catalogue of Somatic Mutations in Cancer (COSMIC) database. Using Fgfr4 single nucleotide polymorphism knock-in mice and transgenic mouse models for breast and lung cancers, we validate the enhanced STAT3 signalling induced by the FGFR4 Arg388-variant in vivo. Thus, our findings elucidate the molecular mechanism behind the genetic association of rs351855 with accelerated cancer progression and suggest that germline variants of cell-surface molecules that recruit STAT3 to the inner cell membrane are a significant risk for cancer prognosis and disease progression.

Identification and in silico characterization of p.G380R substitution in FGFR3, associated with achondroplasia in a non-consanguineous Pakistani family.

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Ajmal, Muhammad; Mir, Asif; Shoaib, Muhammad; Malik, Salman Akbar; Nasir, Muhammad

2017-07-05

The dimerization efficiency of FGFR3 transmembrane domain plays a critical role in the formation of a normal skeleton through the negative regulation of bone development. Recently, gain-of-function mutations in the transmembrane domain of FGFR3 has been described associated with an aberrant negative regulation, leading to the development of achondroplasia-group disorders, including achondroplasia (ACH), hypochondroplasia (HCH) and thanatophoric dysplasia (TD). Here, we describe a non-consanguineous Pakistani family with achondroplasia to explain hereditary basis of the disease. PCR-based linkage analysis using microsatellite markers was employed to localize the disease gene. Gene specific intronic primers were used to amplify the genomic DNA from all affected as well as phenotypically healthy individuals. Amplified PCR products were then subjected to Sanger sequencing and RFLP analysis to identify a potentially pathogenic mutation. The impact of identified mutation on FGFR3 protein's structure and stability was highlighted through different bioinformatics tools. Genetic screening of the family revealed a previously reported heterozygous c.1138 G > A (p.G380R) mutation in the coding exon 8 of FGFR3 gene. Identified genetic variation was confirmed in all affected individuals while healthy individuals and controls were found genotypically normal. The results were further validated by RFLP analysis as c.1138 G > A substitution generates a unique recognition site for SfcI endonuclease. Following SfcI digestion, the electrophoretic pattern of three bands/DNA fragments for each patient is indicative of heterozygous status of the disease allele. In silico studies of the mutant FGFR3 protein predicted to adversely affect the stability of FGFR3 protein. Mutation in the transmembrane domain may adversely affect the dimerization efficiency and overall stability of the FGFR3, leading to a constitutively active protein. As a result, an uncontrolled intracellular signaling

Differential regulation of renal Klotho and FGFR1 in normal and uremic rats.

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Muñoz-Castañeda, Juan R; Herencia, Carmen; Pendón-Ruiz de Mier, Maria Victoria; Rodriguez-Ortiz, Maria Encarnación; Diaz-Tocados, Juan M; Vergara, Noemi; Martínez-Moreno, Julio M; Salmerón, Maria Dolores; Richards, William G; Felsenfeld, Arnold; Kuro-O, Makoto; Almadén, Yolanda; Rodríguez, Mariano

2017-09-01

In renal failure, hyperphosphatemia occurs despite a marked elevation in serum fibroblast growth factor (FGF)-23. Abnormal regulation of the FGFR1-Klotho receptor complex may cause a resistance to the phosphaturic action of FGF23. The purpose of the present study was to investigate the regulation of renal Klotho and FGF receptor (FEFR)-1 in healthy and uremic rats induced by 5/6 nephrectomy. In normal rats, the infusion of rat recombinant FGF23 enhanced phosphaturia and increased renal FGFR1 expression; however, Klotho expression was reduced. Uremic rats on a high-phosphate (HP) diet presented hyperphosphatemia with marked elevation of FGF23 and an increased fractional excretion of phosphate (P) that was associated with a marked reduction of Klotho expression and an increase in FGFR1. After neutralization of FGF23 by anti-FGF23 administration, phosphaturia was still abundant, Klotho expression remained low, and the FGFR1 level was reduced. These results suggest that the expression of renal Klotho is modulated by phosphaturia, whereas the FGFR1 expression is regulated by FGF23. Calcitriol (CTR) administration prevented a decrease in renal Klotho expression. In HEK293 cells HP produced nuclear translocation of β-catenin, together with a reduction in Klotho. Wnt/β-catenin inhibition with Dkk-1 prevented the P-induced down-regulation of Klotho. The addition of CTR to HP medium was able to recover Klotho expression. In summary, high FGF23 levels increase FGFR1, whereas phosphaturia decreases Klotho expression through the activation of Wnt/β-catenin pathway.-Muñoz-Castañeda, J. R., Herencia, C., Pendón-Ruiz de Mier, M. V., Rodriguez-Ortiz, M. E., Diaz-Tocados, J. M., Vergara, N., Martínez-Moreno, J. M., Salmerón, M. D., Richards, W. G., Felsenfeld, A., Kuro-O, M., Almadén, Y., Rodríguez, M. Differential regulation of renal Klotho and FGFR1 in normal and uremic rats. © FASEB.

Comparative functional analysis of two fibroblast growth factor receptor 1 (FGFR1) mutations affecting the same residue (R254W and R254Q) in isolated hypogonadotropic hypogonadism (IHH).

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Koika, Vasiliki; Varnavas, Petros; Valavani, Helen; Sidis, Yisrael; Plummer, Lacey; Dwyer, Andrew; Quinton, Richard; Kanaka-Gantenbein, Christine; Pitteloud, Nelly; Sertedaki, Amalia; Dacou-Voutetakis, Catherine; Georgopoulos, Neoklis A

2013-03-01

FGFR1 mutations have been identified in both Kallmann syndrome and normosmic HH (nIHH). To date, few mutations in the FGFR1 gene have been structurally or functionally characterized in vitro to identify molecular mechanisms that contribute to the disease pathogenesis. We attempted to define the in vitro functionality of two FGFR1 mutants (R254W and R254Q), resulting from two different amino acid substitutions of the same residue, and to correlate the in vitro findings to the patient phenotypes. Two unrelated GnRH deficient probands were found to harbor mutations in FGFR1 (R254W and R254Q). Mutant signaling activity and expression levels were evaluated in vitro and compared to a wild type (WT) receptor. Signaling activity was determined by a FGF2/FGFR1 dependent transcription reporter assay. Receptor total expression levels were assessed by Western blot and cell surface expression was measured by a radiolabeled antibody binding assay. The R254W maximal receptor signaling capacity was reduced by 45% (p<0.01) while R254Q activity was not different from WT. However, both mutants displayed diminished total protein expression levels (40 and 30% reduction relative to WT, respectively), while protein maturation was unaffected. Accordingly, cell surface expression levels of the mutant receptors were also significantly reduced (35% p<0.01 and 15% p<0.05, respectively). The p.R254W and p.R254Q are both loss-of-function mutations as demonstrated by their reduced overall and cell surface expression levels suggesting a deleterious effect on receptor folding and stability. It appears that a tryptophan substitution at R254 is more disruptive to receptor structure than the more conserved glutamine substitution. No clear correlation between the severity of in vitro loss-of-function and phenotypic presentation could be assigned. Copyright © 2012 Elsevier B.V. All rights reserved.

HDAC6 deficiency or inhibition blocks FGFR3 accumulation and improves bone growth in a model of achondroplasia.

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Ota, Sara; Zhou, Zi-Qiang; Romero, Megan P; Yang, Guang; Hurlin, Peter J

2016-10-01

Mutations that cause increased and/or inappropriate activation of FGFR3 are responsible for a collection of short-limbed chondrodysplasias. These mutations can alter receptor trafficking and enhance receptor stability, leading to increased receptor accumulation and activity. Here, we show that wildtype and mutant activated forms of FGFR3 increase expression of the cytoplasmic deacetylase HDAC6 (Histone Deacetylase 6) and that FGFR3 accumulation is compromised in cells lacking HDAC6 or following treatment of fibroblasts or chondrocytes with small molecule inhibitors of HDAC6. The reduced accumulation of FGFR3 was linked to increased FGFR3 degradation that occurred through a lysosome-dependent mechanism. Using a mouse model of Thanatophoric Dysplasia Type II (TDII) we show that both HDAC6 deletion and treatment with the small molecule HDAC6 inhibitor tubacin reduced FGFR3 accumulation in the growth plate and improved endochondral bone growth. Defective endochondral growth in TDII is associated with reduced proliferation and poor hypertrophic differentiation and the improved bone growth was associated with increased chondrocyte proliferation and expansion of the differentiation compartment within the growth plate. These findings further define the mechanisms that control FGFR3 accumulation and contribute to skeletal pathology caused by mutations in FGFR3. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

A meta-analysis of the relationship between FGFR3 and TP53 mutations in bladder cancer.

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Neuzillet, Yann; Paoletti, Xavier; Ouerhani, Slah; Mongiat-Artus, Pierre; Soliman, Hany; de The, Hugues; Sibony, Mathilde; Denoux, Yves; Molinie, Vincent; Herault, Aurélie; Lepage, May-Linda; Maille, Pascale; Renou, Audrey; Vordos, Dimitri; Abbou, Claude-Clément; Bakkar, Ashraf; Asselain, Bernard; Kourda, Nadia; El Gaaied, Amel; Leroy, Karen; Laplanche, Agnès; Benhamou, Simone; Lebret, Thierry; Allory, Yves; Radvanyi, François

2012-01-01

TP53 and FGFR3 mutations are the most common mutations in bladder cancers. FGFR3 mutations are most frequent in low-grade low-stage tumours, whereas TP53 mutations are most frequent in high-grade high-stage tumours. Several studies have reported FGFR3 and TP53 mutations to be mutually exclusive events, whereas others have reported them to be independent. We carried out a meta-analysis of published findings for FGFR3 and TP53 mutations in bladder cancer (535 tumours, 6 publications) and additional unpublished data for 382 tumours. TP53 and FGFR3 mutations were not independent events for all tumours considered together (OR = 0.25 [0.18-0.37], p = 0.0001) or for pT1 tumours alone (OR = 0.47 [0.28-0.79], p = 0.0009). However, if the analysis was restricted to pTa tumours or to muscle-invasive tumours alone, FGFR3 and TP53 mutations were independent events (OR = 0.56 [0.23-1.36] (p = 0.12) and OR = 0.99 [0.37-2.7] (p = 0.35), respectively). After stratification of the tumours by stage and grade, no dependence was detected in the five tumour groups considered (pTaG1 and pTaG2 together, pTaG3, pT1G2, pT1G3, pT2-4). These differences in findings can be attributed to the putative existence of two different pathways of tumour progression in bladder cancer: the CIS pathway, in which FGFR3 mutations are rare, and the Ta pathway, in which FGFR3 mutations are frequent. TP53 mutations occur at the earliest stage of the CIS pathway, whereas they occur would much later in the Ta pathway, at the T1G3 or muscle-invasive stage.

FGF signalling controls the specification of hair placode-derived SOX9 positive progenitors to Merkel cells.

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Nguyen, Minh Binh; Cohen, Idan; Kumar, Vinod; Xu, Zijian; Bar, Carmit; Dauber-Decker, Katherine L; Tsai, Pai-Chi; Marangoni, Pauline; Klein, Ophir D; Hsu, Ya-Chieh; Chen, Ting; Mikkola, Marja L; Ezhkova, Elena

2018-06-13

Merkel cells are innervated mechanosensory cells responsible for light-touch sensations. In murine dorsal skin, Merkel cells are located in touch domes and found in the epidermis around primary hairs. While it has been shown that Merkel cells are skin epithelial cells, the progenitor cell population that gives rise to these cells is unknown. Here, we show that during embryogenesis, SOX9-positive (+) cells inside hair follicles, which were previously known to give rise to hair follicle stem cells (HFSCs) and cells of the hair follicle lineage, can also give rise to Merkel Cells. Interestingly, while SOX9 is critical for HFSC specification, it is dispensable for Merkel cell formation. Conversely, FGFR2 is required for Merkel cell formation but is dispensable for HFSCs. Together, our studies uncover SOX9(+) cells as precursors of Merkel cells and show the requirement for FGFR2-mediated epithelial signalling in Merkel cell specification.

Direct Assessment of the Effect of the Gly380Arg Achondroplasia Mutation on FGFR3 Dimerization Using Quantitative Imaging FRET

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Placone, Jesse; Hristova, Kalina

2012-01-01

The Gly380Arg mutation in FGFR3 is the genetic cause for achondroplasia (ACH), the most common form of human dwarfism. The mutation has been proposed to increase FGFR3 dimerization, but the dimerization propensities of wild-type and mutant FGFR3 have not been compared. Here we use quantitative imaging FRET to characterize the dimerization of wild-type FGFR3 and the ACH mutant in plasma membrane-derived vesicles from HEK293T cells. We demonstrate a small, but statistically significant increase in FGFR3 dimerization due to the ACH mutation. The data are consistent with the idea that the ACH mutation causes a structural change which affects both the stability and the activity of FGFR3 dimers in the absence of ligand. PMID:23056398

FGFR3 regulates brain size by controlling progenitor cell proliferation and apoptosis during embryonic development.

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Inglis-Broadgate, Suzanne L; Thomson, Rachel E; Pellicano, Francesca; Tartaglia, Michael A; Pontikis, Charlie C; Cooper, Jonathan D; Iwata, Tomoko

2005-03-01

Mice with the K644E kinase domain mutation in fibroblast growth factor receptor 3 (Fgfr3) (EIIa;Fgfr3(+/K644E)) exhibited a marked enlargement of the brain. The brain size was increased as early as E11.5, not secondary to the possible effect of Fgfr3 activity in the skeleton. Furthermore, the mutant brains showed a dramatic increase in cortical thickness, a phenotype opposite to that in FGF2 knockout mice. Despite this increased thickness, cortical layer formation was largely unaffected and no cortical folding was observed during embryonic days 11.5-18.5 (E11.5-E18.5). Measurement of cortical thickness revealed an increase of 38.1% in the EIIa;Fgfr3(+/K644E) mice at E14.5 and the advanced appearance of the cortical plate was frequently observed at this stage. Unbiased stereological analysis revealed that the volume of the ventricular zone (VZ) was increased by more than two fold in the EIIa;Fgfr3(+/K644E) mutants at E14.5. A relatively mild increase in progenitor cell proliferation and a profound decrease in developmental apoptosis during E11.5-E14.5 most likely accounts for the dramatic increase in total telecephalic cell number. Taken together, our data suggest a novel function of Fgfr3 in controlling the development of the cortex, by regulating proliferation and apoptosis of cortical progenitors.

Mild achondroplasia/hypochondroplasia with acanthosis nigricans, normal development, and a p.Ser348Cys FGFR3 mutation.

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Couser, Natario L; Pande, Chetna K; Turcott, Christie M; Spector, Elaine B; Aylsworth, Arthur S; Powell, Cynthia M

2017-04-01

Pathogenic allelic variants in the fibroblast growth factor receptor 3 (FGFR3) gene have been associated with a number of phenotypes including achondroplasia, hypochondroplasia, thanatophoric dysplasia, Crouzon syndrome with acanthosis nigricans (Crouzonodermoskeletal syndrome), and SADDAN (severe achondroplasia with developmental delay and acanthosis nigricans). Crouzon syndrome with acanthosis nigricans is caused by the pathogenic variant c.1172C>A (p.Ala391Glu) in the FGFR3 gene. The p.Lys650Thr pathogenic variant in FGFR3 has been linked to acanthosis nigricans without significant craniofacial or skeletal abnormalities. Recently, an infant with achondroplasia and a novel p.Ser348Cys FGFR3 mutation was reported. We describe the clinical history of an 8-year-old child with a skeletal dysplasia in the achondroplasia-hypochondroplasia spectrum, acanthosis nigricans, typical development, and the recently described p.Ser348Cys FGFR3 mutation. © 2017 Wiley Periodicals, Inc.

Direct assessment of the effect of the Gly380Arg achondroplasia mutation on FGFR3 dimerization using quantitative imaging FRET.

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Jesse Placone

Full Text Available The Gly380Arg mutation in FGFR3 is the genetic cause for achondroplasia (ACH, the most common form of human dwarfism. The mutation has been proposed to increase FGFR3 dimerization, but the dimerization propensities of wild-type and mutant FGFR3 have not been compared. Here we use quantitative imaging FRET to characterize the dimerization of wild-type FGFR3 and the ACH mutant in plasma membrane-derived vesicles from HEK293T cells. We demonstrate a small, but statistically significant increase in FGFR3 dimerization due to the ACH mutation. The data are consistent with the idea that the ACH mutation causes a structural change which affects both the stability and the activity of FGFR3 dimers in the absence of ligand.

Fibroblast growth factor receptor 3 interacts with and activates TGFβ-activated kinase 1 tyrosine phosphorylation and NFκB signaling in multiple myeloma and bladder cancer.

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Lisa Salazar

Full Text Available Cancer is a major public health problem worldwide. In the United States alone, 1 in 4 deaths is due to cancer and for 2013 a total of 1,660,290 new cancer cases and 580,350 cancer-related deaths are projected. Comprehensive profiling of multiple cancer genomes has revealed a highly complex genetic landscape in which a large number of altered genes, varying from tumor to tumor, impact core biological pathways and processes. This has implications for therapeutic targeting of signaling networks in the development of treatments for specific cancers. The NFκB transcription factor is constitutively active in a number of hematologic and solid tumors, and many signaling pathways implicated in cancer are likely connected to NFκB activation. A critical mediator of NFκB activity is TGFβ-activated kinase 1 (TAK1. Here, we identify TAK1 as a novel interacting protein and target of fibroblast growth factor receptor 3 (FGFR3 tyrosine kinase activity. We further demonstrate that activating mutations in FGFR3 associated with both multiple myeloma and bladder cancer can modulate expression of genes that regulate NFκB signaling, and promote both NFκB transcriptional activity and cell adhesion in a manner dependent on TAK1 expression in both cancer cell types. Our findings suggest TAK1 as a potential therapeutic target for FGFR3-associated cancers, and other malignancies in which TAK1 contributes to constitutive NFκB activation.

A meta-analysis of the relationship between FGFR3 and TP53 mutations in bladder cancer.

Directory of Open Access Journals (Sweden)

Yann Neuzillet

Full Text Available TP53 and FGFR3 mutations are the most common mutations in bladder cancers. FGFR3 mutations are most frequent in low-grade low-stage tumours, whereas TP53 mutations are most frequent in high-grade high-stage tumours. Several studies have reported FGFR3 and TP53 mutations to be mutually exclusive events, whereas others have reported them to be independent. We carried out a meta-analysis of published findings for FGFR3 and TP53 mutations in bladder cancer (535 tumours, 6 publications and additional unpublished data for 382 tumours. TP53 and FGFR3 mutations were not independent events for all tumours considered together (OR = 0.25 [0.18-0.37], p = 0.0001 or for pT1 tumours alone (OR = 0.47 [0.28-0.79], p = 0.0009. However, if the analysis was restricted to pTa tumours or to muscle-invasive tumours alone, FGFR3 and TP53 mutations were independent events (OR = 0.56 [0.23-1.36] (p = 0.12 and OR = 0.99 [0.37-2.7] (p = 0.35, respectively. After stratification of the tumours by stage and grade, no dependence was detected in the five tumour groups considered (pTaG1 and pTaG2 together, pTaG3, pT1G2, pT1G3, pT2-4. These differences in findings can be attributed to the putative existence of two different pathways of tumour progression in bladder cancer: the CIS pathway, in which FGFR3 mutations are rare, and the Ta pathway, in which FGFR3 mutations are frequent. TP53 mutations occur at the earliest stage of the CIS pathway, whereas they occur would much later in the Ta pathway, at the T1G3 or muscle-invasive stage.

[Mutation analysis of FGFR3 gene in a family featuring hereditary dwarfism].

Science.gov (United States)

Zhang, Qiong; Jiang, Hai-ou; Quan, Qing-li; Li, Jun; He, Ting; Huang, Xue-shuang

2011-12-01

To investigate the clinical symptoms and potential mutation in FGFR3 gene for a family featuring hereditary dwarfism in order to attain diagnosis and provide prenatal diagnosis. Five patients and two unaffected relatives from the family, in addition with 100 healthy controls, were recruited. Genome DNA was extracted. Exons 10 and 13 of the FGFR3 gene were amplified using polymerase chain reaction (PCR). PCR products were sequenced in both directions. All patients had similar features including short stature, short limbs, lumbar hyperlordosis but normal craniofacial features. A heterozygous mutation G1620T (N540K) was identified in the cDNA from all patients but not in the unaffected relatives and 100 control subjects. A heterozygous G380R mutation was excluded. The hereditary dwarfism featured by this family has been caused by hypochondroplasia (HCH) due to a N540K mutation in the FGFR3 gene.

Sixteen Years and Counting: The Current Understanding of Fibroblast Growth Factor Receptor 3 (FGFR3) Signaling in Skeletal Dysplasias

Czech Academy of Sciences Publication Activity Database

Foldynová-Trantírková, Silvie; Wilcox, W. R.; Krejčí, Pavel

2012-01-01

Roč. 33, č. 1 (2012), s. 29-41 ISSN 1059-7794 Grant - others:GA CR(CZ) GAP305/11/0752; GA CR(CZ) GA301/09/0587 Program:GA Institutional research plan: CEZ:AV0Z60220518; CEZ:AV0Z50040507 Keywords : FGFR3 * chondrocyte * skeletal dysplasia * MAP kinase * FGF Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.213, year: 2012 http://onlinelibrary.wiley.com/doi/10.1002/humu.21636/pdf

Suppressed hepatic bile acid signalling despite elevated production of primary and secondary bile acids in NAFLD.

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Jiao, Na; Baker, Susan S; Chapa-Rodriguez, Adrian; Liu, Wensheng; Nugent, Colleen A; Tsompana, Maria; Mastrandrea, Lucy; Buck, Michael J; Baker, Robert D; Genco, Robert J; Zhu, Ruixin; Zhu, Lixin

2017-08-03

Bile acids are regulators of lipid and glucose metabolism, and modulate inflammation in the liver and other tissues. Primary bile acids such as cholic acid and chenodeoxycholic acid (CDCA) are produced in the liver, and converted into secondary bile acids such as deoxycholic acid (DCA) and lithocholic acid by gut microbiota. Here we investigated the possible roles of bile acids in non-alcoholic fatty liver disease (NAFLD) pathogenesis and the impact of the gut microbiome on bile acid signalling in NAFLD. Serum bile acid levels and fibroblast growth factor 19 (FGF19), liver gene expression profiles and gut microbiome compositions were determined in patients with NAFLD, high-fat diet-fed rats and their controls. Serum concentrations of primary and secondary bile acids were increased in patients with NAFLD. In per cent, the farnesoid X receptor (FXR) antagonistic DCA was increased, while the agonistic CDCA was decreased in NAFLD. Increased mRNA expression for cytochrome P450 7A1, Na + -taurocholate cotransporting polypeptide and paraoxonase 1, no change in mRNA expression for small heterodimer partner and bile salt export pump, and reduced serum FGF19 were evidence of impaired FXR and fibroblast growth factor receptor 4 (FGFR4)-mediated signalling in NAFLD. Taurine and glycine metabolising bacteria were increased in the gut of patients with NAFLD, reflecting increased secondary bile acid production. Similar changes in liver gene expression and the gut microbiome were observed in high-fat diet-fed rats. The serum bile acid profile, the hepatic gene expression pattern and the gut microbiome composition consistently support an elevated bile acid production in NAFLD. The increased proportion of FXR antagonistic bile acid explains, at least in part, the suppression of hepatic FXR-mediated and FGFR4-mediated signalling. Our study suggests that future NAFLD intervention may target the components of FXR signalling, including the bile acid converting gut microbiome. © Article

FGFR Family Members Protein Expression as Prognostic Markers in Oral Cavity and Oropharyngeal Squamous Cell Carcinoma

NARCIS (Netherlands)

Koole, Koos; Clausen, Martijn J. A. M.; van Es, Robert J. J.; van Kempen, Pauline M. W.; Melchers, Lieuwe J.; Koole, Ron; Langendijk, Johannes A.; van Diest, Paul J.; Roodenburg, Jan L. N.; Schuuring, Ed; Willems, Stefan M.

Introduction Fibroblast growth factor receptor family member proteins (FGFR1-4) have been identified as promising novel therapeutic targets and prognostic markers in a wide spectrum of solid tumors. The present study investigates the expression and prognostic value of four FGFR family member

Temporal and occipital lobe features in children with hypochondroplasia/FGFR3 gene mutation.

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Philpott, Cristina M; Widjaja, Elysa; Raybaud, Charles; Branson, Helen M; Kannu, Peter; Blaser, Susan

2013-09-01

Thanatophoric dysplasia (TD) and hypochondroplasia are both caused by FGFR3 (fibroblast growth factor receptor 3) gene mutations. Temporal lobe dysplasia has been well described in thanatophoric dysplasia; however, only a couple of anecdotal cases of temporal lobe dysplasia in hypochondroplasia have been described. To define temporal lobe abnormalities in patients with hypochondroplasia, given that they share the same genetic mutation. We identified brain imaging studies of nine children with hypochondroplasia. The temporal lobes were assessed on CT and MRI for size and configuration of the temporal horn and aberrant sulcation of the inferior surface of the temporal lobe. All children had a triangular-shape temporal horn and deep transverse fissures of the inferior temporal lobe surface. Neuroimaging in our cohort revealed enlarged temporal lobes and oversulcation of the mesial temporal and occipital lobes, with abnormal inferomedial orientation of these redundant gyri. Hippocampal dysplasia was also universal. We confirmed frequent inferomesial temporal and occipital lobe abnormalities in our cohort of children with hypochondroplasia. Murine models with mutant fgfr3 display increased neuroprogenitor proliferation, cortical thickness and surface area in the temporo-occipital cortex. This is thought to result in excessive convolution and likely explains the imaging findings in this patient cohort. (Note that fgfr3 is the same genetic mutation in mice as FGFR3 is in humans.).

Early postnatal soluble FGFR3 therapy prevents the atypical development of obesity in achondroplasia.

Science.gov (United States)

Saint-Laurent, Celine; Garcia, Stephanie; Sarrazy, Vincent; Dumas, Karine; Authier, Florence; Sore, Sophie; Tran, Albert; Gual, Philippe; Gennero, Isabelle; Salles, Jean-Pierre; Gouze, Elvire

2018-01-01

Achondroplasia is a rare genetic disease is characterized by abnormal bone development and early obesity. While the bone aspect of the disease has been thoroughly studied, early obesity affecting approximately 50% of them during childhood has been somewhat neglected. It nevertheless represents a major health problem in these patients, and is associated to life-threatening complications including increasing risk of cardiovascular pathologies. We have thus decided to study obesity in patients and to use the mouse model to evaluate if soluble FGFR3 therapy, an innovative treatment approach for achondroplasia, could also impact the development of this significant complication. To achieve this, we have first fully characterized the metabolic deregulations in these patients by conducting a longitudinal retrospective study, in children with achondroplasia Anthropometric, densitometric measures as well as several blood parameters were recorded and compared between three age groups ranging from [0-3], [4-8] and [9-18] years old. Our results show unexpected results with the development of an atypical obesity with preferential fat deposition in the abdomen that is remarkably not associated with classical complications of obesity such as diabetes or hypercholosterolemia. Because it is not associated with diabetes, the atypical obesity has not been studied in the past even though it is recognized as a real problem in these patients. These results were validated in a murine model of achondroplasia (Fgfr3ach/+) where similar visceral adiposity was observed. Unexpected alterations in glucose metabolism were highlighted during high-fat diet. Glucose, insulin or lipid levels remained low, without the development of diabetes. Very interestingly, in achondroplasia mice treated with soluble FGFR3 during the growth period (from D3 to D22), the development of these metabolic deregulations was prevented in adult animals (between 4 and 14 weeks of age). The lean-over-fat tissues ratio was

Fibroblast growth factor-2 up-regulates the expression of nestin through the Ras–Raf–ERK–Sp1 signaling axis in C6 glioma cells

International Nuclear Information System (INIS)

Chang, Kai-Wei; Huang, Yuan-Li; Wong, Zong-Ruei; Su, Peng-Han; Huang, Bu-Miin; Ju, Tsai-Kai; Yang, Hsi-Yuan

2013-01-01

Highlights: •Nestin expression in C6 glioma cells is induced by FGF-2. •Nestin expression is induced by FGF-2 via de novo RNA and protein synthesis. •The FGFR inhibitor SU5402 blocks the FGF-2-induced nestin expression. •The mRNA of FGFR1 and 3 are detected in C6 glioma cells. •Ras–Raf–ERK–Sp1 signaling pathway is responsibe for FGF-2-induced nestin expression. -- Abstract: Nestin is a 240-kDa intermediate filament protein expressed mainly in neural and myogenic stem cells. Although a substantial number of studies have focused on the expression of nestin during development of the central nervous system, little is known about the factors that induce and regulate its expression. Fibroblast growth factor-2 (FGF-2) is an effective mitogen and stimulates the proliferation and differentiation of a subset of nestin-expressing cells, including neural progenitor cells, glial precursor cells, and smooth muscle cells. To assess whether FGF-2 is a potent factor that induces the expression of nestin, C6 glioma cells were used. The results showed that nestin expression was up-regulated by FGF-2 via de novo RNA and protein synthesis. Our RT-PCR results showed that C6 glioma cells express FGFR1/3, and FGFRs is required for FGF-2-induced nestin expression. Further signaling analysis also revealed that FGF-2-induced nestin expression is mediated through FGFR–MAPK–ERK signaling axis and the transcriptional factor Sp1. These findings provide new insight into the regulation of nestin in glial system and enable the further studies on the function of nestin in glial cells

Fibroblast growth factor-2 up-regulates the expression of nestin through the Ras–Raf–ERK–Sp1 signaling axis in C6 glioma cells

Energy Technology Data Exchange (ETDEWEB)

Chang, Kai-Wei [Institute of Molecular and Cellular Biology, National Taiwan University, Taipei 106, Taiwan (China); Huang, Yuan-Li [Department of Biotechnology, College of Health Science, Asia University, Taichung 413, Taiwan (China); Wong, Zong-Ruei; Su, Peng-Han [Institute of Molecular and Cellular Biology, National Taiwan University, Taipei 106, Taiwan (China); Huang, Bu-Miin [Department of Cell Biology and Anatomy, National Cheng-Kung University, Tainan 701, Taiwan (China); Ju, Tsai-Kai [Instrumentation Center, National Taiwan University, Taipei 106, Taiwan (China); Technology Commons, College of Life Science, National Taiwan University, Taipei 106, Taiwan (China); Yang, Hsi-Yuan, E-mail: hyhy@ntu.edu.tw [Institute of Molecular and Cellular Biology, National Taiwan University, Taipei 106, Taiwan (China)

2013-05-17

Highlights: •Nestin expression in C6 glioma cells is induced by FGF-2. •Nestin expression is induced by FGF-2 via de novo RNA and protein synthesis. •The FGFR inhibitor SU5402 blocks the FGF-2-induced nestin expression. •The mRNA of FGFR1 and 3 are detected in C6 glioma cells. •Ras–Raf–ERK–Sp1 signaling pathway is responsibe for FGF-2-induced nestin expression. -- Abstract: Nestin is a 240-kDa intermediate filament protein expressed mainly in neural and myogenic stem cells. Although a substantial number of studies have focused on the expression of nestin during development of the central nervous system, little is known about the factors that induce and regulate its expression. Fibroblast growth factor-2 (FGF-2) is an effective mitogen and stimulates the proliferation and differentiation of a subset of nestin-expressing cells, including neural progenitor cells, glial precursor cells, and smooth muscle cells. To assess whether FGF-2 is a potent factor that induces the expression of nestin, C6 glioma cells were used. The results showed that nestin expression was up-regulated by FGF-2 via de novo RNA and protein synthesis. Our RT-PCR results showed that C6 glioma cells express FGFR1/3, and FGFRs is required for FGF-2-induced nestin expression. Further signaling analysis also revealed that FGF-2-induced nestin expression is mediated through FGFR–MAPK–ERK signaling axis and the transcriptional factor Sp1. These findings provide new insight into the regulation of nestin in glial system and enable the further studies on the function of nestin in glial cells.

Mutational screening of FGFR1, CER1, and CDON in a large cohort of trigonocephalic patients.

Science.gov (United States)

Jehee, Fernanda Sarquis; Alonso, Luis G; Cavalcanti, Denise P; Kim, Chong; Wall, Steven A; Mulliken, John B; Sun, Miao; Jabs, Ethylin Wang; Boyadjiev, Simeon A; Wilkie, Andrew O M; Passos-Bueno, Maria Rita

2006-03-01

Screen the known craniosynostotic related gene, FGFR1 (exon 7), and two new identified potential candidates, CER1 and CDON, in patients with syndromic and nonsyndromic metopic craniosynostosis to determine if they might be causative genes. Using single-strand conformational polymorphisms (SSCPs), denaturing high-performance liquid chromatography, and/or direct sequencing, we analyzed a total of 81 patients for FGFR1 (exon 7), 70 for CER1, and 44 for CDON. Patients were ascertained in the Centro de Estudos do Genoma Humano in São Paulo, Brazil (n = 39), the Craniofacial Unit, Oxford, U.K. (n = 23), and the Johns Hopkins University, Baltimore, Maryland (n = 31). Clinical inclusion criteria included a triangular head and/or forehead, with or without a metopic ridge, and a radiographic documentation of metopic synostosis. Both syndromic and nonsyndromic patients were studied. No sequence alterations were found for FGFR1 (exon 7). Different patterns of SSCP migration for CER1 compatible with the segregation of single nucleotide polymorphisms reported in the region were identified. Seventeen sequence alterations were detected in the coding region of CDON, seven of which are new, but segregation analysis in parents and homology studies did not indicate a pathological role. FGFR1 (exon 7), CER1, and CDON are not related to trigonocephaly in our sample and should not be considered as causative genes for metopic synostosis. Screening of FGFR1 (exon 7) for diagnostic purposes should not be performed in trigonocephalic patients.

Blocking Ihh signaling pathway inhibits the proliferation and promotes the apoptosis of PSCs.

Science.gov (United States)

Xu, Kai; Guo, Fengjing; Zhang, Shuwei; Liu, Cheng; Wang, Feixiong; Zhou, Zhiguo; Chen, Anmin

2009-02-01

The roles of Indian hedgehog (Ihh) signaling pathway in the proliferation and apoptosis of precartilaginous stem cells (PSCs) were investigated. PSCs, labeled with fibroblast growth factor receptor 3 (FGFR-3), were isolated from neonatal rats by immunomagnetic separation. After identification with FGFR-3 and Col II, the cells were incubated with different concentrations of cyclopamine (cyclo), the specific inhibitor of Ihh signaling pathway. The morphologic changes of the cells were observed under the inverted phase contrast microscope. The mRNA expression levels of Ihh, parathyroid hormonerelated peptide (PTHrP), protein Patched (Ptch), Bcl-2 and p21 were detected by RT-PCR. The protein expression levels of Ihh and Ptch were measured by Western blot. MTT assay was used to examine the effects of cyclo on proliferation of PSCs. Apoptosis rate of PSCs was examined by Annexin V/PI assay of flow cytometric analyses. After PSCs were incubated with cyclo, obvious morphologic changes were observed as compared with the control group. The mRNA expression levels of PTHrP, Ptch and Bcl-2 were decreased to varying degrees in a cyclo dose-dependent manner. However, the expression levels of Ihh and p21 mRNA were increased. The protein expression of Ptch and Ihh had the same change as the mRNA expression. Meanwhile, cyclo could obviously inhibit the proliferation and promote the apoptosis of PSCs. The results indicated that Ihh signaling pathway plays an important role in regulating the proliferation and apoptosis of PSCs, which is probably mediated by Bcl-2 and p21.

Apert Syndrome With FGFR2 758 C > G Mutation: A Chinese Case Report

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Yahong Li

2018-05-01

Full Text Available Background: Apert syndrome is considered as one of the most common craniosynostosis syndromes with a prevalence of 1 in 65,000 individuals, and has a close relationship with point mutations in FGFR2 gene.Case report: Here, we described a Apert syndrome case, who was referred to genetic consultation in our hospital with the symptom of craniosynostosis and syndactyly of the hands and feet. Craniosynostosis, midfacial retrusion, steep wide forehead, larger head circumference, marked depression of the nasal bridge, short and wide nose and proptosis could be found obviously, apart from these, ears were mildly low compared with normal children and there was no cleft lip and palate. Mutation was identified by sanger sequencing and a mutation in the exon 7 of FGFR2 gene was detected: p.Pro253Arg (P253R 758 C > G, which was not found in his parents.Conclusion: The baby had Apert syndrome caused by 758 C > G mutation in the exon 7 of FGFR2 gene, considering no this mutation in his parents, it was spontaneous.

Dual Targeting of PDGFRα and FGFR1 Displays Synergistic Efficacy in Malignant Rhabdoid Tumors

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Jocelyn P. Wong

2016-10-01

Full Text Available Subunits of the SWI/SNF chromatin remodeling complex are mutated in a significant proportion of human cancers. Malignant rhabdoid tumors (MRTs are lethal pediatric cancers characterized by a deficiency in the SWI/SNF subunit SMARCB1. Here, we employ an integrated molecular profiling and chemical biology approach to demonstrate that the receptor tyrosine kinases (RTKs PDGFRα and FGFR1 are coactivated in MRT cells and that dual blockade of these receptors has synergistic efficacy. Inhibitor combinations targeting both receptors and the dual inhibitor ponatinib suppress the AKT and ERK1/2 pathways leading to apoptosis. MRT cells that have acquired resistance to the PDGFRα inhibitor pazopanib are susceptible to FGFR inhibitors. We show that PDGFRα levels are regulated by SMARCB1 expression, and assessment of clinical specimens documents the expression of both PDGFRα and FGFR1 in rhabdoid tumor patients. Our findings support a therapeutic approach in cancers with SWI/SNF deficiencies by exploiting RTK coactivation dependencies.

Early postnatal soluble FGFR3 therapy prevents the atypical development of obesity in achondroplasia

Science.gov (United States)

Sarrazy, Vincent; Dumas, Karine; Authier, Florence; Sore, Sophie; Tran, Albert; Gual, Philippe; Gennero, Isabelle; Salles, Jean-Pierre; Gouze, Elvire

2018-01-01

Background Achondroplasia is a rare genetic disease is characterized by abnormal bone development and early obesity. While the bone aspect of the disease has been thoroughly studied, early obesity affecting approximately 50% of them during childhood has been somewhat neglected. It nevertheless represents a major health problem in these patients, and is associated to life-threatening complications including increasing risk of cardiovascular pathologies. We have thus decided to study obesity in patients and to use the mouse model to evaluate if soluble FGFR3 therapy, an innovative treatment approach for achondroplasia, could also impact the development of this significant complication. Methods and findings To achieve this, we have first fully characterized the metabolic deregulations in these patients by conducting a longitudinal retrospective study, in children with achondroplasia Anthropometric, densitometric measures as well as several blood parameters were recorded and compared between three age groups ranging from [0–3], [4–8] and [9–18] years old. Our results show unexpected results with the development of an atypical obesity with preferential fat deposition in the abdomen that is remarkably not associated with classical complications of obesity such as diabetes or hypercholosterolemia. Because it is not associated with diabetes, the atypical obesity has not been studied in the past even though it is recognized as a real problem in these patients. These results were validated in a murine model of achondroplasia (Fgfr3ach/+) where similar visceral adiposity was observed. Unexpected alterations in glucose metabolism were highlighted during high-fat diet. Glucose, insulin or lipid levels remained low, without the development of diabetes. Very interestingly, in achondroplasia mice treated with soluble FGFR3 during the growth period (from D3 to D22), the development of these metabolic deregulations was prevented in adult animals (between 4 and 14 weeks of age

Hypochondroplasia, Acanthosis Nigricans, and Insulin Resistance in a Child with FGFR3 Mutation: Is It Just an Association?

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Manal Mustafa

2014-01-01

Full Text Available FGFR3 mutations cause wide spectrum of disorders ranging from skeletal dysplasias (hypochondroplasia, achondroplasia, and thanatophoric dysplasia, benign skin tumors (epidermal nevi, seborrhaeic keratosis, and acanthosis nigricans, and epithelial malignancies (multiple myeloma and prostate and bladder carcinoma. Hypochondroplasia is the most common type of short-limb dwarfism in children resulting from fibroblast growth factor receptor 3 (FGFR3 mutation. Acanthosis nigricans might be seen in severe skeletal dysplasia, including thanatophoric dysplasia and SADDAN syndrome, without a biochemical evidence of hyperinsulinemia. Insulin insensitivity and acanthosis nigricans are uncommonly seen in hypochondroplasia patients with FGFR3 mutations which may represent a new association. We aim to describe the association of hypochondroplasia, acanthosis nigricans, and insulin resistance in a child harboring FGFR3 mutation. To our knowledge, this is the first case report associating the p.N540 with acanthosis nigricans and the second to describe hyperinsulinemia in hypochondroplasia. This finding demonstrates the possible coexistence of insulin insensitivity and acanthosis nigricans in hypochondroplasia patients.

The clinical pathological characteristics and prognosis of FGFR1 gene amplification in non-small-cell lung cancer: a meta-analysis

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Xie FJ

2016-01-01

Full Text Available Fa-Jun Xie,1,2 Hong-Yang Lu,1,3 Qiu-Qing Zheng,3 Jing Qin,1,3 Yun Gao,3 Yi-Ping Zhang,1,3 Xun Hu,2 Wei-Min Mao3,4 1Department of Medical Oncology, Zhejiang Cancer Hospital, 2Cancer Institute (Key Laboratory for Cancer Intervention and Prevention, China National Ministry of Education, Zhejiang Provincial Key Laboratory of Molecular Biology in Medical Sciences, Second Affiliated Hospital, Zhejiang University School of Medicine,3Zhejiang Key Laboratory of Diagnosis and Treatment Technology on Thoracic Oncology (Lung and Esophagus, Hangzhou, 4Department of Thoracic Surgery, Zhejiang Cancer Hospital, Hangzhou, People’s Republic of China Abstract: FGFR1 amplification is recognized as a novel therapy target for non-small-cell lung cancer (NSCLC, especially in squamous cell carcinoma (SCC. However, the association between FGFR1 amplification and the clinicopathological characteristics of NSCLC remains controversial. We performed a meta-analysis of 17 eligible studies to examine the correlation between FGFR1 gene amplification and clinicopathological characteristics. FGFR1 amplification was closely related to these clinicopathological features, including sex (odds ratio [OR] 2.05, 95% confidence interval [CI] 1.50–2.80, smoking (OR 3.31, 95% CI 2.02–5.44, and histology (OR 3.60, 95% CI 2.82–4.59. FGFR1 amplification was associated with shorter overall survival, and no significant heterogeneity existed between studies (I2=3.8%. We should note that publication bias may partly account for these results, but our findings remained significant after the trim-and-fill method (hazard ratio 1.22, 95% CI 1.06–1.40. However, no significant correlation was found with poor disease-free survival (hazard ratio 1.43, 95% CI 0.96–2.12. In conclusion, this study showed that FGFR1 amplification was significantly associated with sex, smoking, and histology. FGFR1 amplification could be a marker of poor prognosis in NSCLC patients, especially in SCC patients

Tyrosine 769 of the keratinocyte growth factor receptor is required for receptor signaling but not endocytosis

International Nuclear Information System (INIS)

Ceridono, Mara; Belleudi, Francesca; Ceccarelli, Simona; Torrisi, Maria Rosaria

2005-01-01

Keratinocyte growth factor receptor (KGFR) is a receptor tyrosine kinase expressed on epithelial cells which belongs to the family of fibroblast growth factor receptors (FGFRs). Following ligand binding, KGFR is rapidly autophosphorylated on specific tyrosine residues in the intracellular domain, recruits substrate proteins, and is rapidly internalized by clathrin-mediated endocytosis. The role of different autophosphorylation sites in FGFRs, and in particular the role of the tyrosine 766 in FGFR1, first identified as PLCγ binding site, has been extensively studied. We analyzed here the possible role of the tyrosine 769 in KGFR, corresponding to tyrosine 766 in FGFR1, in the regulation of KGFR signal transduction and MAPK activation as well as in the control of the endocytic process of KGFR. A mutant KGFR in which tyrosine 769 was substituted by phenylalanine was generated and transfected in NIH3T3 and HeLa cells. Our results indicate that tyrosine 769 is required for the binding to KGFR and tyrosine phosphorylation of PLCγ as well as for the full activation of MAPKs and for cell proliferation through the regulation of FRS2 tyrosine phosphorylation, suggesting that this residue represents a key regulator of KGFR signal transduction. Our data also show that tyrosine 769 is not involved in the regulation of the endocytic process of KGFR

Negative regulation of RIG-I-mediated antiviral signaling by TRK-fused gene (TFG) protein

International Nuclear Information System (INIS)

Lee, Na-Rae; Shin, Han-Bo; Kim, Hye-In; Choi, Myung-Soo; Inn, Kyung-Soo

2013-01-01

Highlights: •TRK-fused gene product (TFG) interacts with TRIM25 upon viral infection. •TFG negatively regulates RIG-I mediated antiviral signaling. •TFG depletion leads to enhanced viral replication. •TFG act downstream of MAVS. -- Abstract: RIG-I (retinoic acid inducible gene I)-mediated antiviral signaling serves as the first line of defense against viral infection. Upon detection of viral RNA, RIG-I undergoes TRIM25 (tripartite motif protein 25)-mediated K63-linked ubiquitination, leading to type I interferon (IFN) production. In this study, we demonstrate that TRK-fused gene (TFG) protein, previously identified as a TRIM25-interacting protein, binds TRIM25 upon virus infection and negatively regulates RIG-I-mediated type-I IFN signaling. RIG-I-mediated IFN production and nuclear factor (NF)-κB signaling pathways were upregulated by the suppression of TFG expression. Furthermore, vesicular stomatitis virus (VSV) replication was significantly inhibited by small inhibitory hairpin RNA (shRNA)-mediated knockdown of TFG, supporting the suppressive role of TFG in RIG-I-mediated antiviral signaling. Interestingly, suppression of TFG expression increased not only RIG-I-mediated signaling but also MAVS (mitochondrial antiviral signaling protein)-induced signaling, suggesting that TFG plays a pivotal role in negative regulation of RNA-sensing, RIG-I-like receptor (RLR) family signaling pathways

Identification of fibroblast growth factor receptor 3 (FGFR3 as a protein receptor for botulinum neurotoxin serotype A (BoNT/A.

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Birgitte P S Jacky

Full Text Available Botulinum neurotoxin serotype A (BoNT/A causes transient muscle paralysis by entering motor nerve terminals (MNTs where it cleaves the SNARE protein Synaptosomal-associated protein 25 (SNAP25206 to yield SNAP25197. Cleavage of SNAP25 results in blockage of synaptic vesicle fusion and inhibition of the release of acetylcholine. The specific uptake of BoNT/A into pre-synaptic nerve terminals is a tightly controlled multistep process, involving a combination of high and low affinity receptors. Interestingly, the C-terminal binding domain region of BoNT/A, HC/A, is homologous to fibroblast growth factors (FGFs, making it a possible ligand for Fibroblast Growth Factor Receptors (FGFRs. Here we present data supporting the identification of Fibroblast Growth Factor Receptor 3 (FGFR3 as a high affinity receptor for BoNT/A in neuronal cells. HC/A binds with high affinity to the two extra-cellular loops of FGFR3 and acts similar to an agonist ligand for FGFR3, resulting in phosphorylation of the receptor. Native ligands for FGFR3; FGF1, FGF2, and FGF9 compete for binding to FGFR3 and block BoNT/A cellular uptake. These findings show that FGFR3 plays a pivotal role in the specific uptake of BoNT/A across the cell membrane being part of a larger receptor complex involving ganglioside- and protein-protein interactions.

DMPD: Negative regulation of cytoplasmic RNA-mediated antiviral signaling. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 18703349 Negative regulation of cytoplasmic RNA-mediated antiviral signaling. Komur...Show Negative regulation of cytoplasmic RNA-mediated antiviral signaling. PubmedID 18703349 Title Negative r...egulation of cytoplasmic RNA-mediated antiviral signaling. Authors Komuro A, Bamm

The fibroblast growth factor receptor (FGFR) agonist FGF1 and the neural cell adhesion molecule-derived peptide FGL activate FGFR substrate 2alpha differently

DEFF Research Database (Denmark)

Chen, Yongshuo; Li, Shizhong; Berezin, Vladimir

2010-01-01

Activation of fibroblast growth factor (FGF) receptors (FGFRs) both by FGFs and by the neural cell adhesion molecule (NCAM) is crucial in the development and function of the nervous system. We found that FGFR substrate 2alpha (FRS2alpha), Src homologous and collagen A (ShcA), and phospholipase-Cg...

ROS and ROS-Mediated Cellular Signaling

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Jixiang Zhang

2016-01-01

Full Text Available It has long been recognized that an increase of reactive oxygen species (ROS can modify the cell-signaling proteins and have functional consequences, which successively mediate pathological processes such as atherosclerosis, diabetes, unchecked growth, neurodegeneration, inflammation, and aging. While numerous articles have demonstrated the impacts of ROS on various signaling pathways and clarify the mechanism of action of cell-signaling proteins, their influence on the level of intracellular ROS, and their complex interactions among multiple ROS associated signaling pathways, the systemic summary is necessary. In this review paper, we particularly focus on the pattern of the generation and homeostasis of intracellular ROS, the mechanisms and targets of ROS impacting on cell-signaling proteins (NF-κB, MAPKs, Keap1-Nrf2-ARE, and PI3K-Akt, ion channels and transporters (Ca2+ and mPTP, and modifying protein kinase and Ubiquitination/Proteasome System.

Detection of low frequency FGFR3 mutations in the urine of bladder cancer patients using next-generation deep sequencing

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Millholl

2012-06-01

Full Text Available John M Millholland, Shuqiang Li, Cecilia A Fernandez, Anthony P ShuberPredictive Biosciences Inc, Lexington, MA, USAAbstract: Biological fluid-based noninvasive biomarker assays for monitoring and diagnosing disease are clinically powerful. A major technical hurdle for developing these assays is the requirement of high analytical sensitivity so that biomarkers present at very low levels can be consistently detected. In the case of biological fluid-based cancer diagnostic assays, sensitivities similar to those of tissue-based assays are difficult to achieve with DNA markers due to the high abundance of normal DNA background present in the sample. Here we describe a new urine-based assay that uses ultradeep sequencing technology to detect single mutant molecules of fibroblast growth factor receptor 3 (FGFR3 DNA that are indicative of bladder cancer. Detection of FGFR3 mutations in urine would provide clinicians with a noninvasive means of diagnosing early-stage bladder cancer. The single-molecule assay detects FGFR3 mutant DNA when present at as low as 0.02% of total urine DNA and results in 91% concordance with the frequency that FGFR3 mutations are detected in bladder cancer tumors, significantly improving diagnostic performance. To our knowledge, this is the first practical application of next-generation sequencing technology for noninvasive cancer diagnostics.Keywords: FGFR3, mutation, urine, single molecule, sequencing, bladder cancer

Fibroblast growth factor receptor 2 regulates proliferation and Sertoli differentiation during male sex determination

Science.gov (United States)

Kim, Yuna; Bingham, Nathan; Sekido, Ryohei; Parker, Keith L.; Lovell-Badge, Robin; Capel, Blanche

2007-01-01

Targeted mutagenesis of Fgf9 in mice causes male-to-female sex reversal. Among the four FGF receptors, FGFR2 showed two highly specific patterns based on antibody staining, suggesting that it might be the receptor-mediating FGF9 signaling in the gonad. FGFR2 was detected at the plasma membrane in proliferating coelomic epithelial cells and in the nucleus in Sertoli progenitor cells. This expression pattern suggested that Fgfr2 might play more than one role in testis development. To test the hypothesis that Fgfr2 is required for male sex determination, we crossed mice carrying a floxed allele of Fgfr2 with two different Cre lines to induce a temporal or cell-specific deletion of this receptor. Results show that deletion of Fgfr2 in embryonic gonads phenocopies deletion of Fgf9 and leads to male-to-female sex reversal. Using these two Cre lines, we provide the first genetic evidence that Fgfr2 plays distinct roles in proliferation and Sertoli cell differentiation during testis development. PMID:17940049

Cigarette smoke regulates VEGFR2-mediated survival signaling in rat lungs

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Stevenson Christopher S

2010-02-01

Full Text Available Abstract Background Vascular endothelial growth factor (VEGF and VEGF receptor 2 (VEGFR2-mediated survival signaling is critical to endothelial cell survival, maintenance of the vasculature and alveolar structure and regeneration of lung tissue. Reduced VEGF and VEGFR2 expression in emphysematous lungs has been linked to increased endothelial cell death and vascular regression. Previously, we have shown that CS down-regulated the VEGFR2 and its downstream signaling in mouse lungs. However, the VEGFR2-mediated survival signaling in response to oxidants/cigarette smoke (CS is not known. We hypothesized that CS exposure leads to disruption of VEGFR2-mediated endothelial survival signaling in rat lungs. Methods Adult male Sprague-Dawley rats were exposed CS for 3 days, 8 weeks and 6 months to investigate the effect of CS on VEGFR2-mediated survival signaling by measuring the Akt/PI3-kinase/eNOS downstream signaling in rat lungs. Results and Discussion We show that CS disrupts VEGFR2/PI3-kinase association leading to decreased Akt and eNOS phosphorylation. This may further alter the phosphorylation of the pro-apoptotic protein Bad and increase the Bad/Bcl-xl association. However, this was not associated with a significant lung cell death as evidenced by active caspase-3 levels. These data suggest that although CS altered the VEGFR2-mediated survival signaling in the rat lungs, but it was not sufficient to cause lung cell death. Conclusion The rat lungs exposed to CS in acute, sub-chronic and chronic levels may be representative of smokers where survival signaling is altered but was not associated with lung cell death whereas emphysema is known to be associated with lung cell apoptosis.

Phosphoinositide protein kinase PDPK1 is a crucial cell signaling mediator in multiple myeloma.

Science.gov (United States)

Chinen, Yoshiaki; Kuroda, Junya; Shimura, Yuji; Nagoshi, Hisao; Kiyota, Miki; Yamamoto-Sugitani, Mio; Mizutani, Shinsuke; Sakamoto, Natsumi; Ri, Masaki; Kawata, Eri; Kobayashi, Tsutomu; Matsumoto, Yosuke; Horiike, Shigeo; Iida, Shinsuke; Taniwaki, Masafumi

2014-12-15

Multiple myeloma is a cytogenetically/molecularly heterogeneous hematologic malignancy that remains mostly incurable, and the identification of a universal and relevant therapeutic target molecule is essential for the further development of therapeutic strategy. Herein, we identified that 3-phosphoinositide-dependent protein kinase 1 (PDPK1), a serine threonine kinase, is expressed and active in all eleven multiple myeloma-derived cell lines examined regardless of the type of cytogenetic abnormality, the mutation state of RAS and FGFR3 genes, or the activation state of ERK and AKT. Our results revealed that PDPK1 is a pivotal regulator of molecules that are essential for myelomagenesis, such as RSK2, AKT, c-MYC, IRF4, or cyclin Ds, and that PDPK1 inhibition caused the growth inhibition and the induction of apoptosis with the activation of BIM and BAD, and augmented the in vitro cytotoxic effects of antimyeloma agents in myeloma cells. In the clinical setting, PDPK1 was active in myeloma cells of approximately 90% of symptomatic patients at diagnosis, and the smaller population of patients with multiple myeloma exhibiting myeloma cells without active PDPK1 showed a significantly less frequent proportion of the disease stage III by the International Staging System and a significantly more favorable prognosis, including the longer overall survival period and the longer progression-free survival period by bortezomib treatment, than patients with active PDPK1, suggesting that PDPK1 activation accelerates the disease progression and the resistance to treatment in multiple myeloma. Our study demonstrates that PDPK1 is a potent and a universally targetable signaling mediator in multiple myeloma regardless of the types of cytogenetic/molecular profiles. ©2014 American Association for Cancer Research.

DMPD: IRAK1: a critical signaling mediator of innate immunity. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 17890055 IRAK1: a critical signaling mediator of innate immunity. Gottipati S, Rao ...IRAK1: a critical signaling mediator of innate immunity. PubmedID 17890055 Title IRAK1: a critical signaling mediator

Spared pre-irradiated area in pustular lesions induced by icotinib showing decreased expressions of CD1a+ langerhans cells and FGFR2

International Nuclear Information System (INIS)

Zhao Qiong; Wang Yi Na; Wang Bo

2013-01-01

Icotinib hydrochloride, a novel inhibitor of epidermal growth factor receptor tyrosine kinase, has been approved by the State Food and Drug Administration for the treatment of advanced non-small-cell lung cancer. Up to date, cutaneous response to icotinib is largely unknown. Here we report an uncommon lesional phenomenon in a 56-year-old Chinese male with non-small-cell lung cancer, who received icotinib as a second-line treatment. Characteristic papulopustular rash on the chest and back was observed 4 days later. Interestingly, the rash completely spares a pre-irradiated area. The immunohistochemical study in the lesional skin area and spared skin area revealed a significant decrease in CD1a + Langerhans cells, Ki-67 as well as FGFR2 in the spared area than in the lesional area. Thus, the present case indicated that loss of the basal layer of proliferative cells and antigen-presenting cells (Langerhans cell), as well as the down-regulation of FGFR2 signaling in the pre-irradiated skin area, may join forces in inhibiting icotinib-associated cutaneous reactions. To our knowledge, this is the first report of both lesional area and lesion-spared area in a Chinese male receiving treatment with a new epidermal growth factor receptor-tyrosine kinase inhibitor (icotinib). The immunohistochemical reactions described here also provide new insight into the pathogenesis of epidermal growth factor receptor-tyrosine kinase inhibitor-related skin toxicities, and the role that other tyrosine kinase receptors (including FGFR) played in non-small-cell lung cancer. (author)

Spared pre-irradiated area in pustular lesions induced by icotinib showing decreased expressions of CD1a+ langerhans cells and FGFR2.

Science.gov (United States)

Zhao, Qiong; Wang, Yi Na; Wang, Bo

2013-02-01

Icotinib hydrochloride, a novel inhibitor of epidermal growth factor receptor tyrosine kinase, has been approved by the State Food and Drug Administration for the treatment of advanced non-small-cell lung cancer. Up to date, cutaneous response to icotinib is largely unknown. Here we report an uncommon lesional phenomenon in a 56-year-old Chinese male with non-small-cell lung cancer, who received icotinib as a second-line treatment. Characteristic papulopustular rash on the chest and back was observed 4 days later. Interestingly, the rash completely spares a pre-irradiated area. The immunohistochemical study in the lesional skin area and spared skin area revealed a significant decrease in CD1a(+) Langerhans cells, Ki-67 as well as FGFR2 in the spared area than in the lesional area. Thus, the present case indicated that loss of the basal layer of proliferative cells and antigen-presenting cells (Langerhans cell), as well as the down-regulation of FGFR2 signaling in the pre-irradiated skin area, may join forces in inhibiting icotinib-associated cutaneous reactions. To our knowledge, this is the first report of both lesional area and lesion-spared area in a Chinese male receiving treatment with a new epidermal growth factor receptor-tyrosine kinase inhibitor (icotinib). The immunohistochemical reactions described here also provide new insight into the pathogenesis of epidermal growth factor receptor-tyrosine kinase inhibitor-related skin toxicities, and the role that other tyrosine kinase receptors (including FGFR) played in non-small-cell lung cancer.

The role of phosphatidylinositol 3-kinase in neural cell adhesion molecule-mediated neuronal differentiation and survival

DEFF Research Database (Denmark)

Ditlevsen, Dorte K; Køhler, Lene B; Pedersen, Martin Volmer

2003-01-01

The neural cell adhesion molecule, NCAM, is known to stimulate neurite outgrowth from primary neurones and PC12 cells presumably through signalling pathways involving the fibroblast growth factor receptor (FGFR), protein kinase A (PKA), protein kinase C (PKC), the Ras-mitogen activated protein...... kinase (MAPK) pathway and an increase in intracellular Ca2+ levels. Stimulation of neurones with the synthetic NCAM-ligand, C3, induces neurite outgrowth through signalling pathways similar to the pathways activated through physiological, homophilic NCAM-stimulation. We present here data indicating...... that phosphatidylinositol 3-kinase (PI3K) is required for NCAM-mediated neurite outgrowth from PC12-E2 cells and from cerebellar and dopaminergic neurones in primary culture, and that the thr/ser kinase Akt/protein kinase B (PKB) is phosphorylated downstream of PI3K after stimulation with C3. Moreover, we present data...

[Rapid detection of hot spot mutations of FGFR3 gene with PCR-high resolution melting assay].

Science.gov (United States)

Li, Shan; Wang, Han; Su, Hua; Gao, Jinsong; Zhao, Xiuli

2017-08-10

To identify the causative mutations in five individuals affected with dyschondroplasia and develop an efficient procedure for detecting hot spot mutations of the FGFR3 gene. Genomic DNA was extracted from peripheral blood samples with a standard phenol/chloroform method. PCR-Sanger sequencing was used to analyze the causative mutations in the five probands. PCR-high resolution melting (HRM) was developed to detect the identified mutations. A c.1138G>A mutation in exon 8 was found in 4 probands, while a c.1620C>G mutation was found in exon 11 of proband 5 whom had a mild phenotype. All patients were successfully distinguished from healthy controls with the PCR-HRM method. The results of HRM analysis were highly consistent with that of Sanger sequencing. The Gly380Arg and Asn540Lys are hot spot mutations of the FGFR3 gene among patients with ACH/HCH. PCR-HRM analysis is more efficient for detecting hot spot mutations of the FGFR3 gene.

Bent bone dysplasia (BBD)-FGFR2 type: the radiologic manifestations in early gestation

Energy Technology Data Exchange (ETDEWEB)

Handa, Atsuhiko; Okajima, Yuka; Kurihara, Yasuyuki [St. Luke' s International Hospital, Department of Radiology, Tokyo (Japan); Izumi, Noriko; Yamanaka, Michiko [St. Luke' s International Hospital, Department of Integrated Women' s Health, Tokyo (Japan)

2016-02-15

Bent bone dysplasia-fibroblast growth factor receptor 2 type (BBD-FGFR2) is a recently identified skeletal dysplasia caused by specific FGFR2 mutations, characterized by craniosynostosis and prenatal bowing of the long bones. Only a few cases have been published. We report an affected fetus terminated at 21 weeks of gestation. The clinical and radiologic manifestations mostly recapitulate previous descriptions; however we suggest additional hallmarks of this disorder in early gestation. These hallmarks include distinctive short, thick clavicles and wavy ribs, as well as vertebral bodies that showed striking anteroposterior shortening. Femoral fractures were also present in our case. Although craniosynostosis is a hallmark of the disease, clinicians should be aware that craniosynostosis might not be readily apparent on plain films early in gestation. (orig.)

FGF10: A multifunctional mesenchymal-epithelial signaling growth factor in development, health, and disease.

Science.gov (United States)

Itoh, Nobuyuki

2016-04-01

The FGF family comprises 22 members with diverse functions in development and health. FGF10 specifically activates FGFR2b in a paracrine manner with heparan sulfate as a co-factor. FGF10and FGFR2b are preferentially expressed in the mesenchyme and epithelium, respectively. FGF10 is a mesenchymal signaling molecule in the epithelium. FGF10 knockout mice die shortly after birth due to the complete absence of lungs as well as fore- and hindlimbs. FGF10 is also essential for the development of multiple organs. The phenotypes of Fgf10 knockout mice are very similar to those of FGFR2b knockout mice, indicating that FGF10 acts as a ligand that is specific to FGFR2b in mouse multi-organ development. FGF10 also plays roles in epithelial-mesenchymal transition, the repair of tissue injury, and embryonic stem cell differentiation. In humans, FGF10 loss-of-function mutations result in inherited diseases including aplasia of lacrimal and salivary gland, lacrimo-auriculo-dento-digital syndrome, and chronic obstructive pulmonary disease. FGF10 is also involved in the oncogenicity of pancreatic and breast cancers. Single nucleotide polymorphisms in FGF10 are also potential risk factors for limb deficiencies, cleft lip and palate, and extreme myopia. These findings indicate that FGF10 is a crucial paracrine signal from the mesenchyme to epithelium for development, health, and disease. Copyright © 2015. Published by Elsevier Ltd.

Identification of potential pathway mediation targets in Toll-like receptor signaling.

Directory of Open Access Journals (Sweden)

Fan Li

2009-02-01

Full Text Available Recent advances in reconstruction and analytical methods for signaling networks have spurred the development of large-scale models that incorporate fully functional and biologically relevant features. An extended reconstruction of the human Toll-like receptor signaling network is presented herein. This reconstruction contains an extensive complement of kinases, phosphatases, and other associated proteins that mediate the signaling cascade along with a delineation of their associated chemical reactions. A computational framework based on the methods of large-scale convex analysis was developed and applied to this network to characterize input-output relationships. The input-output relationships enabled significant modularization of the network into ten pathways. The analysis identified potential candidates for inhibitory mediation of TLR signaling with respect to their specificity and potency. Subsequently, we were able to identify eight novel inhibition targets through constraint-based modeling methods. The results of this study are expected to yield meaningful avenues for further research in the task of mediating the Toll-like receptor signaling network and its effects.

The Hrs/Stam complex acts as a positive and negative regulator of RTK signaling during Drosophila development.

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Hélène Chanut-Delalande

Full Text Available BACKGROUND: Endocytosis is a key regulatory step of diverse signalling pathways, including receptor tyrosine kinase (RTK signalling. Hrs and Stam constitute the ESCRT-0 complex that controls the initial selection of ubiquitinated proteins, which will subsequently be degraded in lysosomes. It has been well established ex vivo and during Drosophila embryogenesis that Hrs promotes EGFR down regulation. We have recently isolated the first mutations of stam in flies and shown that Stam is required for air sac morphogenesis, a larval respiratory structure whose formation critically depends on finely tuned levels of FGFR activity. This suggest that Stam, putatively within the ESCRT-0 complex, modulates FGF signalling, a possibility that has not been examined in Drosophila yet. PRINCIPAL FINDINGS: Here, we assessed the role of the Hrs/Stam complex in the regulation of signalling activity during Drosophila development. We show that stam and hrs are required for efficient FGFR signalling in the tracheal system, both during cell migration in the air sac primordium and during the formation of fine cytoplasmic extensions in terminal cells. We find that stam and hrs mutant cells display altered FGFR/Btl localisation, likely contributing to impaired signalling levels. Electron microscopy analyses indicate that endosome maturation is impaired at distinct steps by hrs and stam mutations. These somewhat unexpected results prompted us to further explore the function of stam and hrs in EGFR signalling. We show that while stam and hrs together downregulate EGFR signalling in the embryo, they are required for full activation of EGFR signalling during wing development. CONCLUSIONS/SIGNIFICANCE: Our study shows that the ESCRT-0 complex differentially regulates RTK signalling, either positively or negatively depending on tissues and developmental stages, further highlighting the importance of endocytosis in modulating signalling pathways during development.

Fibroblast Growth Factor Receptor 3 (FGFR3–Analyses of the S249C Mutation and Protein Expression in Primary Cervical Carcinomas

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Haiyan Dai

2001-01-01

Full Text Available Fibroblast growth factor receptor 3 (FGFR3 seems to play an inhibitory role in bone development, as activating mutations in the gene underlie disorders such as achondroplasia and thanatophoric dysplasia. Findings from multiple myeloma (MM indicate that FGFR3 also can act as an oncogene, and mutation of codon 249 in the fibroblast growth factor receptor 3 (FGFR3 gene was recently detected in 3/12 primary cervical carcinomas. We have analysed 91 cervical carcinomas for this specific S249C mutation using amplification created restriction site methodology (ACRS, and detected no mutations. Immunohistochemistry was performed on 73 of the tumours. Reduced protein staining was seen in 43 (58.8% samples. Six of the tumours (8.2% revealed increased protein staining compared with normal cervical tissue. These patients had a better prognosis than those with reduced or normal levels, although not statistically significant. This report weakens the hypothesis of FGFR3 as an oncogene of importance in cervical carcinomas.

Fine-Scale Mapping of the FGFR2 Breast Cancer Risk Locus

DEFF Research Database (Denmark)

Meyer, Kerstin B; O'Reilly, Martin; Michailidou, Kyriaki

2013-01-01

The 10q26 locus in the second intron of FGFR2 is the locus most strongly associated with estrogen-receptor-positive breast cancer in genome-wide association studies. We conducted fine-scale mapping in case-control studies genotyped with a custom chip (iCOGS), comprising 41 studies (n = 89,050) of...

Neural cell adhesion molecule induces intracellular signaling via multiple mechanisms of Ca2+ homeostasis

DEFF Research Database (Denmark)

Kiryushko, Darya; Korshunova, Irina; Berezin, Vladimir

2006-01-01

. The first pathway was associated with activation of FGFR, phospholipase Cgamma, and production of diacylglycerol, and the second pathway involved Src-family kinases. Moreover, NCAM-mediated Ca2+ entry required activation of nonselective cation and T-type voltage-gated Ca2+ channels. These channels, together...

Acanthosis nigricans in a Japanese boy with hypochondroplasia due to a K650T mutation in FGFR3

Science.gov (United States)

Hirai, Hiroki; Hamada, Junpei; Hasegawa, Kosei; Ishii, Eiichi

2017-01-01

Abstract. Acanthosis nigricans (AN) is observed in some cases of skeletal dysplasia. However, AN has occasionally been reported in patients with hypochondroplasia (HCH), and a clinical diagnosis is sometimes difficult when its physical and radiological features are mild. Mutations in the gene encoding the fibroblast growth factor receptor 3 (FGFR3) have been identified as the cause of some types of skeletal dysplasia, which is diagnostically useful. Here, we report the case of a 3-yr-old Japanese boy who presented with AN. His height, weight, head circumference, and arm span were 91.7 cm (–1.95 SD), 16.3 kg, 54.0 cm (+2.6 SD), and 88.0 cm, respectively. In addition to the AN, he also exhibited a mild height deficit and macrocephaly, which prompted a search for FGFR3 mutations, although no skeletal disproportion, exaggerated lumbar lordosis, or facial dysmorphism was observed, and only slight radiological abnormalities were noted. A definitive diagnosis of HCH was made based on FGFR3 gene analysis, which detected a heterozygous K650T mutation. Insulin insensitivity was not found to have contributed to the development of AN. In individuals with AN, careful assessments for symptoms of HCH are important, regardless of the presence or absence of a short stature, and FGFR3 gene analysis is recommended in such cases. PMID:29026271

Fibroblast growth factor receptor 1 activation in mammary tumor cells promotes macrophage recruitment in a CX3CL1-dependent manner.

Directory of Open Access Journals (Sweden)

Johanna R Reed

Full Text Available Tumor formation is an extensive process requiring complex interactions that involve both tumor cell-intrinsic pathways and soluble mediators within the microenvironment. Tumor cells exploit the intrinsic functions of many soluble molecules, including chemokines and their receptors, to regulate pro-tumorigenic phenotypes that are required for growth and progression of the primary tumor. Previous studies have shown that activation of inducible FGFR1 (iFGFR1 in mammary epithelial cells resulted in increased proliferation, migration, and invasion in vitro and tumor formation in vivo. These studies also demonstrated that iFGFR1 activation stimulated recruitment of macrophages to the epithelium where macrophages contributed to iFGFR1-mediated epithelial cell proliferation and angiogenesis. The studies presented here further utilize this model to identify the mechanisms that regulate FGFR1-induced macrophage recruitment. Results from this study elucidate a novel role for the inflammatory chemokine CX3CL1 in FGFR1-induced macrophage migration. Specifically, we illustrate that activation of both the inducible FGFR1 construct in mouse mammary epithelial cells and endogenous FGFR in the triple negative breast cancer cell line, HS578T, leads to expression of the chemokine CX3CL1. Furthermore, we demonstrate that FGFR-induced CX3CL1 is sufficient to recruit CX3CR1-expressing macrophages in vitro. Finally, blocking CX3CR1 in vivo leads to decreased iFGFR1-induced macrophage recruitment, which correlates with decreased angiogenesis. While CX3CL1 is a known target of FGF signaling in the wound healing environment, these studies demonstrate that FGFR activation also leads to induction of CX3CL1 in a tumor setting. Furthermore, these results define a novel role for CX3CL1 in promoting macrophage recruitment during mammary tumor formation, suggesting that the CX3CL1/CX3CR1 axis may represent a potential therapeutic approach for targeting breast cancers associated

DMPD: Signalling pathways mediating type I interferon gene expression. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 17904888 Signalling pathways mediating type I interferon gene expression. Edwards M...hways mediating type I interferon gene expression. PubmedID 17904888 Title Signalling pathways...R, Slater L, Johnston SL. Microbes Infect. 2007 Sep;9(11):1245-51. Epub 2007 Jul 1. (.png) (.svg) (.html) (.csml) Show Signalling pat

Plant cell surface receptor-mediated signaling - a common theme amid diversity.

Science.gov (United States)

He, Yunxia; Zhou, Jinggeng; Shan, Libo; Meng, Xiangzong

2018-01-29

Sessile plants employ a diverse array of plasma membrane-bound receptors to perceive endogenous and exogenous signals for regulation of plant growth, development and immunity. These cell surface receptors include receptor-like kinases (RLKs) and receptor-like proteins (RLPs) that harbor different extracellular domains for perception of distinct ligands. Several RLK and RLP signaling pathways converge at the somatic embryogenesis receptor kinases (SERKs), which function as shared co-receptors. A repertoire of receptor-like cytoplasmic kinases (RLCKs) associate with the receptor complexes to relay intracellular signaling. Downstream of the receptor complexes, mitogen-activated protein kinase (MAPK) cascades are among the key signaling modules at which the signals converge, and these cascades regulate diverse cellular and physiological responses through phosphorylation of different downstream substrates. In this Review, we summarize the emerging common theme that underlies cell surface receptor-mediated signaling pathways in Arabidopsis thaliana : the dynamic association of RLKs and RLPs with specific co-receptors and RLCKs for signal transduction. We further discuss how signaling specificities are maintained through modules at which signals converge, with a focus on SERK-mediated receptor signaling. © 2018. Published by The Company of Biologists Ltd.

Common mutations in the fibroblast growth factor receptor 3 (FGFR 3) gene account for achondroplasia, hypochondroplasia, and thanatophoric dwarfism

Energy Technology Data Exchange (ETDEWEB)

Bonaventure, J.; Rousseau, F.; Legeai-Mallet, L.; LeMerrer, M.; Munnich, A.; Maroteaux, P. [INSERM, Paris (France)

1996-05-03

The mapping of the achondroplasia locus to the short arm of chromosome 4 and the subsequent identification of a recurrent missense mutation (G380R) in the fibroblast growth factor receptor 3 (FGFR-3) gene has been followed by the detection of common FGFR-3 mutations in two clinically related disorders: thanatophoric dwarfism (types I and II) and hypochondroplasia. The relative clinical homogeneity of achondroplasia was substantiated by demonstration of its genetic homogeneity as more than 98% of all patients hitherto reported exhibit mutations in the transmembrane receptor domain. Although most hypochondroplasia cases were accounted for by a recurrent missense substitution (N540K) in the first tyrosine kinase (TK 1) domain of the receptor, a significant proportion (40%) of our patients did not harbor the N540K mutation and three hypochondroplasia families were not linked to the FGFR-3 locus, thus supporting clinical heterogeneity of this condition. In thanatophoric dwarfism (TD), a recurrent FGFR-3 mutation located in the second tyrosine kinase (TK 2) domain of the receptor was originally detected in 100% of TD II cases; in our series, seven distinct mutations in three different protein domains were identified in 25 of 26 TD I patients, suggesting that TD, like achondroplasia, is a genetically homogenous skeletal disorder. 31 refs., 4 figs., 2 tabs.

NMR structure of the first Ig module of mouse FGFR1

DEFF Research Database (Denmark)

Kiselyov, V.V.; Bock, Elisabeth Marianne; Berezin, V.

2006-01-01

of this module. We describe here the NMR structure of the Ig1 module of mouse FGFR1. The three-dimensional fold of the module belongs to the intermediate Ig subgroup and can be described as a beta-barrel consisting of two beta-sheets. One sheet is formed by A', G, F, C, and C', and the other by A, B, B', E...

Molecular grading of tumors of the upper urothelial tract using FGFR3 mutation status identifies patients with favorable prognosis

OpenAIRE

Fernandez, Cecilia; Lyle,Stephen; Hsieh,; Shuber,Anthony

2012-01-01

Stephen R Lyle,1 Chung-Cheng Hsieh,1 Cecilia A Fernandez,2 Anthony P Shuber21University of Massachusetts, Worcester, MA, 2Predictive Biosciences Inc., Lexington, MA, USABackground: Mutations in FGFR3 have been shown to occur in tumors of the upper urothelial tract and may be indicative of a good prognosis. In bladder tumors, the combination of FGFR3 mutation status and Ki-67 level has been used to define a tumor's molecular grade and predict survival. Pathological evaluation of upper ...

The mediator complex in genomic and non-genomic signaling in cancer.

Science.gov (United States)

Weber, Hannah; Garabedian, Michael J

2018-05-01

Mediator is a conserved, multi-subunit macromolecular machine divided structurally into head, middle, and tail modules, along with a transiently associating kinase module. Mediator functions as an integrator of transcriptional regulatory activity by interacting with DNA-bound transcription factors and with RNA polymerase II (Pol II) to both activate and repress gene expression. Mediator has been shown to affect multiple steps in transcription, including chromatin looping between enhancers and promoters, pre-initiation complex formation, transcriptional elongation, and mRNA splicing. Individual Mediator subunits participate in regulation of gene expression by the estrogen and androgen receptors and are altered in a number of endocrine cancers, including breast and prostate cancer. In addition to its role in genomic signaling, MED12 has been implicated in non-genomic signaling by interacting with and activating TGF-beta receptor 2 in the cytoplasm. Recent structural studies have revealed extensive inter-domain interactions and complex architecture of the Mediator-Pol II complex, suggesting that Mediator is capable of reorganizing its conformation and composition to fit cellular needs. We propose that alterations in Mediator subunit expression that occur in various cancers could impact the organization and function of Mediator, resulting in changes in gene expression that promote malignancy. A better understanding of the role of Mediator in cancer could reveal new approaches to the diagnosis and treatment of Mediator-dependent endocrine cancers, especially in settings of therapy resistance. Copyright © 2017 Elsevier Inc. All rights reserved.

Allergen-Induced Dermatitis Causes Alterations in Cutaneous Retinoid-Mediated Signaling in Mice

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Gericke, Janine; Ittensohn, Jan; Mihály, Johanna; Dubrac, Sandrine; Rühl, Ralph

2013-01-01

Nuclear receptor-mediated signaling via RARs and PPARδ is involved in the regulation of skin homeostasis. Moreover, activation of both RAR and PPARδ was shown to alter skin inflammation. Endogenous all-trans retinoic acid (ATRA) can activate both receptors depending on specific transport proteins: Fabp5 initiates PPARδ signaling whereas Crabp2 promotes RAR signaling. Repetitive topical applications of ovalbumin (OVA) in combination with intraperitoneal injections of OVA or only intraperitoneal OVA applications were used to induce allergic dermatitis. In our mouse model, expression of IL-4, and Hbegf increased whereas expression of involucrin, Abca12 and Spink5 decreased in inflamed skin, demonstrating altered immune response and epidermal barrier homeostasis. Comprehensive gene expression analysis showed alterations of the cutaneous retinoid metabolism and retinoid-mediated signaling in allergic skin immune response. Notably, ATRA synthesis was increased as indicated by the elevated expression of retinaldehyde dehydrogenases and increased levels of ATRA. Consequently, the expression pattern of genes downstream to RAR was altered. Furthermore, the increased ratio of Fabp5 vs. Crabp2 may indicate an up-regulation of the PPARδ pathway in allergen-induced dermatitis in addition to the altered RAR signaling. Thus, our findings suggest that ATRA levels, RAR-mediated signaling and signaling involved in PPARδ pathways are mainly increased in allergen-induced dermatitis and may contribute to the development and/or maintenance of allergic skin diseases. PMID:23977003

Key mediators of intracellular amino acids signaling to mTORC1 activation.

Science.gov (United States)

Duan, Yehui; Li, Fengna; Tan, Kunrong; Liu, Hongnan; Li, Yinghui; Liu, Yingying; Kong, Xiangfeng; Tang, Yulong; Wu, Guoyao; Yin, Yulong

2015-05-01

Mammalian target of rapamycin complex 1 (mTORC1) is activated by amino acids to promote cell growth via protein synthesis. Specifically, Ras-related guanosine triphosphatases (Rag GTPases) are activated by amino acids, and then translocate mTORC1 to the surface of late endosomes and lysosomes. Ras homolog enriched in brain (Rheb) resides on this surface and directly activates mTORC1. Apart from the presence of intracellular amino acids, Rag GTPases and Rheb, other mediators involved in intracellular amino acid signaling to mTORC1 activation include human vacuolar sorting protein-34 (hVps34) and mitogen-activating protein kinase kinase kinase kinase-3 (MAP4K3). Those molecular links between mTORC1 and its mediators form a complicate signaling network that controls cellular growth, proliferation, and metabolism. Moreover, it is speculated that amino acid signaling to mTORC1 may start from the lysosomal lumen. In this review, we discussed the function of these mediators in mTORC1 pathway and how these mediators are regulated by amino acids in details.

Lunatic fringe-mediated Notch signaling regulates adult hippocampal neural stem cell maintenance.

Science.gov (United States)

Semerci, Fatih; Choi, William Tin-Shing; Bajic, Aleksandar; Thakkar, Aarohi; Encinas, Juan Manuel; Depreux, Frederic; Segil, Neil; Groves, Andrew K; Maletic-Savatic, Mirjana

2017-07-12

Hippocampal neural stem cells (NSCs) integrate inputs from multiple sources to balance quiescence and activation. Notch signaling plays a key role during this process. Here, we report that Lunatic fringe ( Lfng), a key modifier of the Notch receptor, is selectively expressed in NSCs. Further, Lfng in NSCs and Notch ligands Delta1 and Jagged1, expressed by their progeny, together influence NSC recruitment, cell cycle duration, and terminal fate. We propose a new model in which Lfng-mediated Notch signaling enables direct communication between a NSC and its descendants, so that progeny can send feedback signals to the 'mother' cell to modify its cell cycle status. Lfng-mediated Notch signaling appears to be a key factor governing NSC quiescence, division, and fate.

Porcine pluripotency cell signaling develops from the inner cell mass to the epiblast during early development

DEFF Research Database (Denmark)

Hall, Vanessa Jane; Christensen, Josef; Gao, Yu

2009-01-01

  The signaling mechanisms regulating pluripotency in porcine embryonic stem cells and embryos are unknown. In this study, we characterize cell signaling in the in-vivo porcine inner cell mass and later-stage epiblast. We evaluate expression of OCT4, NANOG, SOX2, genes within the JAK/STAT pathway...... pluripotency in human embryonic stem cells is detectable in the porcine epiblast, but not in the inner cell mass. Copyright (c) 2009 Wiley-Liss, Inc.......  The signaling mechanisms regulating pluripotency in porcine embryonic stem cells and embryos are unknown. In this study, we characterize cell signaling in the in-vivo porcine inner cell mass and later-stage epiblast. We evaluate expression of OCT4, NANOG, SOX2, genes within the JAK/STAT pathway...... (LIF, LIFR, GP130), FGF pathway (bFGF, FGFR1, FGFR2), BMP pathway (BMP4), and downstream-activated genes (STAT3, c-Myc, c-Fos, and SMAD4). We discovered two different expression profiles exist in the developing porcine embryo. The D6 porcine blastocyst (inner cell mass stage) is devoid...

Root signals that mediate mutualistic interactions in the rhizosphere.

Science.gov (United States)

Rasmann, Sergio; Turlings, Ted Cj

2016-08-01

A recent boom in research on belowground ecology is rapidly revealing a multitude of fascinating interactions, in particular in the rhizosphere. Many of these interactions are mediated by photo-assimilates that are excreted by plant roots. Root exudates are not mere waste products, but serve numerous functions to control abiotic and biotic processes. These functions range from changing the chemical and physical properties of the soil, inhibiting the growth of competing plants, combatting herbivores, and regulating the microbial community. Particularly intriguing are root-released compounds that have evolved to serve mutualistic interactions with soil-dwelling organisms. These mutually beneficial plant-mediated signals are not only of fundamental ecological interest, but also exceedingly important from an agronomical perspective. Here, we attempt to provide an overview of the plant-produced compounds that have so far been implicated in mutualistic interactions. We propose that these mutualistic signals may have evolved from chemical defenses and we point out that they can be (mis)used by specialized pathogens and herbivores. We speculate that many more signals and interactions remain to be uncovered and that a good understanding of the mechanisms and ecological implications can be the basis for exploitation and manipulation of the signals for crop improvement and protection. Copyright © 2016 Elsevier Ltd. All rights reserved.

Kallmann syndrome: 14 novel mutations in KAL1 and FGFR1 (KAL2).

Science.gov (United States)

Albuisson, Juliette; Pêcheux, Chistophe; Carel, Jean-Claude; Lacombe, Didier; Leheup, Bruno; Lapuzina, Pablo; Bouchard, Philippe; Legius, Eric; Matthijs, Gert; Wasniewska, Malgorzata; Delpech, Marc; Young, Jacques; Hardelin, Jean-Pierre; Dodé, Catherine

2005-01-01

Kallmann syndrome (KAL) combines hypogonadotropic hypogonadism and anosmia. Hypogonadism is due to Gonadotropin Releasing Hormone (GnRH) deficiency and anosmia is related to hypoplasia of the olfactory bulbs. Occasional symptoms include renal agenesis, bimanual synkinesia, cleft lip palate, dental agenesis. KAL is genetically heterogeneous and two genes have so far been identified, namely KAL1 (Xp22.3) and FGFR1/KAL2 (8p12), which underlie the X chromosome-linked form and an autosomal dominant form of the disease, respectively. We studied a cohort of 98 unrelated Caucasian KAL patients. We identified KAL1 mutations in 14 patients, of which 7 (c.3G>A (p.M1?), g.IVS1+1G>T, c.570_571insA (p.R191fsX14), c.784G>C (p.R262P), c.958G>T (p.E320X), c.1651_1654delinsAGCT (p.P551_E552delinsSX), c.1711T>A (p.W571R)) have not been previously reported. In addition, we found FGFR1 mutations in 7 patients, namely c.303G>A (p.V102I), C.385A>C (p.D129A), c.810G>A (p.V273M), c.1093_1094delAG (p.R365fsX41), c.1561G>A (p.A520T), c.1836_1837insT (p.Y613fsX42), c.2190C>G (p.Y730X), all of which were novel mutations. In this study, unilateral renal agenesis and bimanual synkinesia were exclusively found associated with KAL1mutations, cleft palate and dental agenesia with FGFR1mutations. (c) 2004 Wiley-Liss, Inc.

Nitric oxide-mediated bystander signal transduction induced by heavy-ion microbeam irradiation

Science.gov (United States)

Tomita, Masanori; Matsumoto, Hideki; Funayama, Tomoo; Yokota, Yuichiro; Otsuka, Kensuke; Maeda, Munetoshi; Kobayashi, Yasuhiko

2015-07-01

In general, a radiation-induced bystander response is known to be a cellular response induced in non-irradiated cells after receiving bystander signaling factors released from directly irradiated cells within a cell population. Bystander responses induced by high-linear energy transfer (LET) heavy ions at low fluence are an important health problem for astronauts in space. Bystander responses are mediated via physical cell-cell contact, such as gap-junction intercellular communication (GJIC) and/or diffusive factors released into the medium in cell culture conditions. Nitric oxide (NO) is a well-known major initiator/mediator of intercellular signaling within culture medium during bystander responses. In this study, we investigated the NO-mediated bystander signal transduction induced by high-LET argon (Ar)-ion microbeam irradiation of normal human fibroblasts. Foci formation by DNA double-strand break repair proteins was induced in non-irradiated cells, which were co-cultured with those irradiated by high-LET Ar-ion microbeams in the same culture plate. Foci formation was suppressed significantly by pretreatment with an NO scavenger. Furthermore, NO-mediated reproductive cell death was also induced in bystander cells. Phosphorylation of NF-κB and Akt were induced during NO-mediated bystander signaling in the irradiated and bystander cells. However, the activation of these proteins depended on the incubation time after irradiation. The accumulation of cyclooxygenase-2 (COX-2), a downstream target of NO and NF-κB, was observed in the bystander cells 6 h after irradiation but not in the directly irradiated cells. Our findings suggest that Akt- and NF-κB-dependent signaling pathways involving COX-2 play important roles in NO-mediated high-LET heavy-ion-induced bystander responses. In addition, COX-2 may be used as a molecular marker of high-LET heavy-ion-induced bystander cells to distinguish them from directly irradiated cells, although this may depend on the time

A 3-plex methylation assay combined with the FGFR3 mutation assay sensitively detects recurrent bladder cancer in voided urine

DEFF Research Database (Denmark)

Kandimalla, Raju; Masius, Roy; Beukers, Willemien

2013-01-01

is to determine the sensitivity and specificity of a urine assay for the diagnosis of recurrences in patients with a previous primary NMIBC G1/G2 by using cystoscopy as the reference standard. Experimental Design: We selected eight CpG islands (CGI) methylated in bladder cancer from our earlier genome-wide study......Purpose: DNA methylation is associated with bladder cancer and these modifications could serve as useful biomarkers. FGFR3 mutations are present in 60% to 70% of non–muscle invasive bladder cancer (NMIBC). Low-grade bladder cancer recurs in more than 50% of patients. The aim of this study......, and nonmalignant urines (n = 130). Results: The 3-plex assay identified recurrent bladder cancer in voided urine with a sensitivity of 74% in the validation set. In combination with the FGFR3 mutation assay, a sensitivity of 79% was reached (specificity of 77%). Sensitivity of FGFR3 and cytology was 52% and 57...

Inositol trisphosphate receptor mediated spatiotemporal calcium signalling.

Science.gov (United States)

Miyazaki, S

1995-04-01

Spatiotemporal Ca2+ signalling in the cytoplasm is currently understood as an excitation phenomenon by analogy with electrical excitation in the plasma membrane. In many cell types, Ca2+ waves and Ca2+ oscillations are mediated by inositol 1,4,5-trisphosphate (IP3) receptor/Ca2+ channels in the endoplasmic reticulum membrane, with positive feedback between cytosolic Ca2+ and IP3-induced Ca2+ release creating a regenerative process. Remarkable advances have been made in the past year in the analysis of subcellular Ca2+ microdomains using confocal microscopy and of Ca2+ influx pathways that are functionally coupled to IP3-induced Ca2+ release. Ca2+ signals can be conveyed into the nucleus and mitochondria. Ca2+ entry from outside the cell allows repetitive Ca2+ release by providing Ca2+ to refill the endoplasmic reticulum stores, thus giving rise to frequency-encoded Ca2+ signals.

DMPD: Modulation of Toll-interleukin 1 receptor mediated signaling. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 15662540 Modulation of Toll-interleukin 1 receptor mediated signaling. Li X, Qin J.... J Mol Med. 2005 Apr;83(4):258-66. Epub 2005 Jan 21. (.png) (.svg) (.html) (.csml) Show Modulation of Toll-i...nterleukin 1 receptor mediated signaling. PubmedID 15662540 Title Modulation of Toll-interleukin 1 receptor

Single-cell analysis reveals a link between CD3- and CD59-mediated signaling pathways in Jurkat T cells

International Nuclear Information System (INIS)

Lipp, A. M.

2012-01-01

Elevation of intracellular free calcium concentration ([Ca2+]i) is a key signal during T cell activation and is commonly used as a read-out parameter for stimulation of T cell signaling. Upon T cell stimulation a variety of calcium signals is produced by individual cells of the T cell population and the type of calcium signal strongly influences cell fate decisions. The heterogeneous nature of T cells is masked in ensemble measurements, which highlights the need for single-cell measurements. In this study we used single-cell calcium measurements in Jurkat cells to investigate signaling pathways, which are triggered by different proteins, namely CD3 and CD59. By application of an automated cluster algorithm the presented assay provides unbiased analysis of a large data set of individual calcium time traces generated by the whole cell population. By using this method we could demonstrate that the Jurkat population generates heterogeneous calcium signals in a stimulus-dependent manner. Furthermore, our data revealed the existence of a link between CD3- and CD59-mediated signaling pathways. Single-cell calcium measurements in Jurkat cells expressing different levels of the T cell receptor (TCR) complex indicated that CD59-mediated calcium signaling is critically dependent on TCR surface expression levels. In addition, triggering CD59-mediated calcium signaling resulted in down-regulation of TCR surface expression levels, which is known to happen upon direct TCR triggering too. Moreover, by using siRNA-mediated protein knock-downs and protein knock-out Jurkat mutants we could show that CD3- and CD59-mediated calcium signaling require identical key proteins. We therefore explored by which mechanism CD59-mediated signaling couples into TCR-mediated signaling. Fluorescence recovery after photobleaching (FRAP) experiments and live-cell protein-protein interaction assays provided no evidence of a direct physical interaction between CD3- and CD59-mediated signaling pathways

Design, synthesis, and biological evaluation of 3-vinyl-quinoxalin-2(1H-one derivatives as novel antitumor inhibitors of FGFR1

Directory of Open Access Journals (Sweden)

Liu Z

2016-05-01

Full Text Available Zhiguo Liu,1,* Shufang Yu,1,* Di Chen,1 Guoliang Shen,1 Yu Wang,1 Leping Hou,2 Dan Lin,1 Jinsan Zhang,1 Faqing Ye1 1School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, 2Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, People’s Republic of China *These authors contributed equally to this work Abstract: FGFR1 is well known as a molecular target in anticancer drug design. TKI258 plays an important role in RTK inhibitors. Utilizing TKI258 as a lead compound that contains a quinazolinone nucleus, we synthesized four series of 3-vinyl-quinoxalin-2(1H-one derivatives, a total of 27 compounds. We further evaluated these compounds for FGFR1 inhibition ability as well as cytotoxicity against four cancer cell lines (H460, B16-F10, Hela229, and Hct116 in vitro. Some compounds displayed good-to-excellent potency against the four tested cancer cell lines compared with TKI258. Structure–activity relationship analyses indicated that small substituents at the side chain of the 3-vinyl-quinoxalin-2(1H-one were more effective than large substituents. Lastly, we used molecular docking to obtain further insight into the interactions between the compounds and FGFR1. Keywords: FGFR1, synthesis, quinoxaline, antitumor activity, kinase inhibitor

The Role of Cgrp-Receptor Component Protein (Rcp in Cgrp-Mediated Signal Transduction

Directory of Open Access Journals (Sweden)

M. A. Prado

2001-01-01

Full Text Available The calcitonin gene-related peptide (CGRP-receptor component protein (RCP is a 17-kDa intracellular peripheral membrane protein required for signal transduction at CGRP receptors. To determine the role of RCP in CGRP-mediated signal transduction, RCP was depleted from NIH3T3 cells using antisense strategy. Loss of RCP protein correlated with loss of cAMP production by CGRP in the antisense cells. In contrast, loss of RCP had no effect on CGRP-mediated binding; therefore RCP is not acting as a chaperone for the CGRP receptor. Instead, RCP is a novel signal transduction molecule that couples the CGRP receptor to the cellular signal transduction machinery. RCP thus represents a prototype for a new class of signal transduction proteins that are required for regulation of G protein-coupled receptors.

A common FGFR3 gene mutation is present in achondroplasia but not in hypochondroplasia

Energy Technology Data Exchange (ETDEWEB)

Stoilov, I.; Kilpatrick, M.W.; Tsipouras, P. [Univ. of Connecticut Health Center, Farmington, CT (United States)

1995-01-02

Achondroplasia is the most common type of genetic dwarfism. It is characterized by disproportionate short stature and other skeletal anomalies resulting from a defect in the maturation of the chondrocytes in the growth plate of the cartilage. Recent studies mapped the achondroplasia gene on chromosome region 4p16.3 and identified a common mutation in the gene encoding the fibroblast growth factor receptor 3 (FGFR3). In an analysis of 19 achondroplasia families from a variety of ethnic backgrounds we confirmed the presence of the G380R mutation in 21 of 23 achondroplasia chromosomes studied. In contrast, the G380R mutation was not found in any of the 8 hypochondroplasia chromosomes studied. Futhermore, linkage studies in a 3-generation family with hypochondroplasia show discordant segregation with markers in the 4p16.3 region suggesting that at least some cases of hypochondroplasia are caused by mutations in a gene other than FGFR3. 27 refs., 2 figs.

Calcium-mediated signaling and calmodulin-dependent kinase regulate hepatocyte-inducible nitric oxide synthase expression.

Science.gov (United States)

Zhang, Baochun; Crankshaw, Will; Nesemeier, Ryan; Patel, Jay; Nweze, Ikenna; Lakshmanan, Jaganathan; Harbrecht, Brian G

2015-02-01

Induced nitric oxide synthase (iNOS) is induced in hepatocytes by shock and inflammatory stimuli. Excessive NO from iNOS mediates shock-induced hepatic injury and death, so understanding the regulation of iNOS will help elucidate the pathophysiology of septic shock. In vitro, cytokines induce iNOS expression through activation of signaling pathways including mitogen-activated protein kinases and nuclear factor κB. Cytokines also induce calcium (Ca(2+)) mobilization and activate calcium-mediated intracellular signaling pathways, typically through activation of calmodulin-dependent kinases (CaMK). Calcium regulates NO production in macrophages but the role of calcium and calcium-mediated signaling in hepatocyte iNOS expression has not been defined. Primary rat hepatocytes were isolated, cultured, and induced to produce NO with proinflammatory cytokines. Calcium mobilization and Ca(2+)-mediated signaling were altered with ionophore, Ca(2+) channel blockers, and inhibitors of CaMK. The Ca(2+) ionophore A23187 suppressed cytokine-stimulated NO production, whereas Ethylene glycol tetraacetic acid and nifedipine increased NO production, iNOS messenger RNA, and iNOS protein expression. Inhibition of CaMK with KN93 and CBD increased NO production but the calcineurin inhibitor FK 506 decreased iNOS expression. These data demonstrate that calcium-mediated signaling regulates hepatocyte iNOS expression and does so through a mechanism independent of calcineurin. Changes in intracellular calcium levels may regulate iNOS expression during hepatic inflammation induced by proinflammatory cytokines. Copyright © 2015 Elsevier Inc. All rights reserved.

Statins do not inhibit the FGFR signaling in chondrocytes

Czech Academy of Sciences Publication Activity Database

Fafílek, B.; Hampl, Marek; Říčánková, N.; Veselá, Iva; Bálek, L.; Kunová Bosáková, M.; Gudernová, I.; Vařecha, M.; Buchtová, Marcela; Krejčí, P.

2017-01-01

Roč. 25, č. 9 (2017), s. 1522-1530 ISSN 1063-4584 R&D Projects: GA ČR(CZ) GA14-31540S Grant - others:GA MŠk(CZ) LH12004 Institutional support: RVO:67985904 Keywords : statins * FGF signaling * chondrocytes Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Developmental biology Impact factor: 4.742, year: 2016

A de novo missense mutation of FGFR2 causes facial dysplasia syndrome in Holstein cattle.

Science.gov (United States)

Agerholm, Jørgen S; McEvoy, Fintan J; Heegaard, Steffen; Charlier, Carole; Jagannathan, Vidhya; Drögemüller, Cord

2017-08-02

Surveillance for bovine genetic diseases in Denmark identified a hitherto unreported congenital syndrome occurring among progeny of a Holstein sire used for artificial breeding. A genetic aetiology due to a dominant inheritance with incomplete penetrance or a mosaic germline mutation was suspected as all recorded cases were progeny of the same sire. Detailed investigations were performed to characterize the syndrome and to reveal its cause. Seven malformed calves were submitted examination. All cases shared a common morphology with the most striking lesions being severe facial dysplasia and complete prolapse of the eyes. Consequently the syndrome was named facial dysplasia syndrome (FDS). Furthermore, extensive brain malformations, including microencephaly, hydrocephalus, lobation of the cerebral hemispheres and compression of the brain were present. Subsequent data analysis of progeny of the sire revealed that around 0.5% of his offspring suffered from FDS. High density single nucleotide polymorphism (SNP) genotyping data of the seven cases and their parents were used to map the defect in the bovine genome. Significant genetic linkage was obtained for three regions, including chromosome 26 where whole genome sequencing of a case-parent trio revealed two de novo variants perfectly associated with the disease: an intronic SNP in the DMBT1 gene and a single non-synonymous variant in the FGFR2 gene. This FGFR2 missense variant (c.927G>T) affects a gene encoding a member of the fibroblast growth factor receptor family, where amino acid sequence is highly conserved between members and across species. It is predicted to change an evolutionary conserved tryptophan into a cysteine residue (p.Trp309Cys). Both variant alleles were proven to result from de novo mutation events in the germline of the sire. FDS is a novel genetic disorder of Holstein cattle. Mutations in the human FGFR2 gene are associated with various dominant inherited craniofacial dysostosis syndromes. Given

Role of CD137 signaling in dengue virus-mediated apoptosis

International Nuclear Information System (INIS)

Nagila, Amar; Netsawang, Janjuree; Srisawat, Chatchawan; Noisakran, Sansanee; Morchang, Atthapan; Yasamut, Umpa; Puttikhunt, Chunya; Kasinrerk, Watchara

2011-01-01

Highlights: → For the first time the role of CD137 in dengue virus (DENV) infection. → Induction of DENV-mediated apoptosis by CD137 signaling. → Sensitization to CD137-mediated apoptosis by dengue virus capsid protein (DENV C). → Nuclear localization of DENV C is required for CD137-mediated apoptosis. -- Abstract: Hepatic dysfunction is a well recognized feature of dengue virus (DENV) infection. However, molecular mechanisms of hepatic injury are still poorly understood. A complex interaction between DENV and the host immune response contributes to DENV-mediated tissue injury. DENV capsid protein (DENV C) physically interacts with the human death domain-associated protein Daxx. A double substitution mutation in DENV C (R85A/K86A) abrogates Daxx interaction, nuclear localization and apoptosis. Therefore we compared the expression of cell death genes between HepG2 cells expressing DENV C and DENV C (R85A/K86A) using a real-time PCR array. Expression of CD137, which is a member of the tumor necrosis factor receptor family, increased significantly in HepG2 cells expressing DENV C compared to HepG2 cells expressing DENV C (R85A/K86A). In addition, CD137-mediated apoptotic activity in HepG2 cells expressing DENV C was significantly increased by anti-CD137 antibody compared to that of HepG2 cells expressing DENV C (R85A/K86A). In DENV-infected HepG2 cells, CD137 mRNA and CD137 positive cells significantly increased and CD137-mediated apoptotic activity was increased by anti-CD137 antibody. This work is the first to demonstrate the contribution of CD137 signaling to DENV-mediated apoptosis.

Role of CD137 signaling in dengue virus-mediated apoptosis

Energy Technology Data Exchange (ETDEWEB)

Nagila, Amar [Medical Molecular Biology Unit, Office for Research and Development, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Department of Biochemistry, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Netsawang, Janjuree [Faculty of Medical Technology, Rangsit University, Bangkok (Thailand); Srisawat, Chatchawan [Department of Biochemistry, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Noisakran, Sansanee [Dengue Hemorrhagic Fever Research Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok (Thailand); Medical Biotechnology Unit, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Bangkok (Thailand); Morchang, Atthapan; Yasamut, Umpa [Medical Molecular Biology Unit, Office for Research and Development, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Department of Immunology, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Puttikhunt, Chunya [Dengue Hemorrhagic Fever Research Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok (Thailand); Medical Biotechnology Unit, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Bangkok (Thailand); Kasinrerk, Watchara [Division of Clinical Immunology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai (Thailand); Biomedical Technology Research Center, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency at Chiang Mai University, Chiang Mai (Thailand); and others

2011-07-08

Highlights: {yields} For the first time the role of CD137 in dengue virus (DENV) infection. {yields} Induction of DENV-mediated apoptosis by CD137 signaling. {yields} Sensitization to CD137-mediated apoptosis by dengue virus capsid protein (DENV C). {yields} Nuclear localization of DENV C is required for CD137-mediated apoptosis. -- Abstract: Hepatic dysfunction is a well recognized feature of dengue virus (DENV) infection. However, molecular mechanisms of hepatic injury are still poorly understood. A complex interaction between DENV and the host immune response contributes to DENV-mediated tissue injury. DENV capsid protein (DENV C) physically interacts with the human death domain-associated protein Daxx. A double substitution mutation in DENV C (R85A/K86A) abrogates Daxx interaction, nuclear localization and apoptosis. Therefore we compared the expression of cell death genes between HepG2 cells expressing DENV C and DENV C (R85A/K86A) using a real-time PCR array. Expression of CD137, which is a member of the tumor necrosis factor receptor family, increased significantly in HepG2 cells expressing DENV C compared to HepG2 cells expressing DENV C (R85A/K86A). In addition, CD137-mediated apoptotic activity in HepG2 cells expressing DENV C was significantly increased by anti-CD137 antibody compared to that of HepG2 cells expressing DENV C (R85A/K86A). In DENV-infected HepG2 cells, CD137 mRNA and CD137 positive cells significantly increased and CD137-mediated apoptotic activity was increased by anti-CD137 antibody. This work is the first to demonstrate the contribution of CD137 signaling to DENV-mediated apoptosis.

A de novo missense mutation of FGFR2 causes facial dysplasia syndrome in Holstein cattle

DEFF Research Database (Denmark)

Agerholm, Jørgen Steen; McEvoy, Fintan; Heegaard, Steffen

2017-01-01

was suspected as all recorded cases were progeny of the same sire. Detailed investigations were performed to characterize the syndrome and to reveal its cause. Results Seven malformed calves were submitted examination. All cases shared a common morphology with the most striking lesions being severe facial...... chromosome 26 where whole genome sequencing of a case-parent trio revealed two de novo variants perfectly associated with the disease: an intronic SNP in the DMBT1 gene and a single non-synonymous variant in the FGFR2 gene. This FGFR2 missense variant (c.927G>T) affects a gene encoding a member...... of the fibroblast growth factor receptor family, where amino acid sequence is highly conserved between members and across species. It is predicted to change an evolutionary conserved tryptophan into a cysteine residue (p.Trp309Cys). Both variant alleles were proven to result from de novo mutation events...

Endothelial MMP14 is required for endothelial-dependent growth support of human airway basal cells

Science.gov (United States)

Ding, Bi-Sen; Gomi, Kazunori; Rafii, Shahin; Crystal, Ronald G.; Walters, Matthew S.

2015-01-01

ABSTRACT Human airway basal cells are the stem (or progenitor) population of the airway epithelium, and play a central role in anchoring the epithelium to the basement membrane. The anatomic position of basal cells allows for potential paracrine signaling between them and the underlying non-epithelial stromal cells. In support of this, we have previously demonstrated that endothelial cells support growth of basal cells during co-culture through vascular endothelial growth factor A (VEGFA)-mediated signaling. Building on these findings, we found, by RNA sequencing analysis, that basal cells expressed multiple fibroblast growth factor (FGF) ligands (FGF2, FGF5, FGF11 and FGF13) and that only FGF2 and FGF5 were capable of functioning in a paracrine manner to activate classical FGF receptor (FGFR) signaling. Antibody-mediated blocking of FGFR1 during basal-cell–endothelial-cell co-culture significantly reduced the endothelial-cell-dependent basal cell growth. Stimulation of endothelial cells with basal-cell-derived growth factors induced endothelial cell expression of matrix metallopeptidase 14 (MMP14), and short hairpin RNA (shRNA)-mediated knockdown of endothelial cell MMP14 significantly reduced the endothelial-cell-dependent growth of basal cells. Overall, these data characterize a new growth-factor-mediated reciprocal ‘crosstalk’ between human airway basal cells and endothelial cells that regulates proliferation of basal cells. PMID:26116571

Clinical outcomes of myeloid/lymphoid neoplasms with fibroblast growth factor receptor-1 (FGFR1) rearrangement.

Science.gov (United States)

Umino, Kento; Fujiwara, Shin-Ichiro; Ikeda, Takashi; Toda, Yumiko; Ito, Shoko; Mashima, Kiyomi; Minakata, Daisuke; Nakano, Hirofumi; Yamasaki, Ryoko; Kawasaki, Yasufumi; Sugimoto, Miyuki; Yamamoto, Chihiro; Ashizawa, Masahiro; Hatano, Kaoru; Sato, Kazuya; Oh, Iekuni; Ohmine, Ken; Muroi, Kazuo; Kanda, Yoshinobu

2018-02-28

Myeloid/lymphoid neoplasms with fibroblast growth factor receptor-1 (FGFR1) rearrangement are hematopoietic stem cell disorders with a poor prognosis, but no established standard therapy. We experienced a patient with T-lymphoblastic lymphoma (LBL) associated with FGFR1 rearrangement who underwent cord blood transplantation, but died of pulmonary complication. We collected the clinical data of patients with FGFR1 rearrangement from the medical literature and analyzed 45 patients, including our patient. The primary diagnoses were myeloproliferative neoplasm (MPN) or myelodysplastic syndromes (MDS) in 14 and acute leukemia or LBL in 31. In MPN and MDS patients, the cumulative incidence of transformation to blast phase (BP) at 12 months was 46.2%. The 1-year overall survival (OS) from diagnosis in all cases was 43.1%. With regard to the impact of treatment response on survival, the achievement of complete response with a landmark at 2 months after diagnosis of BP was associated with a superior OS (40.0% vs. 26.0% P = 0.011 for 1-year OS from BP). Allogeneic hematopoietic stem cell transplantation (HSCT) was performed in 13 patients, and the 1-year OS from allogeneic HSCT was 61.5%. The hazard ratio for mortality was 0.34 (95% CI, 0.08-1.51, P = 0.15) for allogeneic HSCT treated as a time-dependent covariate, which suggests that allogeneic HSCT may confer a clinical benefit. The further accumulation of clinical data is needed to determine the optimal therapeutic approach for these neoplasms.

BMPRIA mediated signaling is essential for temporomandibular joint development in mice.

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Shuping Gu

Full Text Available The central importance of BMP signaling in the development and homeostasis of synovial joint of appendicular skeleton has been well documented, but its role in the development of temporomandibular joint (TMJ, also classified as a synovial joint, remains completely unknown. In this study, we investigated the function of BMPRIA mediated signaling in TMJ development in mice by transgenic loss-of- and gain-of-function approaches. We found that BMPRIA is expressed in the cranial neural crest (CNC-derived developing condyle and glenoid fossa, major components of TMJ, as well as the interzone mesenchymal cells. Wnt1-Cre mediated tissue specific inactivation of BmprIa in CNC lineage led to defective TMJ development, including failure of articular disc separation from a hypoplastic condyle, persistence of interzone cells, and failed formation of a functional fibrocartilage layer on the articular surface of the glenoid fossa and condyle, which could be at least partially attributed to the down-regulation of Ihh in the developing condyle and inhibition of apoptosis in the interzone. On the other hand, augmented BMPRIA signaling by Wnt1-Cre driven expression of a constitutively active form of BmprIa (caBmprIa inhibited osteogenesis of the glenoid fossa and converted the condylar primordium from secondary cartilage to primary cartilage associated with ectopic activation of Smad-dependent pathway but inhibition of JNK pathway, leading to TMJ agenesis. Our results present unambiguous evidence for an essential role of finely tuned BMPRIA mediated signaling in TMJ development.

Notch Signaling Mediates Skeletal Muscle Atrophy in Cancer Cachexia Caused by Osteosarcoma

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Xiaodong Mu

2016-01-01

Full Text Available Skeletal muscle atrophy in cancer cachexia is mediated by the interaction between muscle stem cells and various tumor factors. Although Notch signaling has been known as a key regulator of both cancer development and muscle stem cell activity, the potential involvement of Notch signaling in cancer cachexia and concomitant muscle atrophy has yet to be elucidated. The murine K7M2 osteosarcoma cell line was used to generate an orthotopic model of sarcoma-associated cachexia, and the role of Notch signaling was evaluated. Skeletal muscle atrophy was observed in the sarcoma-bearing mice, and Notch signaling was highly active in both tumor tissues and the atrophic skeletal muscles. Systemic inhibition of Notch signaling reduced muscle atrophy. In vitro coculture of osteosarcoma cells with muscle-derived stem cells (MDSCs isolated from normal mice resulted in decreased myogenic potential of MDSCs, while the application of Notch inhibitor was able to rescue this repressed myogenic potential. We further observed that Notch-activating factors reside in the exosomes of osteosarcoma cells, which activate Notch signaling in MDSCs and subsequently repress myogenesis. Our results revealed that signaling between tumor and muscle via the Notch pathway may play an important role in mediating the skeletal muscle atrophy seen in cancer cachexia.

A single peroxisomal targeting signal mediates matrix protein import in diatoms.

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Nicola H Gonzalez

Full Text Available Peroxisomes are single membrane bound compartments. They are thought to be present in almost all eukaryotic cells, although the bulk of our knowledge about peroxisomes has been generated from only a handful of model organisms. Peroxisomal matrix proteins are synthesized cytosolically and posttranslationally imported into the peroxisomal matrix. The import is generally thought to be mediated by two different targeting signals. These are respectively recognized by the two import receptor proteins Pex5 and Pex7, which facilitate transport across the peroxisomal membrane. Here, we show the first in vivo localization studies of peroxisomes in a representative organism of the ecologically relevant group of diatoms using fluorescence and transmission electron microscopy. By expression of various homologous and heterologous fusion proteins we demonstrate that targeting of Phaeodactylum tricornutum peroxisomal matrix proteins is mediated only by PTS1 targeting signals, also for proteins that are in other systems imported via a PTS2 mode of action. Additional in silico analyses suggest this surprising finding may also apply to further diatoms. Our data suggest that loss of the PTS2 peroxisomal import signal is not reserved to Caenorhabditis elegans as a single exception, but has also occurred in evolutionary divergent organisms. Obviously, targeting switching from PTS2 to PTS1 across different major eukaryotic groups might have occurred for different reasons. Thus, our findings question the widespread assumption that import of peroxisomal matrix proteins is generally mediated by two different targeting signals. Our results implicate that there apparently must have been an event causing the loss of one targeting signal even in the group of diatoms. Different possibilities are discussed that indicate multiple reasons for the detected targeting switching from PTS2 to PTS1.

­Glial and stem cell expression of murine Fibroblast Growth Factor Receptor 1 in the embryonic and perinatal nervous system

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Jantzen C. Collette

2017-06-01

Full Text Available Background Fibroblast growth factors (FGFs and their receptors (FGFRs are involved in the development and function of multiple organs and organ systems, including the central nervous system (CNS. FGF signaling via FGFR1, one of the three FGFRs expressed in the CNS, stimulates proliferation of stem cells during prenatal and postnatal neurogenesis and participates in regulating cell-type ratios in many developing regions of the brain. Anomalies in FGFR1 signaling have been implicated in certain neuropsychiatric disorders. Fgfr1 expression has been shown, via in situ hybridization, to vary spatially and temporally throughout embryonic and postnatal development of the brain. However, in situ hybridization lacks sufficient resolution to identify which cell-types directly participate in FGF signaling. Furthermore, because antibodies raised against FGFR1 commonly cross-react with other members of the FGFR family, immunocytochemistry is not alone sufficient to accurately document Fgfr1 expression. Here, we elucidate the identity of Fgfr1 expressing cells in both the embryonic and perinatal mouse brain. Methods To do this, we utilized a tgFGFR1-EGFPGP338Gsat BAC line (tgFgfr1-EGFP+ obtained from the GENSAT project. The tgFgfr1-EGFP+ line expresses EGFP under the control of a Fgfr1 promoter, thereby causing cells endogenously expressing Fgfr1 to also present a positive GFP signal. Through simple immunostaining using GFP antibodies and cell-type specific antibodies, we were able to accurately determine the cell-type of Fgfr1 expressing cells. Results This technique revealed Fgfr1 expression in proliferative zones containing BLBP+ radial glial stem cells, such as the cortical and hippocampal ventricular zones, and cerebellar anlage of E14.5 mice, in addition to DCX+ neuroblasts. Furthermore, our data reveal Fgfr1 expression in proliferative zones containing BLBP+ cells of the anterior midline, hippocampus, cortex, hypothalamus, and cerebellum of P0.5 mice

Molecular Signaling Pathways Mediating Osteoclastogenesis Induced by Prostate Cancer Cells

International Nuclear Information System (INIS)

Rafiei, Shahrzad; Komarova, Svetlana V

2013-01-01

Advanced prostate cancer commonly metastasizes to bone leading to osteoblastic and osteolytic lesions. Although an osteolytic component governed by activation of bone resorbing osteoclasts is prominent in prostate cancer metastasis, the molecular mechanisms of prostate cancer-induced osteoclastogenesis are not well-understood. We studied the effect of soluble mediators released from human prostate carcinoma cells on osteoclast formation from mouse bone marrow and RAW 264.7 monocytes. Soluble factors released from human prostate carcinoma cells significantly increased viability of naïve bone marrow monocytes, as well as osteoclastogenesis from precursors primed with receptor activator of nuclear factor κ-B ligand (RANKL). The prostate cancer-induced osteoclastogenesis was not mediated by RANKL as it was not inhibited by osteoprotegerin (OPG). However inhibition of TGFβ receptor I (TβRI), or macrophage-colony stimulating factor (MCSF) resulted in attenuation of prostate cancer-induced osteoclastogenesis. We characterized the signaling pathways induced in osteoclast precursors by soluble mediators released from human prostate carcinoma cells. Prostate cancer factors increased basal calcium levels and calcium fluctuations, induced nuclear localization of nuclear factor of activated t-cells (NFAT)c1, and activated prolonged phosphorylation of ERK1/2 in RANKL-primed osteoclast precursors. Inhibition of calcium signaling, NFATc1 activation, and ERK1/2 phosphorylation significantly reduced the ability of prostate cancer mediators to stimulate osteoclastogenesis. This study reveals the molecular mechanisms underlying the direct osteoclastogenic effect of prostate cancer derived factors, which may be beneficial in developing novel osteoclast-targeting therapeutic approaches

Novel FGFR1 and KISS1R Mutations in Chinese Kallmann Syndrome Males with Cleft Lip/Palate

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Hao Xu

2015-01-01

Full Text Available Kallmann syndrome (KS is characterized by isolated hypogonadotropic hypogonadism (IHH with anosmia and is sometimes associated with cleft lip/palate (CLP. In order to describe the clinical features, genetic etiology, and treatment outcome of KS males with CLP, we performed genetic screening for 15 known causal IHH genes (KAL1, FGFR1, NELF, FGF8, CHD7, WDR11, SEMA3A, KISS1R, KISS1, PROKR2, PROK2, TAC3, TACR3, GNRH1, and GNRHR in four KS with CLP patients and six IHH patients without CLP. Two novel heterozygous missense mutations in FGFR1, (NM_001174066: c.776G>A (p.G259E and (NM_001174066: c.358C>T (p.R120C, were identified in a 23-year-old KS male with cleft lip and an 18-year-old KS patient with cleft lip and palate, dental agenesis, and high arched palate, respectively. These two mutations were not presented in their healthy parents and 200 normal controls. One novel heterozygous missense mutation in KISS1R, (NM_032551: c.587C>A (p.P196H, was identified in an 18-year-old KS male with cleft lip and dental agenesis who developed sperm after being treated with gonadotropin. This mutation was also presented in his healthy father and grandfather. These results have implications for the diagnosis, genetic counseling, and treatment of KS and CLP males with mutations in FGFR1 gene.

Volatile-Mediated within-Plant Signaling in Hybrid Aspen: Required for Systemic Responses.

Science.gov (United States)

Li, Tao; Blande, James D

2017-04-01

Plant volatiles play crucial roles in signaling between plants and their associated community members, but their role in within-plant signaling remains largely unexplored, particularly under field conditions. Using a system comprising the hybrid aspen (Populus tremula x tremuloides) and the specialized herbivorous leaf beetle (Phratora laticollis) and, combining field, greenhouse and laboratory experiments, we examined whether local damage triggered systemic responses in undamaged branches that lack vascular connection to the damaged branches, and to what extent this was caused by airborne volatile signals versus internal signals. An experiment tracing dye through the vasculature of saplings revealed no downward movement of the dye from upper to lower branches, suggesting a lack of vascular connectivity among branches. However, we found under both field and laboratory conditions that herbivore feeding on upper branches elicited volatile emissions by undamaged lower branches. Greenhouse experiments manipulating air contact between damaged and undamaged branches showed that systemic induction of volatiles was almost eliminated when air contact was interrupted. Our findings clearly demonstrate that herbivore-induced volatiles overcome vascular constraints and mediate within-plant signaling. Further, we found that volatile signaling led to induction of different classes of volatiles under field and environment controlled conditions, with a weaker response observed in the field. This difference not only reflects the dose- and time-dependent nature of volatile signaling, but also points out that future studies should focus more on field observations to better understand the ecological role of volatile-mediated within-plant signaling.

Pannexin 1 channels mediate 'find-me' signal release and membrane permeability during apoptosis.

Science.gov (United States)

Chekeni, Faraaz B; Elliott, Michael R; Sandilos, Joanna K; Walk, Scott F; Kinchen, Jason M; Lazarowski, Eduardo R; Armstrong, Allison J; Penuela, Silvia; Laird, Dale W; Salvesen, Guy S; Isakson, Brant E; Bayliss, Douglas A; Ravichandran, Kodi S

2010-10-14

Apoptotic cells release 'find-me' signals at the earliest stages of death to recruit phagocytes. The nucleotides ATP and UTP represent one class of find-me signals, but their mechanism of release is not known. Here, we identify the plasma membrane channel pannexin 1 (PANX1) as a mediator of find-me signal/nucleotide release from apoptotic cells. Pharmacological inhibition and siRNA-mediated knockdown of PANX1 led to decreased nucleotide release and monocyte recruitment by apoptotic cells. Conversely, PANX1 overexpression enhanced nucleotide release from apoptotic cells and phagocyte recruitment. Patch-clamp recordings showed that PANX1 was basally inactive, and that induction of PANX1 currents occurred only during apoptosis. Mechanistically, PANX1 itself was a target of effector caspases (caspases 3 and 7), and a specific caspase-cleavage site within PANX1 was essential for PANX1 function during apoptosis. Expression of truncated PANX1 (at the putative caspase cleavage site) resulted in a constitutively open channel. PANX1 was also important for the 'selective' plasma membrane permeability of early apoptotic cells to specific dyes. Collectively, these data identify PANX1 as a plasma membrane channel mediating the regulated release of find-me signals and selective plasma membrane permeability during apoptosis, and a new mechanism of PANX1 activation by caspases.

PDL1 Signals through Conserved Sequence Motifs to Overcome Interferon-Mediated Cytotoxicity

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Maria Gato-Cañas

2017-08-01

Full Text Available PDL1 blockade produces remarkable clinical responses, thought to occur by T cell reactivation through prevention of PDL1-PD1 T cell inhibitory interactions. Here, we find that PDL1 cell-intrinsic signaling protects cancer cells from interferon (IFN cytotoxicity and accelerates tumor progression. PDL1 inhibited IFN signal transduction through a conserved class of sequence motifs that mediate crosstalk with IFN signaling. Abrogation of PDL1 expression or antibody-mediated PDL1 blockade strongly sensitized cancer cells to IFN cytotoxicity through a STAT3/caspase-7-dependent pathway. Moreover, somatic mutations found in human carcinomas within these PDL1 sequence motifs disrupted motif regulation, resulting in PDL1 molecules with enhanced protective activities from type I and type II IFN cytotoxicity. Overall, our results reveal a mode of action of PDL1 in cancer cells as a first line of defense against IFN cytotoxicity.

Aberrant Receptor Internalization and Enhanced FRS2-dependent Signaling Contribute to the Transforming Activity of the Fibroblast Growth Factor Receptor 2 IIIb C3 Isoform*

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Cha, Jiyoung Y.; Maddileti, Savitri; Mitin, Natalia; Harden, T. Kendall; Der, Channing J.

2009-01-01

Alternative splice variants of fibroblast growth factor receptor 2 (FGFR2) IIIb, designated C1, C2, and C3, possess progressive reduction in their cytoplasmic carboxyl termini (822, 788, and 769 residues, respectively), with preferential expression of the C2 and C3 isoforms in human cancers. We determined that the progressive deletion of carboxyl-terminal sequences correlated with increasing transforming potency. The highly transforming C3 variant lacks five tyrosine residues present in C1, and we determined that the loss of Tyr-770 alone enhanced FGFR2 IIIb C1 transforming activity. Because Tyr-770 may compose a putative YXXL sorting motif, we hypothesized that loss of Tyr-770 in the 770YXXL motif may cause disruption of FGFR2 IIIb C1 internalization and enhance transforming activity. Surprisingly, we found that mutation of Leu-773 but not Tyr-770 impaired receptor internalization and increased receptor stability and activation. Interestingly, concurrent mutations of Tyr-770 and Leu-773 caused 2-fold higher transforming activity than caused by the Y770F or L773A single mutations, suggesting loss of Tyr and Leu residues of the 770YXXL773 motif enhances FGFR2 IIIb transforming activity by distinct mechanisms. We also determined that loss of Tyr-770 caused persistent activation of FRS2 by enhancing FRS2 binding to FGFR2 IIIb. Furthermore, we found that FRS2 binding to FGFR2 IIIb is required for increased FRS2 tyrosine phosphorylation and enhanced transforming activity by Y770F mutation. Our data support a dual mechanism where deletion of the 770YXXL773 motif promotes FGFR2 IIIb C3 transforming activity by causing aberrant receptor recycling and stability and persistent FRS2-dependent signaling. PMID:19103595

FGFR1 tyrosine kinase domain duplication in pilocytic astrocytoma with anaplasia.

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Ballester, Leomar Y; Penas-Prado, Marta; Leeds, Norman E; Huse, Jason T; Fuller, Gregory N

2018-04-01

We report the case of a 27-yr-old male with visual field loss who had a 4.9-cm complex cystic mass in the right occipital lobe. Histologic examination showed pilocytic astrocytoma (PA) with anaplasia, and molecular characterization revealed FGFR1 duplication with additional variants of unknown significance in several genes ( ARID1A, ARID1B, CHEK2, EPHA5, and MLL2 ). This is one of only a very few reported cases of anaplastic PA with characterization of molecular alterations. © 2018 Ballester et al.; Published by Cold Spring Harbor Laboratory Press.

Congenital amegakaryocytic thrombocytopenia iPS cells exhibit defective MPL-mediated signaling.

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Hirata, Shinji; Takayama, Naoya; Jono-Ohnishi, Ryoko; Endo, Hiroshi; Nakamura, Sou; Dohda, Takeaki; Nishi, Masanori; Hamazaki, Yuhei; Ishii, Ei-ichi; Kaneko, Shin; Otsu, Makoto; Nakauchi, Hiromitsu; Kunishima, Shinji; Eto, Koji

2013-09-01

Congenital amegakaryocytic thrombocytopenia (CAMT) is caused by the loss of thrombopoietin receptor-mediated (MPL-mediated) signaling, which causes severe pancytopenia leading to bone marrow failure with onset of thrombocytopenia and anemia prior to leukopenia. Because Mpl(-/-) mice do not exhibit the human disease phenotype, we used an in vitro disease tracing system with induced pluripotent stem cells (iPSCs) derived from a CAMT patient (CAMT iPSCs) and normal iPSCs to investigate the role of MPL signaling in hematopoiesis. We found that MPL signaling is essential for maintenance of the CD34+ multipotent hematopoietic progenitor (MPP) population and development of the CD41+GPA+ megakaryocyte-erythrocyte progenitor (MEP) population, and its role in the fate decision leading differentiation toward megakaryopoiesis or erythropoiesis differs considerably between normal and CAMT cells. Surprisingly, complimentary transduction of MPL into normal or CAMT iPSCs using a retroviral vector showed that MPL overexpression promoted erythropoiesis in normal CD34+ hematopoietic progenitor cells (HPCs), but impaired erythropoiesis and increased aberrant megakaryocyte production in CAMT iPSC-derived CD34+ HPCs, reflecting a difference in the expression of the transcription factor FLI1. These results demonstrate that impaired transcriptional regulation of the MPL signaling that normally governs megakaryopoiesis and erythropoiesis underlies CAMT.

Concurrent inhibition of kit- and FcepsilonRI-mediated signaling: coordinated suppression of mast cell activation

DEFF Research Database (Denmark)

Jensen, Bettina M; Beaven, Michael A; Iwaki, Shoko

2008-01-01

Although primarily required for the growth, differentiation, and survival of mast cells, Kit ligand (stem cell factor) is also required for optimal antigen-mediated mast cell activation. Therefore, concurrent inhibition of Kit- and FcepsilonRI-mediated signaling would be an attractive approach...... characterized Kit inhibitor imatinib mesylate (imatinib). In contrast to imatinib, however, hypothemycin also effectively inhibited FcepsilonRI-mediated degranulation and cytokine production in addition to the potentiation of these responses via Kit. The effect of hypothemycin on Kit-mediated responses could...... be explained by its inhibition of Kit kinase activity, whereas the inhibitory effects on FcepsilonRI-dependent signaling were at the level of Btk activation. Because hypothemycin also significantly reduced the mouse passive cutaneous anaphylaxis response in vivo, these data provide proof of principle...

Proteomic analysis of the signaling pathway mediated by the heterotrimeric Gα protein Pga1 of Penicillium chrysogenum.

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Carrasco-Navarro, Ulises; Vera-Estrella, Rosario; Barkla, Bronwyn J; Zúñiga-León, Eduardo; Reyes-Vivas, Horacio; Fernández, Francisco J; Fierro, Francisco

2016-10-06

The heterotrimeric Gα protein Pga1-mediated signaling pathway regulates the entire developmental program in Penicillium chrysogenum, from spore germination to the formation of conidia. In addition it participates in the regulation of penicillin biosynthesis. We aimed to advance the understanding of this key signaling pathway using a proteomics approach, a powerful tool to identify effectors participating in signal transduction pathways. Penicillium chrysogenum mutants with different levels of activity of the Pga1-mediated signaling pathway were used to perform comparative proteomic analyses by 2D-DIGE and LC-MS/MS. Thirty proteins were identified which showed differences in abundance dependent on Pga1 activity level. By modifying the intracellular levels of cAMP we could establish cAMP-dependent and cAMP-independent pathways in Pga1-mediated signaling. Pga1 was shown to regulate abundance of enzymes in primary metabolic pathways involved in ATP, NADPH and cysteine biosynthesis, compounds that are needed for high levels of penicillin production. An in vivo phosphorylated protein containing a pleckstrin homology domain was identified; this protein is a candidate for signal transduction activity. Proteins with possible roles in purine metabolism, protein folding, stress response and morphogenesis were also identified whose abundance was regulated by Pga1 signaling. Thirty proteins whose abundance was regulated by the Pga1-mediated signaling pathway were identified. These proteins are involved in primary metabolism, stress response, development and signal transduction. A model describing the pathways through which Pga1 signaling regulates different cellular processes is proposed.

Motile cilia of human airway epithelia contain hedgehog signaling components that mediate noncanonical hedgehog signaling.

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Mao, Suifang; Shah, Alok S; Moninger, Thomas O; Ostedgaard, Lynda S; Lu, Lin; Tang, Xiao Xiao; Thornell, Ian M; Reznikov, Leah R; Ernst, Sarah E; Karp, Philip H; Tan, Ping; Keshavjee, Shaf; Abou Alaiwa, Mahmoud H; Welsh, Michael J

2018-02-06

Differentiated airway epithelia produce sonic hedgehog (SHH), which is found in the thin layer of liquid covering the airway surface. Although previous studies showed that vertebrate HH signaling requires primary cilia, as airway epithelia mature, the cells lose primary cilia and produce hundreds of motile cilia. Thus, whether airway epithelia have apical receptors for SHH has remained unknown. We discovered that motile cilia on airway epithelial cells have HH signaling proteins, including patched and smoothened. These cilia also have proteins affecting cAMP-dependent signaling, including Gα i and adenylyl cyclase 5/6. Apical SHH decreases intracellular levels of cAMP, which reduces ciliary beat frequency and pH in airway surface liquid. These results suggest that apical SHH may mediate noncanonical HH signaling through motile cilia to dampen respiratory defenses at the contact point between the environment and the lung, perhaps counterbalancing processes that stimulate airway defenses. Copyright © 2018 the Author(s). Published by PNAS.

Frequency of the allelic variant c.1150T > C in exon 10 of the fibroblast growth factor receptor 3 (FGFR3 gene is not increased in patients with pathogenic mutations and related chondrodysplasia phenotypes

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Thatiane Yoshie Kanazawa

2014-12-01

Full Text Available Mutations in the FGFR3 gene cause the phenotypic spectrum of FGFR3 chondrodysplasias ranging from lethal forms to the milder phenotype seen in hypochondroplasia (Hch. The p.N540K mutation in the FGFR3 gene occurs in ~70% of individuals with Hch, and nearly 30% of individuals with the Hch phenotype have no mutations in the FGFR3, which suggests genetic heterogeneity. The identification of a severe case of Hch associated with the typical mutation c.1620C > A and the occurrence of a c.1150T > C change that resulted in a p.F384L in exon 10, together with the suspicion that this second change could be a modulator of the phenotype, prompted us to investigate this hypothesis in a cohort of patients. An analysis of 48 patients with FGFR3 chondrodysplasia phenotypes and 330 healthy (control individuals revealed no significant difference in the frequency of the C allele at the c.1150 position (p = 0.34. One patient carrying the combination `pathogenic mutation plus the allelic variant c.1150T > C' had a typical achondroplasia (Ach phenotype. In addition, three other patients with atypical phenotypes showed no association with the allelic variant. Together, these results do not support the hypothesis of a modulatory role for the c.1150T > C change in the FGFR3 gene.

Severe complications in a child with achondroplasia and two FGFR3 mutations on the same allele

NARCIS (Netherlands)

Rump, P; Letteboer, TGW; Gille, JJP; Torringa, MJL; Baerts, W; van Gestel, JPJ; Verheij, JBGM; van Essen, AJ

2006-01-01

We describe a unique case of achondroplasia with associated complications, including severe respiratory problems. Molecular analysis of the fibroblast growth factor receptor type 3 (FGFR3) gene in this patient showed the common p.G380R mutation and a second novel p.L377R Mutation. An allele-specific

Reversible Congenital Hypogonadotropic Hypogonadism in Patients with CHD7, FGFR1 or GNRHR Mutations

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Laitinen, Eeva-Maria; Tommiska, Johanna; Sane, Timo; Vaaralahti, Kirsi; Toppari, Jorma; Raivio, Taneli

2012-01-01

Background Congenital hypogonadotropic hypogonadism (HH) is a rare cause for delayed or absent puberty. These patients may recover from HH spontaneously in adulthood. To date, it is not possible to predict who will undergo HH reversal later in life. Herein we investigated whether Finnish patients with reversal of congenital hypogonadotropic hypogonadism (HH) have common phenotypic or genotypic features. Methods and Findings Thirty-two male HH patients with anosmia/hyposmia (Kallmann Syndrome, KS; n = 26) or normal sense of smell (nHH; n = 6) were enrolled (age range, 18–61 yrs). The patients were clinically examined, and reversal of HH was assessed after treatment withdrawal. KAL1, FGFR1, FGF8, PROK2, PROKR2, CHD7, WDR11, GNRHR, GNRH1, KISS1R, KISS1, TAC3, TACR3, and LHβ were screened for mutations. Six HH patients (2 KS, 4 nHH) were verified to have reversal of HH. In the majority of cases, reversal occurred early in adulthood (median age, 23 yrs; range, 21–39 yrs). All had spontaneous testicular growth while on testosterone replacement therapy (TRT). One nHH subject was restarted on TRT due to a decline in serum T. Two reversal variants had a same GNRHR mutation (R262Q), which was accompanied by another GNRHR mutation (R139H or del309F). In addition, both of the KS patients had a mutation in CHD7 (p.Q51X) or FGFR1 (c.91+2T>A). Conclusions Considerable proportion of patients with HH (8% of KS probands) may recover in early adulthood. Spontaneous testicular enlargement during TRT was highly suggestive for reversal of HH. Those with the GNRHR mutation R262Q accompanied by another GNRHR mutation may be prone to reversal, although even patients with a truncating mutation in CHD7 or a splice-site mutation in FGFR1 can recover. We recommend that all adolescents and young adults with congenital HH should be informed on the possibility of reversal. PMID:22724017

Reversible congenital hypogonadotropic hypogonadism in patients with CHD7, FGFR1 or GNRHR mutations.

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Eeva-Maria Laitinen

Full Text Available BACKGROUND: Congenital hypogonadotropic hypogonadism (HH is a rare cause for delayed or absent puberty. These patients may recover from HH spontaneously in adulthood. To date, it is not possible to predict who will undergo HH reversal later in life. Herein we investigated whether Finnish patients with reversal of congenital hypogonadotropic hypogonadism (HH have common phenotypic or genotypic features. METHODS AND FINDINGS: Thirty-two male HH patients with anosmia/hyposmia (Kallmann Syndrome, KS; n = 26 or normal sense of smell (nHH; n = 6 were enrolled (age range, 18-61 yrs. The patients were clinically examined, and reversal of HH was assessed after treatment withdrawal. KAL1, FGFR1, FGF8, PROK2, PROKR2, CHD7, WDR11, GNRHR, GNRH1, KISS1R, KISS1, TAC3, TACR3, and LHβ were screened for mutations. Six HH patients (2 KS, 4 nHH were verified to have reversal of HH. In the majority of cases, reversal occurred early in adulthood (median age, 23 yrs; range, 21-39 yrs. All had spontaneous testicular growth while on testosterone replacement therapy (TRT. One nHH subject was restarted on TRT due to a decline in serum T. Two reversal variants had a same GNRHR mutation (R262Q, which was accompanied by another GNRHR mutation (R139H or del309F. In addition, both of the KS patients had a mutation in CHD7 (p.Q51X or FGFR1 (c.91+2T>A. CONCLUSIONS: Considerable proportion of patients with HH (8% of KS probands may recover in early adulthood. Spontaneous testicular enlargement during TRT was highly suggestive for reversal of HH. Those with the GNRHR mutation R262Q accompanied by another GNRHR mutation may be prone to reversal, although even patients with a truncating mutation in CHD7 or a splice-site mutation in FGFR1 can recover. We recommend that all adolescents and young adults with congenital HH should be informed on the possibility of reversal.

Wnt5a signaling is a substantial constituent in bone morphogenetic protein-2-mediated osteoblastogenesis

International Nuclear Information System (INIS)

Nemoto, Eiji; Ebe, Yukari; Kanaya, Sousuke; Tsuchiya, Masahiro; Nakamura, Takashi; Tamura, Masato; Shimauchi, Hidetoshi

2012-01-01

Highlights: ► Wnt5a is identified in osteoblasts in tibial growth plate and bone marrow. ► Osteoblastic differentiation is associated with increased expression of Wnt5a/Ror2. ► Wnt5a/Ror2 signaling is important for BMP-2-mediated osteoblastic differentiation. ► Wnt5a/Ror2 operates independently of BMP-Smad pathway. -- Abstract: Wnts are secreted glycoproteins that mediate developmental and post-developmental physiology by regulating cellular processes including proliferation, differentiation, and apoptosis through β-catenin-dependent canonical and β-catenin-independent noncanonical pathway. It has been reported that Wnt5a activates noncanonical Wnt signaling through receptor tyrosine kinase-like orphan receptor 2 (Ror2). Although it appears that Wnt5a/Ror2 signaling supports normal bone physiology, the biological significance of noncanonical Wnts in osteogenesis is essentially unknown. In this study, we identified expression of Wnt5a in osteoblasts in the ossification zone of the tibial growth plate as well as bone marrow of the rat tibia as assessed by immunohistochemistry. In addition, we show that osteoblastic differentiation mediated by BMP-2 is associated with increased expression of Wnt5a and Ror2 using cultured pre-osteoblasts, MC3T3-E1 cells. Silencing gene expression of Wnt5a and Ror2 in MC3T3-E1 cells results in suppression of BMP-2-mediated osteoblastic differentiation, suggesting that Wnt5a and Ror2 signaling are of substantial importance for BMP-2-mediated osteoblastic differentiation. BMP-2 stimulation induced phosphorylation of Smad1/5/8 in a similar fashion in both siWnt5a-treated cells and control cells, suggesting that Wnt5a was dispensable for the phosphorylation of Smads by BMP-2. Taken together, our results suggest that Wnt5a/Ror2 signaling appears to be involved in BMP-2-mediated osteoblast differentiation in a Smad independent pathway.

Ciliopathy proteins regulate paracrine signaling by modulating proteasomal degradation of mediators

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Liu, Yangfan P.; Tsai, I-Chun; Morleo, Manuela; Oh, Edwin C.; Leitch, Carmen C.; Massa, Filomena; Lee, Byung-Hoon; Parker, David S.; Finley, Daniel; Zaghloul, Norann A.; Franco, Brunella; Katsanis, Nicholas

2014-01-01

Cilia are critical mediators of paracrine signaling; however, it is unknown whether proteins that contribute to ciliopathies converge on multiple paracrine pathways through a common mechanism. Here, we show that loss of cilopathy-associated proteins Bardet-Biedl syndrome 4 (BBS4) or oral-facial-digital syndrome 1 (OFD1) results in the accumulation of signaling mediators normally targeted for proteasomal degradation. In WT cells, several BBS proteins and OFD1 interacted with proteasomal subunits, and loss of either BBS4 or OFD1 led to depletion of multiple subunits from the centrosomal proteasome. Furthermore, overexpression of proteasomal regulatory components or treatment with proteasomal activators sulforaphane (SFN) and mevalonolactone (MVA) ameliorated signaling defects in cells lacking BBS1, BBS4, and OFD1, in morphant zebrafish embryos, and in induced neurons from Ofd1-deficient mice. Finally, we tested the hypothesis that other proteasome-dependent pathways not known to be associated with ciliopathies are defective in the absence of ciliopathy proteins. We found that loss of BBS1, BBS4, or OFD1 led to decreased NF-κB activity and concomitant IκBβ accumulation and that these defects were ameliorated with SFN treatment. Taken together, our data indicate that basal body proteasomal regulation governs paracrine signaling pathways and suggest that augmenting proteasomal function might benefit ciliopathy patients. PMID:24691443

Low noise signal-to-noise ratio enhancing readout circuit for current-mediated active pixel sensors

International Nuclear Information System (INIS)

Ottaviani, Tony; Karim, Karim S.; Nathan, Arokia; Rowlands, John A.

2006-01-01

Diagnostic digital fluoroscopic applications continuously expose patients to low doses of x-ray radiation, posing a challenge to both the digital imaging pixel and readout electronics when amplifying small signal x-ray inputs. Traditional switch-based amorphous silicon imaging solutions, for instance, have produced poor signal-to-noise ratios (SNRs) at low exposure levels owing to noise sources from the pixel readout circuitry. Current-mediated amorphous silicon pixels are an improvement over conventional pixel amplifiers with an enhanced SNR across the same low-exposure range, but whose output also becomes nonlinear with increasing dosage. A low-noise SNR enhancing readout circuit has been developed that enhances the charge gain of the current-mediated active pixel sensor (C-APS). The solution takes advantage of the current-mediated approach, primarily integrating the signal input at the desired frequency necessary for large-area imaging, while adding minimal noise to the signal readout. Experimental data indicates that the readout circuit can detect pixel outputs over a large bandwidth suitable for real-time digital diagnostic x-ray fluoroscopy. Results from hardware testing indicate that the minimum achievable C-APS output current that can be discerned at the digital fluoroscopic output from the enhanced SNR readout circuit is 0.341 nA. The results serve to highlight the applicability of amorphous silicon current-mediated pixel amplifiers for large-area flat panel x-ray imagers

Identification of a novel Gnao-mediated alternate olfactory signaling pathway in murine OSNs

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Paul eScholz

2016-03-01

Full Text Available It is generally agreed that in olfactory sensory neurons (OSNs, the binding of odorant molecules to their specific olfactory receptor (OR triggers a cAMP-dependent signaling cascade, activating cyclic-nucleotide gated (CNG channels. However, considerable controversy dating back more than 20 years has surrounded the question of whether alternate signaling plays a role in mammalian olfactory transduction. In this study, we demonstrate a specific alternate signaling pathway in Olfr73-expressing OSNs. Methylisoeugenol (MIEG and at least one other known weak Olfr73 agonist (Raspberry Ketone trigger a signaling cascade independent from the canonical pathway, leading to the depolarization of the cell. Interestingly, this pathway is mediated by Gnao activation, leading to Cl- efflux; however, the activation of adenylyl cyclase III (ACIII, the recruitment of Ca2+ from extra-or intracellular stores, and phosphatidylinositol 3-kinase-dependent signaling (PI signaling are not involved. Furthermore, we demonstrated that our newly identified pathway coexists with the canonical olfactory cAMP pathway in the same OSN and can be triggered by the same OR in a ligand-selective manner. We suggest that this pathway might reflect a mechanism for odor recognition predominantly used in early developmental stages before olfactory cAMP signaling is fully developed. Taken together, our findings support the existence of at least one odor-induced alternate signal transduction pathway in native OSNs mediated by Olfr73 in a ligand-selective manner.

Formation of intestinal atresias in the Fgfr2IIIb-/- mice is not associated with defects in notochord development or alterations in Shh expression.

Science.gov (United States)

Reeder, Amy L; Botham, Robert A; Franco, Marta; Zaremba, Krzysztof M; Nichol, Peter F

2012-09-01

The etiology of intestinal atresia remains elusive but has been ascribed to a number of possible events including in utero vascular accidents, failure of recanalization of the intestinal lumen, and mechanical compression. Another such event that has been postulated to be a cause in atresia formation is disruption in notochord development. This hypothesis arose from clinical observations of notochord abnormalities in patients with intestinal atresias as well as abnormal notochord development observed in a pharmacologic animal model of intestinal atresia. Atresias in this model result from in utero exposure to Adriamycin, wherein notochord defects were noted in up to 80% of embryos that manifested intestinal atresias. Embryos with notochord abnormalities were observed to have ectopic expression of Sonic Hedgehog (Shh), which in turn was postulated to be causative in atresia formation. We were interested in determining whether disruptions in notochord development or Shh expression occurred in an established genetic model of intestinal atresia and used the fibroblast growth factor receptor 2IIIb homozygous mutant (Fgfr2IIIb-/-) mouse model. These embryos develop colonic atresias (100% penetrance) and duodenal atresias (42% penetrance). Wild-type and Fgfr2IIIb-/- mouse embryos were harvested at embryonic day (E) 10.5, E11.5, E12.5, and E13.5. Whole-mount in situ hybridization was performed on E10.5 embryos for Shh. Embryos at each time point were harvested and sectioned for hematoxylin-eosin staining. Sections were photographed specifically for the notochord and resulting images reconstructed in 3-D using Amira software. Colons were isolated from wild-type and Fgfr2IIIb-/- embryos at E10.5, then cultured for 48 hours in Matrigel with FGF10 in the presence or absence of exogenous Shh protein. Explants were harvested, fixed in formalin, and photographed. Fgfr2IIIb-/- mouse embryos exhibit no disruptions in Shh expression at E10.5, when the first events in atresia

Methods for the Analysis of Protein Phosphorylation-Mediated Cellular Signaling Networks

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White, Forest M.; Wolf-Yadlin, Alejandro

2016-06-01

Protein phosphorylation-mediated cellular signaling networks regulate almost all aspects of cell biology, including the responses to cellular stimulation and environmental alterations. These networks are highly complex and comprise hundreds of proteins and potentially thousands of phosphorylation sites. Multiple analytical methods have been developed over the past several decades to identify proteins and protein phosphorylation sites regulating cellular signaling, and to quantify the dynamic response of these sites to different cellular stimulation. Here we provide an overview of these methods, including the fundamental principles governing each method, their relative strengths and weaknesses, and some examples of how each method has been applied to the analysis of complex signaling networks. When applied correctly, each of these techniques can provide insight into the topology, dynamics, and regulation of protein phosphorylation signaling networks.

Deformed Skull Morphology Is Caused by the Combined Effects of the Maldevelopment of Calvarias, Cranial Base and Brain in FGFR2-P253R Mice Mimicking Human Apert Syndrome.

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Luo, Fengtao; Xie, Yangli; Xu, Wei; Huang, Junlan; Zhou, Siru; Wang, Zuqiang; Luo, Xiaoqing; Liu, Mi; Chen, Lin; Du, Xiaolan

2017-01-01

Apert syndrome (AS) is a common genetic syndrome in humans characterized with craniosynostosis. Apert patients and mouse models showed abnormalities in sutures, cranial base and brain, that may all be involved in the pathogenesis of skull malformation of Apert syndrome. To distinguish the differential roles of these components of head in the pathogenesis of the abnormal skull morphology of AS, we generated mouse strains specifically expressing mutant FGFR2 in chondrocytes, osteoblasts, and progenitor cells of central nervous system (CNS) by crossing Fgfr2 +/P253R-Neo mice with Col2a1-Cre, Osteocalcin-Cre (OC-Cre), and Nestin-Cre mice, respectively. We then quantitatively analyzed the skull and brain morphology of these mutant mice by micro-CT and micro-MRI using Euclidean distance matrix analysis (EDMA). Skulls of Col2a1-Fgfr2 +/P253R mice showed Apert syndrome-like dysmorphology, such as shortened skull dimensions along the rostrocaudal axis, shortened nasal bone, and evidently advanced ossification of cranial base synchondroses. The OC-Fgfr2 +/P253R mice showed malformation in face at 8-week stage. Nestin-Fgfr2 +/P253R mice exhibited increased dorsoventral height and rostrocaudal length on the caudal skull and brain at 8 weeks. Our study indicates that the abnormal skull morphology of AS is caused by the combined effects of the maldevelopment in calvarias, cranial base, and brain tissue. These findings further deepen our knowledge about the pathogenesis of the abnormal skull morphology of AS, and provide new clues for the further analyses of skull phenotypes and clinical management of AS.

Adaptive and innate immune reactions regulating mast cell activation: from receptor-mediated signaling to responses

DEFF Research Database (Denmark)

Tkaczyk, Christine; Jensen, Bettina M; Iwaki, Shoko

2006-01-01

differentially activate multiple signaling pathways within the mast cells required for the generation and/or release of inflammatory mediators. Thus, the composition of the suite of mediators released and the physiologic ramifications of these responses are dependent on the stimuli and the microenvironment...

Proteomic analysis of cAMP-mediated signaling during differentiation of 3 T3-L1 preadipocytes

DEFF Research Database (Denmark)

Borkowski, Kamil; Wrzesinski, Krzysztow; Rogowska-Wrzesinska, Adelina

2014-01-01

Initiation of adipocyte differentiation is promoted by the synergistic action of insulin/insulin-like growth factor, glucocorticoids, and agents activating cAMP-dependent signaling. The action of cAMP is mediated via PKA and Epac, where at least part of the PKA function relates to strong repression...... a comprehensive evaluation of Epac-mediated processes and their interplay with PKA during the initiation of 3 T3-L1 preadipocyte differentiation using a combination of proteomics, molecular approaches, and bioinformatics. Proteomic analyses revealed 7 proteins specifically regulated in response to Epac activation......-dependent signaling thereby adding a novel facet to our understanding of cAMP-mediated potentiation of adipocyte differentiation....

The carboxyl terminal tyrosine 417 residue of NOK has an autoinhibitory effect on NOK-mediated signaling transductions

International Nuclear Information System (INIS)

Li Yinghua; Zhong Shan; Rong Zhili; Ren Yongming; Li Zhiyong; Zhang Shuping; Chang Zhijie; Liu Li

2007-01-01

Receptor protein tyrosine kinases (RPTKs) are essential mediators of cell growth, differentiation, migration, and metabolism. Recently, a novel RPTK named NOK has been cloned and characterized. In current study, we investigated the role of the carboxyl terminal tyrosine 417 residue of NOK in the activations of different signaling pathways. A single tyrosine to phenylalanine point mutation at Y417 site (Y417 F) not only dramatically enhanced the NOK-induced activation of extracellular signal-regulated kinase (ERK), but also markedly promoted the NOK-mediated activation of both signal transducer and activator of transcription 1 and 3 (STAT1 and 3). Moreover, the proliferation potential of NIH3T3-NOK (Y417F) stable cells were significantly elevated as compared with that of NIH3T3-NOK. Overall, our results demonstrate that the tyrosine Y417 residue at the carboxyl tail of NOK exhibits an autoinhibitory role in NOK-mediated signaling transductions

Ketogenic Diet Impairs FGF21 Signaling and Promotes Differential Inflammatory Responses in the Liver and White Adipose Tissue.

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Mohamed Asrih

Full Text Available Beside its beneficial effects on weight loss, ketogenic diet (KD causes dyslipidemia, a pro-inflammatory state involved in the development of hepatic steatosis, glucose intolerance and insulin resistance, although the latter is still being debated. Additionally, KD is known to increase fibroblast growth factor 21 (FGF21 plasma levels. However, FGF21 cannot initiate its beneficial actions on metabolism in these conditions. We therefore hypothesized and tested in the present study that KD may impair FGF21 signaling.Using indirect calorimetry, we found that KD-fed mice exhibited higher energy expenditure than regular chow (RC-fed mice associated with increased Ucp1 levels in white adipose tissue (WAT, along with increased plasma FGF21 levels. We then assessed the effect of KD on FGF21 signaling in both the liver and WAT. We found that Fgfr4 and Klb (β-klotho were downregulated in the liver, while Fgfr1 was downregulated in WAT of KD-fed mice. Because inflammation could be one of the mechanisms linking KD to impaired FGF21 signaling, we measured the expression levels of inflammatory markers and macrophage accumulation in WAT and liver and found an increased inflammation and macrophage accumulation in the liver, but surprisingly, a reduction of inflammation in WAT.We also showed that KD enhances lipid accumulation in the liver, which may explain hepatic inflammation and impaired Fgfr4 and Klb expression. In contrast, import of lipids from the circulation was significantly reduced in WAT of KD-fed mice, as suggested by a downregulation of Lpl and Cd36. This was further associated with reduced inflammation in WAT.Altogether, these results indicate that KD could be beneficial for a given tissue but deleterious for another.

The Bucket System – A computer mediated signaling system for group improvisation

DEFF Research Database (Denmark)

Dahlstedt, Palle; Nilsson, Per Anders; Robair, Gino

2015-01-01

The Bucket System is a new system for computer-mediated ensemble improvisation, designed by improvisers for improvisers. Coming from a tradition of structured free ensemble improvisation practices (comprovisation), influenced by post-WW2 experimental music practices, it is a signaling system...

Neuronal Orphan G-Protein Coupled Receptor Proteins Mediate Plasmalogens-Induced Activation of ERK and Akt Signaling.

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Md Shamim Hossain

Full Text Available The special glycerophospholipids plasmalogens (Pls are enriched in the brain and reported to prevent neuronal cell death by enhancing phosphorylation of Akt and ERK signaling in neuronal cells. Though the activation of Akt and ERK was found to be necessary for the neuronal cells survival, it was not known how Pls enhanced cellular signaling. To answer this question, we searched for neuronal specific orphan GPCR (G-protein coupled receptor proteins, since these proteins were believed to play a role in cellular signal transduction through the lipid rafts, where both Pls and some GPCRs were found to be enriched. In the present study, pan GPCR inhibitor significantly reduced Pls-induced ERK signaling in neuronal cells, suggesting that Pls could activate GPCRs to induce signaling. We then checked mRNA expression of 19 orphan GPCRs and 10 of them were found to be highly expressed in neuronal cells. The knockdown of these 10 neuronal specific GPCRs by short hairpin (sh-RNA lentiviral particles revealed that the Pls-mediated phosphorylation of ERK was inhibited in GPR1, GPR19, GPR21, GPR27 and GPR61 knockdown cells. We further found that the overexpression of these GPCRs enhanced Pls-mediated phosphorylation of ERK and Akt in cells. Most interestingly, the GPCRs-mediated cellular signaling was reduced significantly when the endogenous Pls were reduced. Our cumulative data, for the first time, suggest a possible mechanism for Pls-induced cellular signaling in the nervous system.

Silver Nanoparticles Induce HePG-2 Cells Apoptosis Through ROS-Mediated Signaling Pathways

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Zhu, Bing; Li, Yinghua; Lin, Zhengfang; Zhao, Mingqi; Xu, Tiantian; Wang, Changbing; Deng, Ning

2016-04-01

Recently, silver nanoparticles (AgNPs) have been shown to provide a novel approach to overcome tumors, especially those of hepatocarcinoma. However, the anticancer mechanism of silver nanoparticles is unclear. Thus, the purpose of this study was to estimate the effect of AgNPs on proliferation and activation of ROS-mediated signaling pathway on human hepatocellular carcinoma HePG-2 cells. A simple chemical method for preparing AgNPs with superior anticancer activity has been showed in this study. AgNPs were detected by transmission electronic microscopy (TEM) and energy dispersive X-ray (EDX). The size distribution and zeta potential of silver nanoparticles were detected by Zetasizer Nano. The average size of AgNPs (2 nm) observably increased the cellular uptake by endocytosis. AgNPs markedly inhibited the proliferation of HePG-2 cells through induction of apoptosis with caspase-3 activation and PARP cleavage. AgNPs with dose-dependent manner significantly increased the apoptotic cell population (sub-G1). Furthermore, AgNP-induced apoptosis was found dependent on the overproduction of reactive oxygen species (ROS) and affecting of MAPKs and AKT signaling and DNA damage-mediated p53 phosphorylation to advance HePG-2 cells apoptosis. Therefore, our results show that the mechanism of ROS-mediated signaling pathways may provide useful information in AgNP-induced HePG-2 cell apoptosis.

Diet-induced obesity mediated by the JNK/DIO2 signal transduction pathway

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Vernia, Santiago; Cavanagh-Kyros, Julie; Barrett, Tamera; Jung, Dae Young; Kim, Jason K.; Davis, Roger J.

2013-01-01

The cJun N-terminal kinase (JNK) signaling pathway is a key mediator of metabolic stress responses caused by consuming a high-fat diet, including the development of obesity. To test the role of JNK, we examined diet-induced obesity in mice with targeted ablation of Jnk genes in the anterior pituitary gland. These mice exhibited an increase in the pituitary expression of thyroid-stimulating hormone (TSH), an increase in the blood concentration of thyroid hormone (T4), increased energy expenditure, and markedly reduced obesity compared with control mice. The increased amount of pituitary TSH was caused by reduced expression of type 2 iodothyronine deiodinase (Dio2), a gene that is required for T4-mediated negative feedback regulation of TSH expression. These data establish a molecular mechanism that accounts for the regulation of energy expenditure and the development of obesity by the JNK signaling pathway. PMID:24186979

Changes in FGFR2 amino-acid residue Asn549 lead to Crouzon and Pfeiffer syndrome with hydrocephalus

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Caroline Apra

2016-10-01

Full Text Available Mutations in Fibroblast Growth Factor Receptor II (FGFR2 have been identified in patients with Crouzon and Pfeiffer syndrome, among which rare mutations of the intracellular tyrosine kinase domain. Correlating subtle phenotypes with each rare mutation is still in progress. In Necker-Enfants Malades Hospital, we identified three patients harboring three different pathogenic variants of the same amino acid residue Asn-549 located in this domain: in addition to a very typical crouzonoid appearance, they all developed clinically relevant hydrocephalus, which is an inconstant feature of Crouzon and Pfeiffer syndrome. Overall, FGFR2 tyrosine kinase domain mutations account for 5/67 (7.4% cases in our hospital. We describe a novel mutation, p.Asn549Ser, and new cases of p.Asn549His and p.Asn549Thr mutations, each reported once before. Our three cases of Asn-549 mutations, alongside with rare previously reported cases, show that these patients are at higher risk of hydrocephalus. Clinical and imaging follow-up, with possible early surgery, may help prevent secondary intellectual disability.

Detection of a de novo Y278C mutation in FGFR3 in a pregnancy with severe fetal hypochondroplasia: prenatal diagnosis and literature review.

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Chen, Chih-Ping; Su, Yi-Ning; Lin, Tzu-Hung; Chang, Tung-Yao; Su, Jun-Wei; Wang, Wayseen

2013-12-01

We describe a prenatal molecular diagnosis of hypochondroplasia (HCH) in a pregnancy not at risk of HCH and review the literature on prenatal diagnosis of HCH. A 28-year-old primigravid woman was referred for genetic counseling at 30 weeks of gestation because of short-limbed dwarfism in the fetus. The woman had a body height of 152 cm. Her husband had a body height of 180 cm. Level II ultrasound showed a normal amount of amniotic fluid and a singleton fetus with fetal biometry equivalent to 30 weeks except for short limbs. Fetal biometry measurements were as follows: biparietal diameter = 7.38 cm (30 weeks); head circumference = 28.14 cm (30 weeks); abdominal circumference (AC) = 24.64 cm (30 weeks); femur length (FL) = 3.97 cm ( 0.18); humerus = 3.64 cm (diagnosis of achondroplasia (ACH) was made. DNA testing for the FGFR3 gene and whole-genome array comparative genomic hybridization (aCGH) analysis were performed using cord blood DNA obtained by cordocentesis. FGFR3 mutation analysis revealed a de novo heterozygous c.833A > G, TAC > TGC transversion in exon 7 leading to a p.Tyr278Cys (Y278C) mutation in the FGFR3 protein. aCGH analysis revealed no genomic imbalance in cord blood. After delivery, the fetus had short limbs, a narrow thorax, brachydactyly, and relative macrocephaly. Cytogenetic analysis of cultured placental cells revealed a karyotype of 46,XX. Prenatal diagnosis of abnormal ultrasound findings suspicious of ACH should include a differential diagnosis of HCH by molecular analysis of FGFR3. Copyright © 2013. Published by Elsevier B.V.

Lentiviral-Mediated Transgene Expression Can Potentiate Intestinal Mesenchymal-Epithelial Signaling

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Kohn Aimee

2009-01-01

Full Text Available Abstract Mesenchymal-epithelial signaling is essential for the development of many organs and is often disrupted in disease. In this study, we demonstrate the use of lentiviral-mediated transgene delivery as an effective approach for ectopic transgene expression and an alternative to generation of transgenic animals. One benefit to this approach is that it can be used independently or in conjunction with established transgenic or knockout animals for studying modulation of mesenchymal-epithelial interactions. To display the power of this approach, we explored ectopic expression of a Wnt ligand in the mouse intestinal mesenchyme and demonstrate its functional influence on the adjacent epithelium. Our findings highlight the efficient use of lentiviral-mediated transgene expression for modulating mesenchymal-epithelial interactions in vivo.

Lentiviral-Mediated Transgene Expression Can Potentiate Intestinal Mesenchymal-Epithelial Signaling

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Dismuke Adria D

2009-07-01

Full Text Available Abstract Mesenchymal-epithelial signaling is essential for the development of many organs and is often disrupted in disease. In this study, we demonstrate the use of lentiviral-mediated transgene delivery as an effective approach for ectopic transgene expression and an alternative to generation of transgenic animals. One benefit to this approach is that it can be used independently or in conjunction with established transgenic or knockout animals for studying modulation of mesenchymal-epithelial interactions. To display the power of this approach, we explored ectopic expression of a Wnt ligand in the mouse intestinal mesenchyme and demonstrate its functional influence on the adjacent epithelium. Our findings highlight the efficient use of lentiviral-mediated transgene expression for modulating mesenchymal-epithelial interactions in vivo.

Intercellular and intracellular signaling pathways mediating ionizing radiation-induced bystander effects

International Nuclear Information System (INIS)

Hamada, Nobuyuki; Hara, Takamitsu; Kobayashi, Yasuhiko; Matsumoto, Hideki

2007-01-01

A rapidly growing body of experimental evidence indicates that ionizing radiation induces biological effects in non-irradiated bystander cells that have received signals from adjacent or distant irradiated cells. This phenomenon, which has been termed the ionizing radiation-induced bystander effect, challenges the long-standing paradigm that radiation traversal through the nucleus of a cell is a prerequisite to elicit genetic damage or a biological response. Bystander effects have been observed in a number of experimental systems, and cells whose nucleus or cytoplasm is irradiated exert bystander responses. Bystander cells manifest a multitude of biological consequences, such as genetic and epigenetic changes, alterations in gene expression, activation of signal transduction pathways, and delayed effects in their progeny. Several mediating mechanisms have been proposed. These involve gap junction-mediated intercellular communication, secreted soluble factors, oxidative metabolism, plasma membrane-bound lipid rafts, and calcium fluxes. This paper reviews briefly the current knowledge of the bystander effect with a focus on proposed mechanisms. The potential benefit of bystander effects to cancer radiotherapy will also be discussed. (author)

Essential roles of Gab1 tyrosine phosphorylation in growth factor-mediated signaling and angiogenesis.

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Wang, Weiye; Xu, Suowen; Yin, Meimei; Jin, Zheng Gen

2015-02-15

Growth factors and their downstream receptor tyrosine kinases (RTKs) mediate a number of biological processes controlling cell function. Adaptor (docking) proteins, which consist exclusively of domains and motifs that mediate molecular interactions, link receptor activation to downstream effectors. Recent studies have revealed that Grb2-associated-binders (Gab) family members (including Gab1, Gab2, and Gab3), when phosphorylated on tyrosine residues, provide binding sites for multiple effector proteins, such as Src homology-2 (SH2)-containing protein tyrosine phosphatase 2 (SHP2) and phosphatidylinositol 3-kinase (PI3K) regulatory subunit p85, thereby playing important roles in transducing RTKs-mediated signals into pathways with diversified biological functions. Here, we provide an up-to-date overview on the domain structure and biological functions of Gab1, the most intensively studied Gab family protein, in growth factor signaling and biological functions, with a special focus on angiogenesis. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Screening of mutations affecting protein stability and dynamics of FGFR1—A simulation analysis

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C. George Priya Doss

2012-12-01

Full Text Available Single amino acid substitutions in Fibroblast Growth Factor Receptor 1 (FGFR1 destabilize protein and have been implicated in several genetic disorders like various forms of cancer, Kallamann syndrome, Pfeiffer syndrome, Jackson Weiss syndrome, etc. In order to gain functional insight into mutation caused by amino acid substitution to protein function and expression, special emphasis was laid on molecular dynamics simulation techniques in combination with in silico tools such as SIFT, PolyPhen 2.0, I-Mutant 3.0 and SNAP. It has been estimated that 68% nsSNPs were predicted to be deleterious by I-Mutant, slightly higher than SIFT (37%, PolyPhen 2.0 (61% and SNAP (58%. From the observed results, P722S mutation was found to be most deleterious by comparing results of all in silico tools. By molecular dynamics approach, we have shown that P722S mutation leads to increase in flexibility, and deviated more from the native structure which was supported by the decrease in the number of hydrogen bonds. In addition, biophysical analysis revealed a clear insight of stability loss due to P722S mutation in FGFR1 protein. Majority of mutations predicted by these in silico tools were in good concordance with the experimental results.

β-Arrestin-2-Dependent Signaling Promotes CCR4-mediated Chemotaxis of Murine T-Helper Type 2 Cells.

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Lin, Rui; Choi, Yeon Ho; Zidar, David A; Walker, Julia K L

2018-06-01

Allergic asthma is a complex inflammatory disease that leads to significant healthcare costs and reduction in quality of life. Although many cell types are implicated in the pathogenesis of asthma, CD4 + T-helper cell type 2 (Th2) cells are centrally involved. We previously reported that the asthma phenotype is virtually absent in ovalbumin-sensitized and -challenged mice that lack global expression of β-arrestin (β-arr)-2 and that CD4 + T cells from these mice displayed significantly reduced CCL22-mediated chemotaxis. Because CCL22-mediated activation of CCR4 plays a role in Th2 cell regulation in asthmatic inflammation, we hypothesized that CCR4-mediated migration of CD4 + Th2 cells to the lung in asthma may use β-arr-dependent signaling. To test this hypothesis, we assessed the effect of various signaling inhibitors on CCL22-induced chemotaxis using in vitro-polarized primary CD4 + Th2 cells from β-arr2-knockout and wild-type mice. Our results show, for the first time, that CCL22-induced, CCR4-mediated Th2 cell chemotaxis is dependent, in part, on a β-arr2-dependent signaling pathway. In addition, we show that this chemotactic signaling mechanism involves activation of P-p38 and Rho-associated protein kinase. These findings point to a proinflammatory role for β-arr2-dependent signaling and support β-arr2 as a novel therapeutic target in asthma.

The effects of tissue-non-specific alkaline phosphatase gene therapy on craniosynostosis and craniofacial morphology in the FGFR2C342Y/+ mouse model of Crouzon craniosynostosis.

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Wang, E; Nam, H K; Liu, J; Hatch, N E

2015-04-01

Craniosynostosis, the premature fusion of cranial bones, has traditionally been described as a disease of increased bone mineralization. However, multiple mouse models of craniosynostosis display craniosynostosis simultaneously with diminished cranial bone volume and/or density. We propose an alternative hypothesis that craniosynostosis results from abnormal tissue mineralization through the downregulation of tissue-non-specific alkaline phosphatase (TNAP) enzyme downstream of activating mutations in FGFRs. Neonatal Crouzon (FGFRC342Y/+) and wild-type (FGFR+/+) mice were injected with lentivirus to deliver a recombinant form of TNAP. Mice were sacrificed at 4 weeks postnatal. Serum was collected to test for alkaline phosphatase (AP), phosphorus, and calcium levels. Craniofacial bone fusion and morphology were assessed by micro-computed tomography. Injection with the TNAP lentivirus significantly increased serum AP levels (increased serum AP levels are indicative of efficient transduction and production of the recombinant protein), but results were variable and dependent upon viral lot and the litter of mice injected. Morphological analysis revealed craniofacial form differences for inferior surface (p=0.023) and cranial height (p=0.014) regions between TNAP lentivirus-injected and vehicle-injected Crouzon mice. With each unit increase in AP level, the odds of lambdoid suture fusion decreased by 84.2% and these results came close to statistical significance (p=0.068). These results suggest that TNAP deficiency may mediate FGFR2-associated craniosynostosis. Future studies should incorporate injection of recombinant TNAP protein, to avoid potential side effects and variable efficacy of lentiviral gene delivery. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Antioxidant role of glutathione S-transferases: 4-Hydroxynonenal, a key molecule in stress-mediated signaling

International Nuclear Information System (INIS)

Singhal, Sharad S.; Singh, Sharda P.; Singhal, Preeti; Horne, David; Singhal, Jyotsana; Awasthi, Sanjay

2015-01-01

4-Hydroxy-2-trans-nonenal (4HNE), one of the major end products of lipid peroxidation (LPO), has been shown to induce apoptosis in a variety of cell lines. It appears to modulate signaling processes in more than one way because it has been suggested to have a role in signaling for differentiation and proliferation. It has been known that glutathione S-transferases (GSTs) can reduce lipid hydroperoxides through their Se-independent glutathione-peroxidase activity and that these enzymes can also detoxify LPO end-products such as 4HNE. Available evidence from earlier studies together with results of recent studies in our laboratories strongly suggests that LPO products, particularly hydroperoxides and 4HNE, are involved in the mechanisms of stress-mediated signaling and that it can be modulated by the alpha-class GSTs through the regulation of the intracellular concentrations of 4HNE. We demonstrate that 4HNE induced apoptosis in various cell lines is accompanied with c-Jun-N-terminal kinase (JNK) and caspase-3 activation. Cells exposed to mild, transient heat or oxidative stress acquire the capacity to exclude intracellular 4HNE at a faster rate by inducing GSTA4-4 which conjugates 4HNE to glutathione (GSH), and RLIP76 which mediates the ATP-dependent transport of the GSH-conjugate of 4HNE (GS-HNE). The balance between formation and exclusion promotes different cellular processes — higher concentrations of 4HNE promote apoptosis; whereas, lower concentrations promote proliferation. In this article, we provide a brief summary of the cellular effects of 4HNE, followed by a review of its GST-catalyzed detoxification, with an emphasis on the structural attributes that play an important role in the interactions with alpha-class GSTA4-4. Taken together, 4HNE is a key signaling molecule and that GSTs being determinants of its intracellular concentrations, can regulate stress-mediated signaling, are reviewed in this article. - Highlights: • GSTs are the major

Construction and Deciphering of Human Phosphorylation-Mediated Signaling Transduction Networks.

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Zhang, Menghuan; Li, Hong; He, Ying; Sun, Han; Xia, Li; Wang, Lishun; Sun, Bo; Ma, Liangxiao; Zhang, Guoqing; Li, Jing; Li, Yixue; Xie, Lu

2015-07-02

Protein phosphorylation is the most abundant reversible covalent modification. Human protein kinases participate in almost all biological pathways, and approximately half of the kinases are associated with disease. PhoSigNet was designed to store and display human phosphorylation-mediated signal transduction networks, with additional information related to cancer. It contains 11 976 experimentally validated directed edges and 216 871 phosphorylation sites. Moreover, 3491 differentially expressed proteins in human cancer from dbDEPC, 18 907 human cancer variation sites from CanProVar, and 388 hyperphosphorylation sites from PhosphoSitePlus were collected as annotation information. Compared with other phosphorylation-related databases, PhoSigNet not only takes the kinase-substrate regulatory relationship pairs into account, but also extends regulatory relationships up- and downstream (e.g., from ligand to receptor, from G protein to kinase, and from transcription factor to targets). Furthermore, PhoSigNet allows the user to investigate the impact of phosphorylation modifications on cancer. By using one set of in-house time series phosphoproteomics data, the reconstruction of a conditional and dynamic phosphorylation-mediated signaling network was exemplified. We expect PhoSigNet to be a useful database and analysis platform benefiting both proteomics and cancer studies.

Computational mouse atlases and their application to automatic assessment of craniofacial dysmorphology caused by the Crouzon mutation Fgfr2

DEFF Research Database (Denmark)

Ólafsdóttir, Hildur; Darvann, Tron Andre; Hermann, Nuno V.

2007-01-01

Crouzon syndrome is characterised by premature fusion of sutures and synchondroses. Recently the first mouse model of the syndrome was generated, having the mutation Cys342Tyr in Fgfr2c, equivalent to the most common human Crouzon/Pfeiffer syndrome mutation. In this study, a set of Micro CT scann....... Furthermore, the nonrigid approach is essential when it comes to analysing local, nonlinear shape differences.......Crouzon syndrome is characterised by premature fusion of sutures and synchondroses. Recently the first mouse model of the syndrome was generated, having the mutation Cys342Tyr in Fgfr2c, equivalent to the most common human Crouzon/Pfeiffer syndrome mutation. In this study, a set of Micro CT....... Subsequently, the atlas was deformed to match each subject from the two groups of mice. The accuracy of these registrations was measured by a comparison of manually placed landmarks from two different observers and automatically assessed landmarks. Both of the automatic approaches were within the inter...

Varicose and cheerio collaborate with pebble to mediate semaphorin-1a reverse signaling in Drosophila.

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Jeong, Sangyun; Yang, Da-Som; Hong, Young Gi; Mitchell, Sarah P; Brown, Matthew P; Kolodkin, Alex L

2017-09-26

The transmembrane semaphorin Sema-1a acts as both a ligand and a receptor to regulate axon-axon repulsion during neural development. Pebble (Pbl), a Rho guanine nucleotide exchange factor, mediates Sema-1a reverse signaling through association with the N-terminal region of the Sema-1a intracellular domain (ICD), resulting in cytoskeletal reorganization. Here, we uncover two additional Sema-1a interacting proteins, varicose (Vari) and cheerio (Cher), each with neuronal functions required for motor axon pathfinding. Vari is a member of the membrane-associated guanylate kinase (MAGUK) family of proteins, members of which can serve as scaffolds to organize signaling complexes. Cher is related to actin filament cross-linking proteins that regulate actin cytoskeleton dynamics. The PDZ domain binding motif found in the most C-terminal region of the Sema-1a ICD is necessary for interaction with Vari, but not Cher, indicative of distinct binding modalities. Pbl/Sema-1a-mediated repulsive guidance is potentiated by both vari and cher Genetic analyses further suggest that scaffolding functions of Vari and Cher play an important role in Pbl-mediated Sema-1a reverse signaling. These results define intracellular components critical for signal transduction from the Sema-1a receptor to the cytoskeleton and provide insight into mechanisms underlying semaphorin-induced localized changes in cytoskeletal organization.

Interference with Intraepithelial TNF-α Signaling Inhibits CD8+ T-Cell-Mediated Lung Injury in Influenza Infection

OpenAIRE

Srikiatkhachorn, Anon; Chintapalli, Jyothi; Liu, Jun; Jamaluddin, Mohammad; Harrod, Kevin S.; Whitsett, Jeffrey A.; Enelow, Richard I.; Ramana, Chilakamarti V.

2010-01-01

CD8+ T-cell-mediated pulmonary immunopathology in respiratory virus infection is mediated in large part by antigen-specific TNF-α expression by antiviral effector T cells, which results in epithelial chemokine expression and inflammatory infiltration of the lung. To further define the signaling events leading to lung epithelial chemokine production in response to CD8+ T-cell antigen recognition, we expressed the adenoviral 14.7K protein, a putative inhibitor of TNF-α signaling, in the distal ...

Event generation and production of signal inputs for the search of dark matter mediator signal at a future hadron collider

CERN Document Server

Chalise, Darshan

2017-01-01

The interaction between Dark Matter particles and Standard Model particles is possible through a force mediated by a Dark Matter(DM) - Standard Model(SM) mediator. If that mediator decays through a dijet event, the reconstructed invariant mass of the jets will peak at a specific value, in contrast to the smooth QCD background. This analysis is a preliminary work towards the understanding of how changes in detector conditions at the Future Circular Collider affect the sensitivity of the mediator signal. MadGraph 5 was used to produce events with 30 TeV DM mediator and Heppy was used to produce flat n-tuples for ROOT analysis. MadAnalysis 5 was then used to produce histograms of MadGraph events and PyRoot was used to analyze Heppy output. Histograms of invariant mass of the jets after event production through MadGraph as well as after Heppy analysis showed a peak at 30 TeV. This verified the production of a 30 TeV mediator during event production.

Nitro-fatty acids in plant signaling: New key mediators of nitric oxide metabolism

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Capilla Mata-Pérez

2017-04-01

Full Text Available Recent studies in animal systems have shown that NO can interact with fatty acids to generate nitro-fatty acids (NO2-FAs. They are the product of the reaction between reactive nitrogen species and unsaturated fatty acids, and are considered novel mediators of cell signaling based mainly on a proven anti-inflammatory response. Although these signaling mediators have been described widely in animal systems, NO2-FAs have scarcely been studied in plants. Preliminary data have revealed the endogenous presence of free and protein-adducted NO2-FAs in extra-virgin olive oil (EVOO, which appear to be contributing to the cardiovascular benefits associated with the Mediterranean diet. Importantly, new findings have displayed the endogenous occurrence of nitro-linolenic acid (NO2-Ln in the model plant Arabidopsis thaliana and the modulation of NO2-Ln levels throughout this plant's development. Furthermore, a transcriptomic analysis by RNA-seq technology established a clear signaling role for this molecule, demonstrating that NO2-Ln was involved in plant-defense response against different abiotic-stress conditions, mainly by inducing the chaperone network and supporting a conserved mechanism of action in both animal and plant defense processes. Thus, NO2-Ln levels significantly rose under several abiotic-stress conditions, highlighting the strong signaling role of these molecules in the plant-protection mechanism. Finally, the potential of NO2-Ln as a NO donor has recently been described both in vitro and in vivo. Jointly, this ability gives NO2-Ln the potential to act as a signaling molecule by the direct release of NO, due to its capacity to induce different changes mediated by NO or NO-related molecules such as nitration and S-nitrosylation, or by the electrophilic capacity of these molecules through a nitroalkylation mechanism. Here, we describe the current state of the art regarding the advances performed in the field of NO2-FAs in plants and their

Augmented BMPRIA-mediated BMP signaling in cranial neural crest lineage leads to cleft palate formation and delayed tooth differentiation.

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Lu Li

Full Text Available The importance of BMP receptor Ia (BMPRIa mediated signaling in the development of craniofacial organs, including the tooth and palate, has been well illuminated in several mouse models of loss of function, and by its mutations associated with juvenile polyposis syndrome and facial defects in humans. In this study, we took a gain-of-function approach to further address the role of BMPR-IA-mediated signaling in the mesenchymal compartment during tooth and palate development. We generated transgenic mice expressing a constitutively active form of BmprIa (caBmprIa in cranial neural crest (CNC cells that contributes to the dental and palatal mesenchyme. Mice bearing enhanced BMPRIa-mediated signaling in CNC cells exhibit complete cleft palate and delayed odontogenic differentiation. We showed that the cleft palate defect in the transgenic animals is attributed to an altered cell proliferation rate in the anterior palatal mesenchyme and to the delayed palatal elevation in the posterior portion associated with ectopic cartilage formation. Despite enhanced activity of BMP signaling in the dental mesenchyme, tooth development and patterning in transgenic mice appeared normal except delayed odontogenic differentiation. These data support the hypothesis that a finely tuned level of BMPRIa-mediated signaling is essential for normal palate and tooth development.

Endothelial Dysfunction in Human Diabetes Is Mediated by Wnt5a-JNK Signaling.

Science.gov (United States)

Bretón-Romero, Rosa; Feng, Bihua; Holbrook, Monika; Farb, Melissa G; Fetterman, Jessica L; Linder, Erika A; Berk, Brittany D; Masaki, Nobuyuki; Weisbrod, Robert M; Inagaki, Elica; Gokce, Noyan; Fuster, Jose J; Walsh, Kenneth; Hamburg, Naomi M

2016-03-01

Endothelial dysfunction is linked to insulin resistance, inflammatory activation, and increased cardiovascular risk in diabetes mellitus; however, the mechanisms remain incompletely understood. Recent studies have identified proinflammatory signaling of wingless-type family member (Wnt) 5a through c-jun N-terminal kinase (JNK) as a regulator of metabolic dysfunction with potential relevance to vascular function. We sought to gain evidence that increased activation of Wnt5a-JNK signaling contributes to impaired endothelial function in patients with diabetes mellitus. We measured flow-mediated dilation of the brachial artery and characterized freshly isolated endothelial cells by protein expression, eNOS activation, and nitric oxide production in 85 subjects with type 2 diabetes mellitus (n=42) and age- and sex-matched nondiabetic controls (n=43) and in human aortic endothelial cells treated with Wnt5a. Endothelial cells from patients with diabetes mellitus displayed 1.3-fold higher Wnt5a levels (P=0.01) along with 1.4-fold higher JNK activation (P<0.01) without a difference in total JNK levels. Higher JNK activation was associated with lower flow-mediated dilation, consistent with endothelial dysfunction (r=0.53, P=0.02). Inhibition of Wnt5a and JNK signaling restored insulin and A23187-mediated eNOS activation and improved nitric oxide production in endothelial cells from patients with diabetes mellitus. In endothelial cells from nondiabetic controls, rWnt5a treatment inhibited eNOS activation replicating the diabetic endothelial phenotype. In human aortic endothelial cells, Wnt5a-induced impairment of eNOS activation and nitric oxide production was reversed by Wnt5a and JNK inhibition. Our findings demonstrate that noncanonical Wnt5a signaling and JNK activity contribute to vascular insulin resistance and endothelial dysfunction and may represent a novel therapeutic opportunity to protect the vasculature in patients with diabetes mellitus. © 2016 American Heart

DMPD: Signal transduction pathways mediated by the interaction of CpG DNA withToll-like receptor 9. [Dynamic Macrophage Pathway CSML Database

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Full Text Available 14751759 Signal transduction pathways mediated by the interaction of CpG DNA withTo...;16(1):17-22. (.png) (.svg) (.html) (.csml) Show Signal transduction pathways mediated by the interaction of... CpG DNA withToll-like receptor 9. PubmedID 14751759 Title Signal transduction pathways media

Electrochemical DNA biosensor based on MNAzyme-mediated signal amplification

International Nuclear Information System (INIS)

Diao, Wei; Tang, Min; Ding, Xiaojuan; Zhang, Ye; Yang, Jianru; Cheng, Wenbin; Mo, Fei; Wen, Bo; Xu, Lulu; Yan, Yurong

2016-01-01

The authors describe an electrochemical sensing strategy for highly sensitive and specific detection of target (analyte) DNA based on an amplification scheme mediated by a multicomponent nucleic acid enzyme (MNAzyme). MNAzymes were formed by multicomponent complexes which produce amplified “output” signals in response to specific “input” signal. In the presence of target nucleic acid, multiple partial enzymes (partzymes) oligonucleotides are assembled to form active MNAzymes. These can cleave H0 substrate into two pieces, thereby releasing the activated MNAzyme to undergo an additional cycle of amplification. Here, the two pieces contain a biotin-tagged sequence and a byproduct. The biotin-tagged sequences are specifically captured by the detection probes immobilized on the gold electrode. By employing streptavidinylated alkaline phosphatase as an enzyme label, an electrochemical signal is obtained. The electrode, if operated at a working potential of 0.25 V (vs. Ag/AgCl) in solution of pH 7.5, covers the 100 pM to 0.25 μM DNA concentration range, with a 79 pM detection limit. In our perception, the strategy introduced here has a wider potential in that it may be applied to molecular diagnostics and pathogen detection. (author)

Insulin signaling mediates sexual attractiveness in Drosophila.

Directory of Open Access Journals (Sweden)

Tsung-Han Kuo

Full Text Available Sexually attractive characteristics are often thought to reflect an individual's condition or reproductive potential, but the underlying molecular mechanisms through which they do so are generally unknown. Insulin/insulin-like growth factor signaling (IIS is known to modulate aging, reproduction, and stress resistance in several species and to contribute to variability of these traits in natural populations. Here we show that IIS determines sexual attractiveness in Drosophila through transcriptional regulation of genes involved in the production of cuticular hydrocarbons (CHC, many of which function as pheromones. Using traditional gas chromatography/mass spectrometry (GC/MS together with newly introduced laser desorption/ionization orthogonal time-of-flight mass spectrometry (LDI-MS we establish that CHC profiles are significantly affected by genetic manipulations that target IIS. Manipulations that reduce IIS also reduce attractiveness, while females with increased IIS are significantly more attractive than wild-type animals. IIS effects on attractiveness are mediated by changes in CHC profiles. Insulin signaling influences CHC through pathways that are likely independent of dFOXO and that may involve the nutrient-sensing Target of Rapamycin (TOR pathway. These results suggest that the activity of conserved molecular regulators of longevity and reproductive output may manifest in different species as external characteristics that are perceived as honest indicators of fitness potential.

Protein kinase D1 signaling in angiogenic gene expression and VEGF-mediated angiogenesis

Directory of Open Access Journals (Sweden)

Bin eRen MD, Phd, FAHA

2016-05-01

Full Text Available Protein kinase D 1 (PKD-1 is a signaling kinase important in fundamental cell functions including migration, proliferation and differentiation. PKD-1 is also a key regulator of gene expression and angiogenesis that is essential for cardiovascular development and tumor progression. Further understanding molecular aspects of PKD-1 signaling in the regulation of angiogenesis may have translational implications in obesity, cardiovascular disease and cancer. The author will summarize and provide the insights into molecular mechanisms by which PKD-1 regulates transcriptional expression of angiogenic genes, focusing on the transcriptional regulation of CD36 by PKD-1-FoxO1 signaling axis along with the potential implications of this axis in arterial differentiation and morphogenesis. He will also discuss a new concept of dynamic balance between proangiogenic and antiangiogenic signaling in determining angiogenic switch, and stress how PKD-1 signaling regulates VEGF signaling-mediated angiogenesis.

Inhibition of fibroblast growth factor receptor 3-dependent lung adenocarcinoma with a human monoclonal antibody

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Yongjun Yin

2016-05-01

Full Text Available Activating mutations in fibroblast growth factor receptor 3 (FGFR3 have been identified in multiple types of human cancer and in congenital birth defects. In human lung cancer, fibroblast growth factor 9 (FGF9, a high-affinity ligand for FGFR3, is overexpressed in 10% of primary resected non-small cell lung cancer (NSCLC specimens. Furthermore, in a mouse model where FGF9 can be induced in lung epithelial cells, epithelial proliferation and ensuing tumorigenesis is dependent on FGFR3. To develop new customized therapies for cancers that are dependent on FGFR3 activation, we have used this mouse model to evaluate a human monoclonal antibody (D11 with specificity for the extracellular ligand-binding domain of FGFR3, that recognizes both human and mouse forms of the receptor. Here, we show that D11 effectively inhibits signaling through FGFR3 in vitro, inhibits the growth of FGFR3-dependent FGF9-induced lung adenocarcinoma in mice, and reduces tumor-associated morbidity. Given the potency of FGF9 in this mouse model and the absolute requirement for signaling through FGFR3, this study validates the D11 antibody as a potentially useful and effective reagent for treating human cancers or other pathologies that are dependent on activation of FGFR3.

Differential effects of FGFR2 mutations on syndactyly and cleft palate in Apert syndrome

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Slaney, S.F.; Oldridge, M.; Wilkie, A.O.M. [Univ. of Oxford (United Kingdom)] [and others

1996-05-01

Apert syndrome is a distinctive human malformation characterized by craniosynostosis and severe syndactyly of the hands and feet. It is caused by specific missense substitutions involving adjacent amino acids (Ser252Trp or Pro253Arg) in the linker between the second and third extracellular immunoglobulin domains of fibroblast growth factor receptor 2 (FGFR2). We have developed a simple PCR assay for these mutations in genomic DNA, based on the creation of novel SfiI and BstUI restriction sites. Analysis of DNA from 70 unrelated patients with Apert syndrome showed that 45 had the Ser252Trp mutation and 25 had the Pro253Arg mutation. Phenotypic differences between these two groups of patients were investigated. Significant differences were found for severity of syndactyly and presence of cleft palate. The syndactyly was more severe with the Pro253Arg mutation, for both the hands and the feet. In contrast, cleft palate was significantly more common in the Ser252Trp patients. No convincing differences were found in the prevalence of other malformations associated with Apert syndrome. We conclude that, although the phenotype attributable to the two mutations is very similar, there are subtle differences. The opposite trends for severity of syndactyly and cleft palate in relation to the two mutations may relate to the varying patterns of temporal and tissue-specific expression of different fibroblast growth factors, the ligands for FGFR2. 54 refs., 5 figs., 3 tabs.

The Mediator Kinase Module Restrains Epidermal Growth Factor Receptor Signaling and Represses Vulval Cell Fate Specification in Caenorhabditis elegans.

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Grants, Jennifer M; Ying, Lisa T L; Yoda, Akinori; You, Charlotte C; Okano, Hideyuki; Sawa, Hitoshi; Taubert, Stefan

2016-02-01

Cell signaling pathways that control proliferation and determine cell fates are tightly regulated to prevent developmental anomalies and cancer. Transcription factors and coregulators are important effectors of signaling pathway output, as they regulate downstream gene programs. In Caenorhabditis elegans, several subunits of the Mediator transcriptional coregulator complex promote or inhibit vulva development, but pertinent mechanisms are poorly defined. Here, we show that Mediator's dissociable cyclin dependent kinase 8 (CDK8) module (CKM), consisting of cdk-8, cic-1/Cyclin C, mdt-12/dpy-22, and mdt-13/let-19, is required to inhibit ectopic vulval cell fates downstream of the epidermal growth factor receptor (EGFR)-Ras-extracellular signal-regulated kinase (ERK) pathway. cdk-8 inhibits ectopic vulva formation by acting downstream of mpk-1/ERK, cell autonomously in vulval cells, and in a kinase-dependent manner. We also provide evidence that the CKM acts as a corepressor for the Ets-family transcription factor LIN-1, as cdk-8 promotes transcriptional repression by LIN-1. In addition, we find that CKM mutation alters Mediator subunit requirements in vulva development: the mdt-23/sur-2 subunit, which is required for vulva development in wild-type worms, is dispensable for ectopic vulva formation in CKM mutants, which instead display hallmarks of unrestrained Mediator tail module activity. We propose a model whereby the CKM controls EGFR-Ras-ERK transcriptional output by corepressing LIN-1 and by fine tuning Mediator specificity, thus balancing transcriptional repression vs. activation in a critical developmental signaling pathway. Collectively, these data offer an explanation for CKM repression of EGFR signaling output and ectopic vulva formation and provide the first evidence of Mediator CKM-tail module subunit crosstalk in animals. Copyright © 2016 by the Genetics Society of America.

Notch signaling mediates the age-associated decrease in adhesion of germline stem cells to the niche.

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Chen-Yuan Tseng

2014-12-01

Full Text Available Stem cells have an innate ability to occupy their stem cell niche, which in turn, is optimized to house stem cells. Organ aging is associated with reduced stem cell occupancy in the niche, but the mechanisms involved are poorly understood. Here, we report that Notch signaling is increased with age in Drosophila female germline stem cells (GSCs, and this results in their removal from the niche. Clonal analysis revealed that GSCs with low levels of Notch signaling exhibit increased adhesiveness to the niche, thereby out-competing their neighbors with higher levels of Notch; adhesiveness is altered through regulation of E-cadherin expression. Experimental enhancement of Notch signaling in GSCs hastens their age-dependent loss from the niche, and such loss is at least partially mediated by Sex lethal. However, disruption of Notch signaling in GSCs does not delay GSC loss during aging, and nor does it affect BMP signaling, which promotes self-renewal of GSCs. Finally, we show that in contrast to GSCs, Notch activation in the niche (which maintains niche integrity, and thus mediates GSC retention is reduced with age, indicating that Notch signaling regulates GSC niche occupancy both intrinsically and extrinsically. Our findings expose a novel role of Notch signaling in controlling GSC-niche adhesion in response to aging, and are also of relevance to metastatic cancer cells, in which Notch signaling suppresses cell adhesion.

Heterogeneity of Breast Cancer Associations with Common Genetic Variants in FGFR2 according to the Intrinsic Subtypes in Southern Han Chinese Women

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Huiying Liang

2015-01-01

Full Text Available GWAS have identified variation in the FGFR2 locus as risk factors for breast cancer. Validation studies, however, have shown inconsistent results by ethnics and pathological characteristics. To further explore this inconsistency and investigate the associations of FGFR2 variants with breast cancer according to intrinsic subtype (Luminal-A, Luminal-B, ER−&PR−&HER2+, and triple negative among Southern Han Chinese women, we genotyped rs1078806, rs1219648, rs2420946, rs2981579, and rs2981582 polymorphisms in 609 patients and 882 controls. Significant associations with breast cancer risk were observed for rs2420946, rs2981579, and rs2981582 with OR (95% CI per risk allele of 1.19 (1.03–1.39, 1.24 (1.07–1.43, and 1.17 (1.01–1.36, respectively. In subtype specific analysis, above three SNPs were significantly associated with increased Luminal-A risk in a dose-dependent manner Ptrend<0.01; however, only rs2981579 was associated with Luminal-B, and none were linked to ER−&PR− subtypes (ER−&PR−&HER2+ and triple negative. Haplotype analyses also identified common haplotypes significantly associated with luminal-like subtypes (Luminal-A and Luminal-B, but not with ER−&PR− subtypes. Our results suggest that associations of FGFR2 SNPs with breast cancer were heterogeneous according to intrinsic subtype. Future studies stratifying patients by their intrinsic subtypes will provide new insights into the complex genetic mechanisms underlying breast cancer.

Transcription Profiles Reveal Sugar and Hormone Signaling Pathways Mediating Flower Induction in Apple (Malus domestica Borkh.).

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Xing, Li-Bo; Zhang, Dong; Li, You-Mei; Shen, Ya-Wen; Zhao, Cai-Ping; Ma, Juan-Juan; An, Na; Han, Ming-Yu

2015-10-01

Flower induction in apple (Malus domestica Borkh.) is regulated by complex gene networks that involve multiple signal pathways to ensure flower bud formation in the next year, but the molecular determinants of apple flower induction are still unknown. In this research, transcriptomic profiles from differentiating buds allowed us to identify genes potentially involved in signaling pathways that mediate the regulatory mechanisms of flower induction. A hypothetical model for this regulatory mechanism was obtained by analysis of the available transcriptomic data, suggesting that sugar-, hormone- and flowering-related genes, as well as those involved in cell-cycle induction, participated in the apple flower induction process. Sugar levels and metabolism-related gene expression profiles revealed that sucrose is the initiation signal in flower induction. Complex hormone regulatory networks involved in cytokinin (CK), abscisic acid (ABA) and gibberellic acid pathways also induce apple flower formation. CK plays a key role in the regulation of cell formation and differentiation, and in affecting flowering-related gene expression levels during these processes. Meanwhile, ABA levels and ABA-related gene expression levels gradually increased, as did those of sugar metabolism-related genes, in developing buds, indicating that ABA signals regulate apple flower induction by participating in the sugar-mediated flowering pathway. Furthermore, changes in sugar and starch deposition levels in buds can be affected by ABA content and the expression of the genes involved in the ABA signaling pathway. Thus, multiple pathways, which are mainly mediated by crosstalk between sugar and hormone signals, regulate the molecular network involved in bud growth and flower induction in apple trees. © The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

IGF-1 signaling mediated cell-specific skeletal mechano-transduction.

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Tian, Faming; Wang, Yongmei; Bikle, Daniel D

2018-02-01

Mechanical loading preserves bone mass and stimulates bone formation, whereas skeletal unloading leads to bone loss. In addition to osteocytes, which are considered the primary sensor of mechanical load, osteoblasts, and bone specific mesenchymal stem cells also are involved. The skeletal response to mechanical signals is a complex process regulated by multiple signaling pathways including that of insulin-like growth factor-1 (IGF-1). Conditional osteocyte deletion of IGF-1 ablates the osteogenic response to mechanical loading. Similarly, osteocyte IGF-1 receptor (IGF-1R) expression is necessary for reloading-induced periosteal bone formation. Transgenic overexpression of IGF-1 in osteoblasts results in enhanced responsiveness to in vivo mechanical loading in mice, a response which is eliminated by osteoblastic conditional disruption of IGF-1 in vivo. Bone marrow derived stem cells (BMSC) from unloaded bone fail to respond to IGF-1 in vitro. IGF-1R is required for the transduction of a mechanical stimulus to downstream effectors, transduction which is lost when the IGF-1R is deleted. Although the molecular mechanisms are not yet fully elucidated, the IGF signaling pathway and its interactions with potentially interlinked signaling cascades involving integrins, the estrogen receptor, and wnt/β-catenin play an important role in regulating adaptive response of cancer bone cells to mechanical stimuli. In this review, we discuss recent advances investigating how IGF-1 and other interlinked molecules and signaling pathways regulate skeletal mechano-transduction involving different bone cells, providing an overview of the IGF-1 signaling mediated cell-specific response to mechanical stimuli. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:576-583, 2018. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

Calpain-mediated proteolysis of polycystin-1 C-terminus induces JAK2 and ERK signal alterations

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Kim, Hyunho [Transplantation Research Institute, Seoul National University Medical Research Center, Seoul (Korea, Republic of); Department of Medicine, University of Maryland, Baltimore, MD (United States); Kang, Ah-Young [Transplantation Research Institute, Seoul National University Medical Research Center, Seoul (Korea, Republic of); Department of Medicine, Program of Immunology, Graduate School, Seoul National University, Seoul (Korea, Republic of); Ko, Ah-ra [Clinical Research Center, Samsung Biomedical Research Institute, Seoul (Korea, Republic of); Park, Hayne Cho [Transplantation Research Institute, Seoul National University Medical Research Center, Seoul (Korea, Republic of); Research Coordination Center for Rare Diseases, Seoul National University Hospital, Seoul (Korea, Republic of); So, Insuk [Department of Physiology, Seoul National University College of Medicine, Seoul (Korea, Republic of); Park, Jong Hoon [Department of Biological Science, Sookmyung Women’s University, Seoul (Korea, Republic of); Cheong, Hae Il [Research Coordination Center for Rare Diseases, Seoul National University Hospital, Seoul (Korea, Republic of); Department of Pediatrics, Seoul National University Children’s Hospital, Seoul (Korea, Republic of); Kidney Research Institute, Medical Research Center, Seoul National University College of Medicine, Seoul (Korea, Republic of); Hwang, Young-Hwan [Research Coordination Center for Rare Diseases, Seoul National University Hospital, Seoul (Korea, Republic of); Department of Internal Medicine, Eulji General Hospital, Eulji University College of Medicine, Seoul (Korea, Republic of); and others

2014-01-01

Autosomal dominant polycystic kidney disease (ADPKD), a hereditary renal disease caused by mutations in PKD1 (85%) or PKD2 (15%), is characterized by the development of gradually enlarging multiple renal cysts and progressive renal failure. Polycystin-1 (PC1), PKD1 gene product, is an integral membrane glycoprotein which regulates a number of different biological processes including cell proliferation, apoptosis, cell polarity, and tubulogenesis. PC1 is a target of various proteolytic cleavages and proteosomal degradations, but its role in intracellular signaling pathways remains poorly understood. Herein, we demonstrated that PC1 is a novel substrate for μ- and m-calpains, which are calcium-dependent cysteine proteases. Overexpression of PC1 altered both Janus-activated kinase 2 (JAK2) and extracellular signal-regulated kinase (ERK) signals, which were independently regulated by calpain-mediated PC1 degradation. They suggest that the PC1 function on JAK2 and ERK signaling pathways might be regulated by calpains in response to the changes in intracellular calcium concentration. - Highlights: • Polycystin-1 is a target of ubiquitin-independent degradation by calpains. • The PEST domain is required for calpain-mediated degradation of polycystin-1. • Polycystin-1 may independently regulate JAK2 and ERK signaling pathways.

[Children with idiopathic hypogonadotropic hypogonadism: clinical data analysis and mutations analysis of KAL1 and FGFR1 gene].

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Qin, Miao; Gong, Chunxiu; Qi, Zhan; Wu, Di; Liu, Min; Gu, Yi; Cao, Bingyan; Li, Wenjing; Liang, Xuejun

2014-12-01

To summarize the clinical features of idiopathic hypogonadotropic hypogonadism (IHH) diagnosed during childhood, and detect mutations in KAL1 and FGFR1, acting as key clues for diagnoses. We collected and analyzed clinical data of 21 cases (including demographic data, chief complaint, history of present illness, family history, physical examination, laboratory tests and imaging studies, etc.) diagnosed with IHH from December 2008 to February 2013. Polymerase chain reaction and gene sequencing was applied to detect mutations on KAL1 and FGFR1. Fifty healthy unrelated individuals were choosen as controls. Of 21 patients with IHH, 19 were males and 2 females, they visited us initially from 8-17 years old, with an average of (13.58 ± 2.38) years old. Sixteen cases were KS patients (76%). One boy reported abnormal sense of smelling but having olfactory perfect picture on MRI; 2/19 male cases had no puberty when they were over 13-14 years old without abnormal external genitalia. 8/19 cases only had small penis, 8/19 had both of cryptorchidism and small penis, and the Case 2 also had hypospadias. One boy had cryptorchidism combined with a normal penis. Only 2 girls diagnosed as IHH who visited us because of no puberty signs when they were 13 and 16 years old, respectively. Other clinical manifestations included: one with gynecomastia, 2 had mental retardation, and one was deaf; one with high palatal arch; one with mirror-movement and one with left renal agenesis but normal renal function respectively. Laboratory tests showed that the basic testosterone (T) is low and with inappropriately low or normal gonadotropin hormones. The results of cases of standard human chorionic gonadotropin (HCG) test of 7 cases out of 19 male children's were normal (testosterone>1 100 ng/L), and another nine cases continued to complete the extended HCG test, and the testosterone levels of two of them (cases 6, 8) were still lower than 1 000 ng/L. Family history: the parents in 9/21 family had

Regulator of G-protein signaling-5 is a marker of hepatic stellate cells and expression mediates response to liver injury.

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Arya J Bahrami

Full Text Available Liver fibrosis is mediated by hepatic stellate cells (HSCs, which respond to a variety of cytokine and growth factors to moderate the response to injury and create extracellular matrix at the site of injury. G-protein coupled receptor (GPCR-mediated signaling, via endothelin-1 (ET-1 and angiotensin II (AngII, increases HSC contraction, migration and fibrogenesis. Regulator of G-protein signaling-5 (RGS5, an inhibitor of vasoactive GPCR agonists, functions to control GPCR-mediated contraction and hypertrophy in pericytes and smooth muscle cells (SMCs. Therefore we hypothesized that RGS5 controls GPCR signaling in activated HSCs in the context of liver injury. In this study, we localize RGS5 to the HSCs and demonstrate that Rgs5 expression is regulated during carbon tetrachloride (CCl4-induced acute and chronic liver injury in Rgs5LacZ/LacZ reporter mice. Furthermore, CCl4 treated RGS5-null mice develop increased hepatocyte damage and fibrosis in response to CCl4 and have increased expression of markers of HSC activation. Knockdown of Rgs5 enhances ET-1-mediated signaling in HSCs in vitro. Taken together, we demonstrate that RGS5 is a critical regulator of GPCR signaling in HSCs and regulates HSC activation and fibrogenesis in liver injury.

Mast cell chymase induces smooth muscle cell apoptosis by disrupting NF-κB-mediated survival signaling

International Nuclear Information System (INIS)

Leskinen, Markus J.; Heikkilae, Hanna M.; Speer, Mei Y.; Hakala, Jukka K.; Laine, Mika; Kovanen, Petri T.; Lindstedt, Ken A.

2006-01-01

Chymase released from activated mast cells induces apoptosis of vascular smooth muscle cells (SMCs) in vitro by degrading the pericellular matrix component fibronectin, so causing disruption of focal adhesion complexes and Akt dephosphorylation, which are necessary for cell adhesion and survival. However, the molecular mechanisms of chymase-mediated apoptosis downstream of Akt have remained elusive. Here, we show by means of RT-PCR, Western blotting, EMSA, immunocytochemistry and confocal microscopy, that chymase induces SMC apoptosis by disrupting NF-κB-mediated survival signaling. Following chymase treatment, the translocation of active NF-κB/p65 to the nucleus was partly abolished and the amount of nuclear p65 was reduced. Pretreatment of SMCs with chymase also inhibited LPS- and IL-1β-induced nuclear translocation of p65. The chymase-induced degradation of p65 was mediated by active caspases. Loss of NF-κB-mediated transactivation resulted in downregulation of bcl-2 mRNA and protein expression, leading to mitochondrial swelling and release of cytochrome c. The apoptotic process involved activation of both caspase 9 and caspase 8. The results reveal that, by disrupting the NF-κB-mediated survival-signaling pathway, activated chymase-secreting mast cells can mediate apoptosis of cultured arterial SMCs. Since activated mast cells colocalize with apoptotic SMCs in vulnerable areas of human atherosclerotic plaques, they may participate in the weakening and rupture of atherosclerotic plaques

A pivotal role of the jasmonic acid signal pathway in mediating radiation-induced bystander effects in Arabidopsis thaliana

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Wang, Ting; Xu, Wei; Deng, Chenguang; Xu, Shaoxin; Li, Fanghua; Wu, Yuejin; Wu, Lijun [Key Laboratory of Ion Beam Bioengineering, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031 (China); Key Laboratory of Environmental Toxicology and Pollution Control Technology of Anhui Province, Hefei 230031 (China); Bian, Po, E-mail: bianpo@ipp.ac.cn [Key Laboratory of Ion Beam Bioengineering, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031 (China); Key Laboratory of Environmental Toxicology and Pollution Control Technology of Anhui Province, Hefei 230031 (China)

2016-09-15

Highlights: • The JA signal pathway plays a pivotal role in mediating radiation-induced bystander effects in Arabidopsis thaliana. • The JA signal pathway is involved in both the generation of bystander signals in irradiated roots and radiation responses in bystander aerial plants. • Over-accumulation of endogenous JA enhances the radiosensitivity of plants in terms of RIBE. - Abstract: Although radiation-induced bystander effects (RIBE) in Arabidopsis thaliana have been well demonstrated in vivo, little is known about their underlying mechanisms, particularly with regard to the participating signaling molecules and signaling pathways. In higher plants, jasmonic acid (JA) and its bioactive derivatives are well accepted as systemic signal transducers that are produced in response to various environmental stresses. It is therefore speculated that the JA signal pathway might play a potential role in mediating radiation-induced bystander signaling of root-to-shoot. In the present study, pretreatment of seedlings with Salicylhydroxamic acid, an inhibitor of lipoxigenase (LOX) in JA biosynthesis, significantly suppressed RIBE-mediated expression of the AtRAD54 gene. After root irradiation, the aerial parts of A. thaliana mutants deficient in JA biosynthesis (aos) and signaling cascades (jar1-1) showed suppressed induction of the AtRAD54 and AtRAD51 genes and TSI and 180-bp repeats, which have been extensively used as endpoints of bystander genetic and epigenetic effects in plants. These results suggest an involvement of the JA signal pathway in the RIBE of plants. Using the root micro-grafting technique, the JA signal pathway was shown to participate in both the generation of bystander signals in irradiated root cells and radiation responses in the bystander aerial parts of plants. The over-accumulation of endogenous JA in mutant fatty acid oxygenation up-regulated 2 (fou2), in which mutation of the Two Pore Channel 1 (TPC1) gene up-regulates expression of the LOX

Convergent RANK- and c-Met-mediated signaling components predict survival of patients with prostate cancer: an interracial comparative study.

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Hu, Peizhen; Chung, Leland W K; Berel, Dror; Frierson, Henry F; Yang, Hua; Liu, Chunyan; Wang, Ruoxiang; Li, Qinlong; Rogatko, Andre; Zhau, Haiyen E

2013-01-01

We reported (PLoS One 6 (12):e28670, 2011) that the activation of c-Met signaling in RANKL-overexpressing bone metastatic LNCaP cell and xenograft models increased expression of RANK, RANKL, c-Met, and phosphorylated c-Met, and mediated downstream signaling. We confirmed the significance of the RANK-mediated signaling network in castration resistant clinical human prostate cancer (PC) tissues. In this report, we used a multispectral quantum dot labeling technique to label six RANK and c-Met convergent signaling pathway mediators simultaneously in formalin fixed paraffin embedded (FFPE) tissue specimens, quantify the intensity of each expression at the sub-cellular level, and investigated their potential utility as predictors of patient survival in Caucasian-American, African-American and Chinese men. We found that RANKL and neuropilin-1 (NRP-1) expression predicts survival of Caucasian-Americans with PC. A Gleason score ≥ 8 combined with nuclear p-c-Met expression predicts survival in African-American PC patients. Neuropilin-1, p-NF-κB p65 and VEGF are predictors for the overall survival of Chinese men with PC. These results collectively support interracial differences in cell signaling networks that can predict the survival of PC patients.

Distinct signalling properties of insulin receptor substrate (IRS)-1 and IRS-2 in mediating insulin/IGF-1 action.

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Rabiee, Atefeh; Krüger, Marcus; Ardenkjær-Larsen, Jacob; Kahn, C Ronald; Emanuelli, Brice

2018-07-01

Insulin/IGF-1 action is driven by a complex and highly integrated signalling network. Loss-of-function studies indicate that the major insulin/IGF-1 receptor substrate (IRS) proteins, IRS-1 and IRS-2, mediate different biological functions in vitro and in vivo, suggesting specific signalling properties despite their high degree of homology. To identify mechanisms contributing to the differential signalling properties of IRS-1 and IRS-2 in the mediation of insulin/IGF-1 action, we performed comprehensive mass spectrometry (MS)-based phosphoproteomic profiling of brown preadipocytes from wild type, IRS-1 -/- and IRS-2 -/- mice in the basal and IGF-1-stimulated states. We applied stable isotope labeling by amino acids in cell culture (SILAC) for the accurate quantitation of changes in protein phosphorylation. We found ~10% of the 6262 unique phosphorylation sites detected to be regulated by IGF-1. These regulated sites included previously reported substrates of the insulin/IGF-1 signalling pathway, as well as novel substrates including Nuclear Factor I X and Semaphorin-4B. In silico prediction suggests the protein kinase B (PKB), protein kinase C (PKC), and cyclin-dependent kinase (CDK) as the main mediators of these phosphorylation events. Importantly, we found preferential phosphorylation patterns depending on the presence of either IRS-1 or IRS-2, which was associated with specific sets of kinases involved in signal transduction downstream of these substrates such as PDHK1, MAPK3, and PKD1 for IRS-1, and PIN1 and PKC beta for IRS-2. Overall, by generating a comprehensive phosphoproteomic profile from brown preadipocyte cells in response to IGF-1 stimulation, we reveal both common and distinct insulin/IGF-1 signalling events mediated by specific IRS proteins. Copyright © 2018 Elsevier Inc. All rights reserved.

Covalent Targeting of Fibroblast Growth Factor Receptor Inhibits Metastatic Breast Cancer.

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Brown, Wells S; Tan, Li; Smith, Andrew; Gray, Nathanael S; Wendt, Michael K

2016-09-01

Therapeutic targeting of late-stage breast cancer is limited by an inadequate understanding of how tumor cell signaling evolves during metastatic progression and by the currently available small molecule inhibitors capable of targeting these processes. Herein, we demonstrate that both β3 integrin and fibroblast growth factor receptor-1 (FGFR1) are part of an epithelial-mesenchymal transition (EMT) program that is required to facilitate metastatic outgrowth in response to fibroblast growth factor-2 (FGF2). Mechanistically, β3 integrin physically disrupts an interaction between FGFR1 and E-cadherin, leading to a dramatic redistribution of FGFR1 subcellular localization, enhanced FGF2 signaling and increased three-dimensional (3D) outgrowth of metastatic breast cancer cells. This ability of β3 integrin to drive FGFR signaling requires the enzymatic activity of focal adhesion kinase (FAK). Consistent with these mechanistic data, we demonstrate that FGFR, β3 integrin, and FAK constitute a molecular signature capable of predicting decreased survival of patients with the basal-like subtype of breast cancer. Importantly, covalent targeting of a conserved cysteine in the P-loop of FGFR1-4 with our newly developed small molecule, FIIN-4, more effectively blocks 3D metastatic outgrowth as compared with currently available FGFR inhibitors. In vivo application of FIIN-4 potently inhibited the growth of metastatic, patient-derived breast cancer xenografts and murine-derived metastases growing within the pulmonary microenvironment. Overall, the current studies demonstrate that FGFR1 works in concert with other EMT effector molecules to drive aberrant downstream signaling, and that these events can be effectively targeted using our novel therapeutics for the treatment of the most aggressive forms of breast cancer. Mol Cancer Ther; 15(9); 2096-106. ©2016 AACR. ©2016 American Association for Cancer Research.

DNA and protein co-administration induces tolerogenic dendritic cells through DC-SIGN mediated negative signals.

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Li, Jinyao; Geng, Shuang; Liu, Xiuping; Liu, Hu; Jin, Huali; Liu, Chang-Gong; Wang, Bin

2013-10-01

We previously demonstrated that DNA and protein co-administration induced differentiation of immature dendritic cells (iDCs) into CD11c(+)CD40(low)IL-10(+) regulatory DCs (DCregs) via the caveolin-1 (Cav-1) -mediated signal pathway. Here, we demonstrate that production of IL-10 and the low expression of CD40 play a critical role in the subsequent induction of regulatory T cells (Tregs) by the DCregs. We observed that DNA and protein were co-localized with DC-SIGN in caveolae and early lysosomes in the treated DCs, as indicated by co-localization with Cav-1 and EEA-1 compartment markers. DNA and protein also co-localized with LAMP-2. Gene-array analysis of gene expression showed that more than a thousand genes were significantly changed by the DC co-treatment with DNA + protein compared with controls. Notably, the level of DC-SIGN expression was dramatically upregulated in pOVA + OVA co-treated DCs. The expression levels of Rho and Rho GNEF, the down-stream molecules of DC-SIGN mediated signal pathway, were also greatly upregulated. Further, the level of TLR9, the traditional DNA receptor, was significantly downregulated. These results suggest that DC-SIGN as the potential receptor for DNA and protein might trigger the negative pathway to contribute the induction of DCreg combining with Cav-1 mediated negative signal pathway.

Getting the Message? Native Reactive Electrophiles Pass Two Out of Three Thresholds to be Bona Fide Signaling Mediators.

Science.gov (United States)

Poganik, Jesse R; Long, Marcus J C; Aye, Yimon

2018-05-01

Precision cell signaling activities of reactive electrophilic species (RES) are arguably among the most poorly-understood means to transmit biological messages. Latest research implicates native RES to be a chemically-distinct subset of endogenous redox signals that influence cell decision making through non-enzyme-assisted modifications of specific proteins. Yet, fundamental questions remain regarding the role of RES as bona fide second messengers. Here, we lay out three sets of criteria we feel need to be met for RES to be considered as true cellular signals that directly mediate information transfer by modifying "first-responding" sensor proteins. We critically assess the available evidence and define the extent to which each criterion has been fulfilled. Finally, we offer some ideas on the future trajectories of the electrophile signaling field taking inspiration from work that has been done to understand canonical signaling mediators. Also see the video abstract here: https://youtu.be/rG7o0clVP0c. © 2018 WILEY Periodicals, Inc.

Proteomic analysis of the signaling pathway mediated by the heterotrimeric G? protein Pga1 of Penicillium chrysogenum

OpenAIRE

Carrasco-Navarro, Ulises; Vera-Estrella, Rosario; Barkla, Bronwyn J.; Z??iga-Le?n, Eduardo; Reyes-Vivas, Horacio; Fern?ndez, Francisco J.; Fierro, Francisco

2016-01-01

Background The heterotrimeric G? protein Pga1-mediated signaling pathway regulates the entire developmental program in Penicillium chrysogenum, from spore germination to the formation of conidia. In addition it participates in the regulation of penicillin biosynthesis. We aimed to advance the understanding of this key signaling pathway using a proteomics approach, a powerful tool to identify effectors participating in signal transduction pathways. Results Penicillium chrysogenum mutants with ...

Amyloid β production is regulated by β2-adrenergic signaling-mediated post-translational modifications of the ryanodine receptor.

Science.gov (United States)

Bussiere, Renaud; Lacampagne, Alain; Reiken, Steven; Liu, Xiaoping; Scheuerman, Valerie; Zalk, Ran; Martin, Cécile; Checler, Frederic; Marks, Andrew R; Chami, Mounia

2017-06-16

Alteration of ryanodine receptor (RyR)-mediated calcium (Ca 2+ ) signaling has been reported in Alzheimer disease (AD) models. However, the molecular mechanisms underlying altered RyR-mediated intracellular Ca 2+ release in AD remain to be fully elucidated. We report here that RyR2 undergoes post-translational modifications (phosphorylation, oxidation, and nitrosylation) in SH-SY5Y neuroblastoma cells expressing the β-amyloid precursor protein (βAPP) harboring the familial double Swedish mutations (APPswe). RyR2 macromolecular complex remodeling, characterized by depletion of the regulatory protein calstabin2, resulted in increased cytosolic Ca 2+ levels and mitochondrial oxidative stress. We also report a functional interplay between amyloid β (Aβ), β-adrenergic signaling, and altered Ca 2+ signaling via leaky RyR2 channels. Thus, post-translational modifications of RyR occur downstream of Aβ through a β2-adrenergic signaling cascade that activates PKA. RyR2 remodeling in turn enhances βAPP processing. Importantly, pharmacological stabilization of the binding of calstabin2 to RyR2 channels, which prevents Ca 2+ leakage, or blocking the β2-adrenergic signaling cascade reduced βAPP processing and the production of Aβ in APPswe-expressing SH-SY5Y cells. We conclude that targeting RyR-mediated Ca 2+ leakage may be a therapeutic approach to treat AD. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Phospho-Tyrosine(s) vs. Phosphatidylinositol Binding in Shc Mediated Integrin Signaling

OpenAIRE

Lin, Xiaochen; Vinogradova, Olga

2015-01-01

The Shc adaptor protein, particularly its p52 isoform, has been identified as a primary signaling partner for the tyrosine(s)-phosphorylated cytoplasmic tails of activated ? 3 integrins. Inspired by our recent structure of the Shc PTB domain in complex with a bi-phosphorylated peptide derived from ? 3 cytoplasmic tail, we have initiated the investigation of Shc interaction with phospholipids of the membrane. We are particularly focused on PtdIns and their effects on Shc mediated integrin sign...

FGFR3 gene mutation plus GRB10 gene duplication in a patient with achondroplasia plus growth delay with prenatal onset.

Science.gov (United States)

Yuan, Haiming; Huang, Linhuan; Hu, Xizi; Li, Qian; Sun, Xiaofang; Xie, Yingjun; Kong, Shu; Wang, Xiaoman

2016-07-02

Achondroplasia is a well-defined and common bone dysplasia. Genotype- and phenotype-level correlations have been found between the clinical symptoms of achondroplasia and achondroplasia-specific FGFR3 mutations. A 2-year-old boy with clinical features consistent with achondroplasia and Silver-Russell syndrome-like symptoms was found to carry a mutation in the fibroblast growth factor receptor-3 (FGFR3) gene at c.1138G > A (p.Gly380Arg) and a de novo 574 kb duplication at chromosome 7p12.1 that involved the entire growth-factor receptor bound protein 10 (GRB10) gene. Using quantitative real-time PCR analysis, GRB10 was over-expressed, and, using enzyme-linked immunosorbent assays for IGF1 and IGF-binding protein-3 (IGFBP3), we found that IGF1 and IGFBP3 were low-expressed in this patient. We demonstrate that a combination of uncommon, rare and exceptional molecular defects related to the molecular bases of particular birth defects can be analyzed and diagnosed to potentially explain the observed variability in the combination of molecular defects.

Unfolded protein response (UPR) signaling regulates arsenic trioxide-mediated macrophage innate immune function disruption

International Nuclear Information System (INIS)

Srivastava, Ritesh K.; Li, Changzhao; Chaudhary, Sandeep C.; Ballestas, Mary E.; Elmets, Craig A.; Robbins, David J.; Matalon, Sadis; Deshane, Jessy S.; Afaq, Farrukh; Bickers, David R.; Athar, Mohammad

2013-01-01

Arsenic exposure is known to disrupt innate immune functions in humans and in experimental animals. In this study, we provide a mechanism by which arsenic trioxide (ATO) disrupts macrophage functions. ATO treatment of murine macrophage cells diminished internalization of FITC-labeled latex beads, impaired clearance of phagocytosed fluorescent bacteria and reduced secretion of pro-inflammatory cytokines. These impairments in macrophage functions are associated with ATO-induced unfolded protein response (UPR) signaling pathway characterized by the enhancement in proteins such as GRP78, p-PERK, p-eIF2α, ATF4 and CHOP. The expression of these proteins is altered both at transcriptional and translational levels. Pretreatment with chemical chaperon, 4-phenylbutyric acid (PBA) attenuated the ATO-induced activation in UPR signaling and afforded protection against ATO-induced disruption of macrophage functions. This treatment also reduced ATO-mediated reactive oxygen species (ROS) generation. Interestingly, treatment with antioxidant N-acetylcysteine (NAC) prior to ATO exposure, not only reduced ROS production and UPR signaling but also improved macrophage functions. These data demonstrate that UPR signaling and ROS generation are interdependent and are involved in the arsenic-induced pathobiology of macrophage. These data also provide a novel strategy to block the ATO-dependent impairment in innate immune responses. - Highlights: • Inorganic arsenic to humans and experimental animals disrupt innate immune responses. • The mechanism underlying arsenic impaired macrophage functions involves UPR signaling. • Chemical chaperon attenuates arsenic-mediated macrophage function impairment. • Antioxidant, NAC blocks impairment in arsenic-treated macrophage functions

Unfolded protein response (UPR) signaling regulates arsenic trioxide-mediated macrophage innate immune function disruption

Energy Technology Data Exchange (ETDEWEB)

Srivastava, Ritesh K.; Li, Changzhao; Chaudhary, Sandeep C. [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Ballestas, Mary E. [Department of Pediatrics Infectious Disease, Children' s of Alabama, School of Medicine, University of Alabama at Birmingham, AL (United States); Elmets, Craig A. [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Robbins, David J. [Department of Surgery, Molecular Oncology Program, Miller School of Medicine, University of Miami, Miami (United States); Matalon, Sadis [Department of Anesthesiology, University of Alabama at Birmingham, Birmingham, AL (United States); Deshane, Jessy S. [Department of Medicine, Division of Pulmonary, Allergy and Critical Care Medicine, University of Alabama at Birmingham, Birmingham, AL (United States); Afaq, Farrukh [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Bickers, David R. [Department of Dermatology, Columbia University Medical Center, New York (United States); Athar, Mohammad, E-mail: mathar@uab.edu [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States)

2013-11-01

Arsenic exposure is known to disrupt innate immune functions in humans and in experimental animals. In this study, we provide a mechanism by which arsenic trioxide (ATO) disrupts macrophage functions. ATO treatment of murine macrophage cells diminished internalization of FITC-labeled latex beads, impaired clearance of phagocytosed fluorescent bacteria and reduced secretion of pro-inflammatory cytokines. These impairments in macrophage functions are associated with ATO-induced unfolded protein response (UPR) signaling pathway characterized by the enhancement in proteins such as GRP78, p-PERK, p-eIF2α, ATF4 and CHOP. The expression of these proteins is altered both at transcriptional and translational levels. Pretreatment with chemical chaperon, 4-phenylbutyric acid (PBA) attenuated the ATO-induced activation in UPR signaling and afforded protection against ATO-induced disruption of macrophage functions. This treatment also reduced ATO-mediated reactive oxygen species (ROS) generation. Interestingly, treatment with antioxidant N-acetylcysteine (NAC) prior to ATO exposure, not only reduced ROS production and UPR signaling but also improved macrophage functions. These data demonstrate that UPR signaling and ROS generation are interdependent and are involved in the arsenic-induced pathobiology of macrophage. These data also provide a novel strategy to block the ATO-dependent impairment in innate immune responses. - Highlights: • Inorganic arsenic to humans and experimental animals disrupt innate immune responses. • The mechanism underlying arsenic impaired macrophage functions involves UPR signaling. • Chemical chaperon attenuates arsenic-mediated macrophage function impairment. • Antioxidant, NAC blocks impairment in arsenic-treated macrophage functions.

Cre-mediated stress affects sirtuin expression levels, peroxisome biogenesis and metabolism, antioxidant and proinflammatory signaling pathways.

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Yu Xiao

Full Text Available Cre-mediated excision of loxP sites is widely used in mice to manipulate gene function in a tissue-specific manner. To analyze phenotypic alterations related to Cre-expression, we have used AMH-Cre-transgenic mice as a model system. Different Cre expression levels were obtained by investigation of C57BL/6J wild type as well as heterozygous and homozygous AMH-Cre-mice. Our results indicate that Cre-expression itself in Sertoli cells already has led to oxidative stress and lipid peroxidation (4-HNE lysine adducts, inducing PPARα/γ, peroxisome proliferation and alterations of peroxisome biogenesis (PEX5, PEX13 and PEX14 as well as metabolic proteins (ABCD1, ABCD3, MFP1, thiolase B, catalase. In addition to the strong catalase increase, a NRF2- and FOXO3-mediated antioxidative response (HMOX1 of the endoplasmic reticulum and mitochondrial SOD2 and a NF-κB activation were noted. TGFβ1 and proinflammatory cytokines like IL1, IL6 and TNFα were upregulated and stress-related signaling pathways were induced. Sertoli cell mRNA-microarray analysis revealed an increase of TNFR2-signaling components. 53BP1 recruitment and expression levels for DNA repair genes as well as for p53 were elevated and the ones for related sirtuin deacetylases affected (SIRT 1, 3-7 in Sertoli cells. Under chronic Cre-mediated DNA damage conditions a strong downregulation of Sirt1 was observed, suggesting that the decrease of this important coordinator between DNA repair and metabolic signaling might induce the repression release of major transcription factors regulating metabolic and cytokine-mediated stress pathways. Indeed, caspase-3 was activated and increased germ cell apoptosis was observed, suggesting paracrine effects. In conclusion, the observed wide stress-induced effects and metabolic alterations suggest that it is essential to use the correct control animals (Cre/Wt with matched Cre expression levels to differentiate between Cre-mediated and specific gene-knock out-mediated

Cre-Mediated Stress Affects Sirtuin Expression Levels, Peroxisome Biogenesis and Metabolism, Antioxidant and Proinflammatory Signaling Pathways

Science.gov (United States)

Xiao, Yu; Karnati, Srikanth; Qian, Guofeng; Nenicu, Anca; Fan, Wei; Tchatalbachev, Svetlin; Höland, Anita; Hossain, Hamid; Guillou, Florian; Lüers, Georg H.; Baumgart-Vogt, Eveline

2012-01-01

Cre-mediated excision of loxP sites is widely used in mice to manipulate gene function in a tissue-specific manner. To analyze phenotypic alterations related to Cre-expression, we have used AMH-Cre-transgenic mice as a model system. Different Cre expression levels were obtained by investigation of C57BL/6J wild type as well as heterozygous and homozygous AMH-Cre-mice. Our results indicate that Cre-expression itself in Sertoli cells already has led to oxidative stress and lipid peroxidation (4-HNE lysine adducts), inducing PPARα/γ, peroxisome proliferation and alterations of peroxisome biogenesis (PEX5, PEX13 and PEX14) as well as metabolic proteins (ABCD1, ABCD3, MFP1, thiolase B, catalase). In addition to the strong catalase increase, a NRF2- and FOXO3-mediated antioxidative response (HMOX1 of the endoplasmic reticulum and mitochondrial SOD2) and a NF-κB activation were noted. TGFβ1 and proinflammatory cytokines like IL1, IL6 and TNFα were upregulated and stress-related signaling pathways were induced. Sertoli cell mRNA-microarray analysis revealed an increase of TNFR2-signaling components. 53BP1 recruitment and expression levels for DNA repair genes as well as for p53 were elevated and the ones for related sirtuin deacetylases affected (SIRT 1, 3-7) in Sertoli cells. Under chronic Cre-mediated DNA damage conditions a strong downregulation of Sirt1 was observed, suggesting that the decrease of this important coordinator between DNA repair and metabolic signaling might induce the repression release of major transcription factors regulating metabolic and cytokine-mediated stress pathways. Indeed, caspase-3 was activated and increased germ cell apoptosis was observed, suggesting paracrine effects. In conclusion, the observed wide stress-induced effects and metabolic alterations suggest that it is essential to use the correct control animals (Cre/Wt) with matched Cre expression levels to differentiate between Cre-mediated and specific gene-knock out-mediated

Convergent RANK- and c-Met-mediated signaling components predict survival of patients with prostate cancer: an interracial comparative study.

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Peizhen Hu

Full Text Available We reported (PLoS One 6 (12:e28670, 2011 that the activation of c-Met signaling in RANKL-overexpressing bone metastatic LNCaP cell and xenograft models increased expression of RANK, RANKL, c-Met, and phosphorylated c-Met, and mediated downstream signaling. We confirmed the significance of the RANK-mediated signaling network in castration resistant clinical human prostate cancer (PC tissues. In this report, we used a multispectral quantum dot labeling technique to label six RANK and c-Met convergent signaling pathway mediators simultaneously in formalin fixed paraffin embedded (FFPE tissue specimens, quantify the intensity of each expression at the sub-cellular level, and investigated their potential utility as predictors of patient survival in Caucasian-American, African-American and Chinese men. We found that RANKL and neuropilin-1 (NRP-1 expression predicts survival of Caucasian-Americans with PC. A Gleason score ≥ 8 combined with nuclear p-c-Met expression predicts survival in African-American PC patients. Neuropilin-1, p-NF-κB p65 and VEGF are predictors for the overall survival of Chinese men with PC. These results collectively support interracial differences in cell signaling networks that can predict the survival of PC patients.

Downregulation of toll-like receptor-mediated signalling pathways in oral lichen planus.

Science.gov (United States)

Sinon, Suraya H; Rich, Alison M; Parachuru, Venkata P B; Firth, Fiona A; Milne, Trudy; Seymour, Gregory J

2016-01-01

The objective of this study was to investigate the expression of Toll-like receptors (TLR) and TLR-associated signalling pathway genes in oral lichen planus (OLP). Initially, immunohistochemistry was used to determine TLR expression in 12 formalin-fixed archival OLP tissues with 12 non-specifically inflamed oral tissues as controls. RNA was isolated from further fresh samples of OLP and non-specifically inflamed oral tissue controls (n = 6 for both groups) and used in qRT(2)-PCR focused arrays to determine the expression of TLRs and associated signalling pathway genes. Genes with a statistical significance of ±two-fold regulation (FR) and a P-value < 0.05 were considered as significantly regulated. Significantly more TLR4(+) cells were present in the inflammatory infiltrate in OLP compared with the control tissues (P < 0.05). There was no statistically significant difference in the numbers of TLR2(+) and TLR8(+) cells between the groups. TLR3 was significantly downregulated in OLP (P < 0.01). TLR8 was upregulated in OLP, but the difference between the groups was not statistically significant. The TLR-mediated signalling-associated protein genes MyD88 and TIRAP were significantly downregulated (P < 0.01 and P < 0.05), as were IRAK1 (P < 0.05), MAPK8 (P < 0.01), MAP3K1 (P < 0.05), MAP4K4 (P < 0.05), REL (P < 0.01) and RELA (P < 0.01). Stress proteins HMGB1 and the heat shock protein D1 were significantly downregulated in OLP (P < 0.01). These findings suggest a downregulation of TLR-mediated signalling pathways in OLP lesions. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Uncovering molecular structural mechanisms of signaling mediated by the prion protein

International Nuclear Information System (INIS)

Romano, Sebastian A.; Linden, Rafael; Silva, Jerson L.; Foguel, Debora

2009-01-01

The glycosyl phosphatidylinositol (GPI) - anchored prion protein (PrP c ), usually associated with neurodegenerative diseases, modulates various cellular responses and may scaffold multiprotein cell surface signaling complexes. Engagement of PrP c with the secretable cochaperone hop/STI 1 induces neurotrophic transmembrane signals through unknown molecular mechanisms. We addressed whether interaction of Pr P c and hop STI 1 entails structural rearrangements relevant for signaling. Circular dichroism and fluorescence spectroscopy showed that PrP c :hop/STI 1 interaction triggers loss of PrP helical structures, involving at least a perturbation of the Pr P c 143-153 beta-helix. Novel SAXS models revealed a significant C-terminal compaction of hop/STI 1 when bound to PrP c . Differing from a recent dimeric model of human hop/STI 1, both size exclusion chromatography and SAXS data support a monomeric form of free murine hop/STI 1. Changes in the Pr P c 143-153 beta-helix may engage the transmembrane signaling protein laminin receptor precursor and neural cell adhesion molecule, both of which bind that domain of Pr P c , and further ligands may be engaged by the tertiary structural changes of hop/STI 1. These reciprocal structural modifications indicate a versatile mechanism for signaling mediated by Pr P c :hop/STI 1 interaction, consistent with the hypothesis that Pr P c scaffolds multiprotein signaling complexes at the cell surface. (author)

Uncovering molecular structural mechanisms of signaling mediated by the prion protein

Energy Technology Data Exchange (ETDEWEB)

Romano, Sebastian A.; Linden, Rafael [Universidade Federal do Rio de Janeiro (IBCCF/UFRl), RJ (Brazil). Inst. de Biofisica Carlos Chagas Filho; Cordeiro, Yraima; Rocha e Lima, Luis M.T. da [Universidade Federal do Rio de Janeiro (FF/UFRl), RJ (Brazil). Fac. de Farmacia; Lopes, Marilene H. [Instituto Ludwig de Pesquisa de Cancer, Sao Paulo, SP (Brazil); Silva, Jerson L.; Foguel, Debora [Universidade Federal do Rio de Janeiro (IBqM/UFRl), RJ (Brazil). Inst. de Bioquimica Medica

2009-07-01

The glycosyl phosphatidylinositol (GPI) - anchored prion protein (PrP{sup c}), usually associated with neurodegenerative diseases, modulates various cellular responses and may scaffold multiprotein cell surface signaling complexes. Engagement of PrP{sup c} with the secretable cochaperone hop/STI 1 induces neurotrophic transmembrane signals through unknown molecular mechanisms. We addressed whether interaction of Pr P{sup c} and hop STI 1 entails structural rearrangements relevant for signaling. Circular dichroism and fluorescence spectroscopy showed that PrP{sup c}:hop/STI 1 interaction triggers loss of PrP helical structures, involving at least a perturbation of the Pr P{sup c}{sub 143-153} beta-helix. Novel SAXS models revealed a significant C-terminal compaction of hop/STI 1 when bound to PrP{sup c}. Differing from a recent dimeric model of human hop/STI 1, both size exclusion chromatography and SAXS data support a monomeric form of free murine hop/STI 1. Changes in the Pr P{sup c}{sub 143-153} beta-helix may engage the transmembrane signaling protein laminin receptor precursor and neural cell adhesion molecule, both of which bind that domain of Pr P{sup c}, and further ligands may be engaged by the tertiary structural changes of hop/STI 1. These reciprocal structural modifications indicate a versatile mechanism for signaling mediated by Pr P{sup c}:hop/STI 1 interaction, consistent with the hypothesis that Pr P{sup c} scaffolds multiprotein signaling complexes at the cell surface. (author)

Signals in hepatitis A virus P3 region proteins recognized by the ubiquitin-mediated proteolytic system

International Nuclear Information System (INIS)

Losick, Vicki P.; Schlax, Peter E.; Emmons, Rebecca A.; Lawson, T. Glen

2003-01-01

The hepatitis A virus 3C protease and 3D RNA polymerase are present in low concentrations in infected cells. The 3C protease was previously shown to be rapidly degraded by the ubiquitin/26S proteasome system and we present evidence here that the 3D polymerase is also subject to ubiquitination-mediated proteolysis. Our results show that the sequence 32 LGVKDDWLLV 41 in the 3C protease serves as a protein destruction signal recognized by the ubiquitin-protein ligase E3α and that the destruction signal for the RNA polymerase does not require the carboxyl-terminal 137 amino acids. Both the viral 3ABCD polyprotein and the 3CD diprotein were also found to be substrates for ubiquitin-mediated proteolysis. Attempts to determine if the 3C protease or the 3D polymerase destruction signals trigger the ubiquitination and degradation of these precursors yielded evidence suggesting, but not unequivocally proving, that the recognition of the 3D polymerase by the ubiquitin system is responsible

Progesterone receptors (PR) mediate STAT actions: PR and prolactin receptor signaling crosstalk in breast cancer models.

Science.gov (United States)

Leehy, Katherine A; Truong, Thu H; Mauro, Laura J; Lange, Carol A

2018-02-01

Estrogen is the major mitogenic stimulus of mammary gland development during puberty wherein ER signaling acts to induce abundant PR expression. PR signaling, in contrast, is the primary driver of mammary epithelial cell proliferation in adulthood. The high circulating levels of progesterone during pregnancy signal through PR, inducing expression of the prolactin receptor (PRLR). Cooperation between PR and prolactin (PRL) signaling, via regulation of downstream components in the PRL signaling pathway including JAKs and STATs, facilitates the alveolar morphogenesis observed during pregnancy. Indeed, these pathways are fully integrated via activation of shared signaling pathways (i.e. JAKs, MAPKs) as well as by the convergence of PRs and STATs at target genes relevant to both mammary gland biology and breast cancer progression (i.e. proliferation, stem cell outgrowth, tissue cell type heterogeneity). Thus, rather than a single mediator such as ER, transcription factor cascades (ER>PR>STATs) are responsible for rapid proliferative and developmental programming in the normal mammary gland. It is not surprising that these same mediators typify uncontrolled proliferation in a majority of breast cancers, where ER and PR are most often co-expressed and may cooperate to drive malignant tumor progression. This review will primarily focus on the integration of PR and PRL signaling in breast cancer models and the importance of this cross-talk in cancer progression in the context of mammographic density. Components of these PR/PRL signaling pathways could offer alternative drug targets and logical complements to anti-ER or anti-estrogen-based endocrine therapies. Copyright © 2017 Elsevier Ltd. All rights reserved.

SPRY4-mediated ERK1/2 signaling inhibition abolishes 17β-estradiol-induced cell growth in endometrial adenocarcinoma cell.

Science.gov (United States)

Li, Mingjiang; Zhang, Hui; Zhao, Xingbo; Yan, Lei; Wang, Chong; Li, Chunyan; Li, Changzhong

2014-08-01

Basic fibroblast growth factor (FGF2)-mediated Extracellular signal-regulated kinases1/2 (ERK1/2) signaling is a critical modulator in angiogenesis. SPRY4 has been reported to be a feedback negative regulator of FGFs-induced ERK1/2 signaling. The aim of this study was to explore the role of SPRY4 in endometrial adenocarcinoma cell. The effect of SPRY4 expression on FGF2-mediated ERK1/2 signaling was detected by luciferase assay and Western blot analysis. The growth of Ishikawa cells was detected using colony formation assay and cell number counting experiment. We found that plasmid-driven SPRY4 expression efficiently blocked the activity of FGF2-induced ERK1/2 signaling in Ishikawa cells. SPRY4 expression significantly reduced the proliferation and 17β-estradiol-induced proliferation of Ishikawa cells. SPRY4 may function as a tumor suppressor in endometrial adenocarcinoma.

Transforming growth factor β-induced expression of chondroitin sulfate proteoglycans is mediated through non-Smad signaling pathways.

Science.gov (United States)

Jahan, Naima; Hannila, Sari S

2015-01-01

The expression of chondroitin sulfate proteoglycans (CSPGs) by reactive astrocytes is a major factor contributing to glial scarring and regenerative failure after spinal cord injury, but the molecular mechanisms underlying CSPG expression remain largely undefined. One contributing factor is transforming growth factor β (TGFβ), which is upregulated after injury and has been shown to induce expression of CSPGs in vitro. TGFβ typically mediates its effects through the Smad2/3 signaling pathway, and it has been suggested that this pathway is responsible for CSPG expression. However, there is evidence that TGFβ can also activate non-Smad signaling pathways. In this study, we report that TGFβ-induced expression of three different CSPGs--neurocan, brevican, and aggrecan--is mediated through non-Smad signaling pathways. We observed significant increases in TGFβ-induced expression of neurocan, brevican, and aggrecan following siRNA knockdown of Smad2 or Smad4, which indicates that Smad signaling is not required for the expression of these CSPGs. In addition, we show that neurocan, aggrecan, and brevican levels are significantly reduced when TGFβ is administered in the presence of either the PI3K inhibitor LY294002 or the mTOR inhibitor rapamycin, but not the MEK1/2 inhibitor U0126. This suggests that TGFβ mediates this effect through non-Smad-dependent activation of the PI3K-Akt-mTOR signaling pathway, and targeting this pathway may therefore be an effective means of reducing CSPG expression in the injured CNS. Copyright © 2014 Elsevier Inc. All rights reserved.

SPATA2-Mediated Binding of CYLD to HOIP Enables CYLD Recruitment to Signaling Complexes

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Sebastian Kupka

2016-08-01

Full Text Available Recruitment of the deubiquitinase CYLD to signaling complexes is mediated by its interaction with HOIP, the catalytically active component of the linear ubiquitin chain assembly complex (LUBAC. Here, we identify SPATA2 as a constitutive direct binding partner of HOIP that bridges the interaction between CYLD and HOIP. SPATA2 recruitment to TNFR1- and NOD2-signaling complexes is dependent on HOIP, and loss of SPATA2 abolishes CYLD recruitment. Deficiency in SPATA2 exerts limited effects on gene activation pathways but diminishes necroptosis induced by tumor necrosis factor (TNF, resembling loss of CYLD. In summary, we describe SPATA2 as a previously unrecognized factor in LUBAC-dependent signaling pathways that serves as an adaptor between HOIP and CYLD, thereby enabling recruitment of CYLD to signaling complexes.

Glucose intolerance induced by blockade of central FGF receptors is linked to an acute stress response

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Jennifer M. Rojas

2015-08-01

Conclusions: The effect of acute inhibition of central FGFR signaling to impair glucose tolerance likely involves a stress response associated with pronounced, but transient, sympathoadrenal activation and an associated reduction of insulin secretion. Whether this effect is a true consequence of FGFR blockade or involves an off-target effect of the FGFR inhibitor requires additional study.

Extracellular signal regulated kinase 5 mediates signals triggered by the novel tumor promoter palytoxin

International Nuclear Information System (INIS)

Charlson, Aaron T.; Zeliadt, Nicholette A.; Wattenberg, Elizabeth V.

2009-01-01

Palytoxin is classified as a non-12-O-tetradecanoylphorbol-13-acetate (TPA)-type skin tumor because it does not bind to or activate protein kinase C. Palytoxin is thus a novel tool for investigating alternative signaling pathways that may affect carcinogenesis. We previously showed that palytoxin activates three major members of the mitogen activated protein kinase (MAPK) family, extracellular signal regulated kinase 1 and 2 (ERK1/2), c-Jun N-terminal kinase (JNK), and p38. Here we report that palytoxin also activates another MAPK family member, called ERK5, in HeLa cells and in keratinocytes derived from initiated mouse skin (308 cells). By contrast, TPA does not activate ERK5 in these cell lines. The major cell surface receptor for palytoxin is the Na+,K+-ATPase. Accordingly, ouabain blocked the ability of palytoxin to activate ERK5. Ouabain alone did not activate ERK5. ERK5 thus represents a divergence in the signaling pathways activated by these two agents that bind to the Na+,K+-ATPase. Cycloheximide, okadaic acid, and sodium orthovanadate did not mimic the effect of palytoxin on ERK5. These results indicate that the stimulation of ERK5 by palytoxin is not simply due to inhibition of protein synthesis or inhibition of serine/threonine or tyrosine phosphatases. Therefore, the mechanism by which palytoxin activates ERK5 differs from that by which it activates ERK1/2, JNK, and p38. Finally, studies that used pharmacological inhibitors and shRNA to block ERK5 action indicate that ERK5 contributes to palytoxin-stimulated c-Fos gene expression. These results suggest that ERK5 can act as an alternative mediator for transmitting diverse tumor promoter-stimulated signals.

Antioxidant role of glutathione S-transferases: 4-Hydroxynonenal, a key molecule in stress-mediated signaling.

Science.gov (United States)

Singhal, Sharad S; Singh, Sharda P; Singhal, Preeti; Horne, David; Singhal, Jyotsana; Awasthi, Sanjay

2015-12-15

4-Hydroxy-2-trans-nonenal (4HNE), one of the major end products of lipid peroxidation (LPO), has been shown to induce apoptosis in a variety of cell lines. It appears to modulate signaling processes in more than one way because it has been suggested to have a role in signaling for differentiation and proliferation. It has been known that glutathione S-transferases (GSTs) can reduce lipid hydroperoxides through their Se-independent glutathione-peroxidase activity and that these enzymes can also detoxify LPO end-products such as 4HNE. Available evidence from earlier studies together with results of recent studies in our laboratories strongly suggests that LPO products, particularly hydroperoxides and 4HNE, are involved in the mechanisms of stress-mediated signaling and that it can be modulated by the alpha-class GSTs through the regulation of the intracellular concentrations of 4HNE. We demonstrate that 4HNE induced apoptosis in various cell lines is accompanied with c-Jun-N-terminal kinase (JNK) and caspase-3 activation. Cells exposed to mild, transient heat or oxidative stress acquire the capacity to exclude intracellular 4HNE at a faster rate by inducing GSTA4-4 which conjugates 4HNE to glutathione (GSH), and RLIP76 which mediates the ATP-dependent transport of the GSH-conjugate of 4HNE (GS-HNE). The balance between formation and exclusion promotes different cellular processes - higher concentrations of 4HNE promote apoptosis; whereas, lower concentrations promote proliferation. In this article, we provide a brief summary of the cellular effects of 4HNE, followed by a review of its GST-catalyzed detoxification, with an emphasis on the structural attributes that play an important role in the interactions with alpha-class GSTA4-4. Taken together, 4HNE is a key signaling molecule and that GSTs being determinants of its intracellular concentrations, can regulate stress-mediated signaling, are reviewed in this article. Copyright © 2015 Elsevier Inc. All rights

BMP signaling mediates effects of exercise on hippocampal neurogenesis and cognition in mice.

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Kevin T Gobeske

2009-10-01

Full Text Available Exposure to exercise or to environmental enrichment increases the generation of new neurons in the adult hippocampus and promotes certain kinds of learning and memory. While the precise role of neurogenesis in cognition has been debated intensely, comparatively few studies have addressed the mechanisms linking environmental exposures to cellular and behavioral outcomes. Here we show that bone morphogenetic protein (BMP signaling mediates the effects of exercise on neurogenesis and cognition in the adult hippocampus. Elective exercise reduces levels of hippocampal BMP signaling before and during its promotion of neurogenesis and learning. Transgenic mice with decreased BMP signaling or wild type mice infused with a BMP inhibitor both exhibit remarkable gains in hippocampal cognitive performance and neurogenesis, mirroring the effects of exercise. Conversely, transgenic mice with increased BMP signaling have diminished hippocampal neurogenesis and impaired cognition. Exercise exposure does not rescue these deficits, suggesting that reduced BMP signaling is required for environmental effects on neurogenesis and learning. Together, these observations show that BMP signaling is a fundamental mechanism linking environmental exposure with changes in cognitive function and cellular properties in the hippocampus.

The Signaling Pathways Involved in Chondrocyte Differentiation and Hypertrophic Differentiation

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Jianmei Li

2016-01-01

Full Text Available Chondrocytes communicate with each other mainly via diffusible signals rather than direct cell-to-cell contact. The chondrogenic differentiation of mesenchymal stem cells (MSCs is well regulated by the interactions of varieties of growth factors, cytokines, and signaling molecules. A number of critical signaling molecules have been identified to regulate the differentiation of chondrocyte from mesenchymal progenitor cells to their terminal maturation of hypertrophic chondrocytes, including bone morphogenetic proteins (BMPs, SRY-related high-mobility group-box gene 9 (Sox9, parathyroid hormone-related peptide (PTHrP, Indian hedgehog (Ihh, fibroblast growth factor receptor 3 (FGFR3, and β-catenin. Except for these molecules, other factors such as adenosine, O2 tension, and reactive oxygen species (ROS also have a vital role in cartilage formation and chondrocyte maturation. Here, we outlined the complex transcriptional network and the function of key factors in this network that determine and regulate the genetic program of chondrogenesis and chondrocyte differentiation.

Prenatal and postnatal presentation of severe achondroplasia with developmental delay and acanthosis nigricans (SADDAN) due to the FGFR3 Lys650Met mutation.

NARCIS (Netherlands)

Zankl, A.; Elakis, G.; Susman, R.D.; Inglis, G.; Gardener, G.; Buckley, M.F.; Roscioli, T.

2008-01-01

We present prenatal and postnatal features of a patient with severe achondroplasia with developmental delay and acanthosis nigricans (SADDAN). Mutation analysis confirmed the clinical diagnosis by detecting the FGFR3 Lys650Met mutation. This case, one of only six with molecular analysis reported in

Phospho-Tyrosine(s) vs. Phosphatidylinositol Binding in Shc Mediated Integrin Signaling.

Science.gov (United States)

Lin, Xiaochen; Vinogradova, Olga

2015-04-01

The Shc adaptor protein, particularly its p52 isoform, has been identified as a primary signaling partner for the tyrosine(s)-phosphorylated cytoplasmic tails of activated β 3 integrins. Inspired by our recent structure of the Shc PTB domain in complex with a bi-phosphorylated peptide derived from β 3 cytoplasmic tail, we have initiated the investigation of Shc interaction with phospholipids of the membrane. We are particularly focused on PtdIns and their effects on Shc mediated integrin signaling in vitro . Here we present thermodynamic profiles and molecular details of the interactions between Shc, integrin, and PtdIns, all of which have been studied by ITC and solution NMR methods. A model of p52 Shc interaction with phosphorylated β 3 integrin cytoplasmic tail at the cytosolic face of the plasma membrane is proposed based on these data.

A pivotal role of the jasmonic acid signal pathway in mediating radiation-induced bystander effects in Arabidopsis thaliana.

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Wang, Ting; Xu, Wei; Deng, Chenguang; Xu, Shaoxin; Li, Fanghua; Wu, Yuejin; Wu, Lijun; Bian, Po

Although radiation-induced bystander effects (RIBE) in Arabidopsis thaliana have been well demonstrated in vivo, little is known about their underlying mechanisms, particularly with regard to the participating signaling molecules and signaling pathways. In higher plants, jasmonic acid (JA) and its bioactive derivatives are well accepted as systemic signal transducers that are produced in response to various environmental stresses. It is therefore speculated that the JA signal pathway might play a potential role in mediating radiation-induced bystander signaling of root-to-shoot. In the present study, pretreatment of seedlings with Salicylhydroxamic acid, an inhibitor of lipoxigenase (LOX) in JA biosynthesis, significantly suppressed RIBE-mediated expression of the AtRAD54 gene. After root irradiation, the aerial parts of A. thaliana mutants deficient in JA biosynthesis (aos) and signaling cascades (jar1-1) showed suppressed induction of the AtRAD54 and AtRAD51 genes and TSI and 180-bp repeats, which have been extensively used as endpoints of bystander genetic and epigenetic effects in plants. These results suggest an involvement of the JA signal pathway in the RIBE of plants. Using the root micro-grafting technique, the JA signal pathway was shown to participate in both the generation of bystander signals in irradiated root cells and radiation responses in the bystander aerial parts of plants. The over-accumulation of endogenous JA in mutant fatty acid oxygenation up-regulated 2 (fou2), in which mutation of the Two Pore Channel 1 (TPC1) gene up-regulates expression of the LOX and allene oxide synthase (AOS) genes, inhibited RIBE-mediated expression of the AtRAD54 gene, but up-regulated expression of the AtKU70 and AtLIG4 genes in the non-homologous end joining (NHEJ) pathway. Considering that NHEJ is employed by plants with increased DNA damage, the switch from HR to NHEJ suggests that over-accumulation of endogenous JA might enhance the radiosensitivity of plants

Influence of ER leak on resting cytoplasmic Ca2+ and receptor-mediated Ca2+ signalling in human macrophage.

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Layhadi, Janice A; Fountain, Samuel J

2017-06-03

Mechanisms controlling endoplasmic reticulum (ER) Ca 2+ homeostasis are important regulators of resting cytoplasmic Ca 2+ concentration ([Ca 2+ ] cyto ) and receptor-mediated Ca 2+ signalling. Here we investigate channels responsible for ER Ca 2+ leak in THP-1 macrophage and human primary macrophage. In the absence of extracellular Ca 2+ we employ ionomycin action at the plasma membrane to stimulate ER Ca 2+ leak. Under these conditions ionomycin elevates [Ca 2+ ] cyto revealing a Ca 2+ leak response which is abolished by thapsigargin. IP 3 receptors (Xestospongin C, 2-APB), ryanodine receptors (dantrolene), and translocon (anisomycin) inhibition facilitated ER Ca 2+ leak in model macrophage, with translocon inhibition also reducing resting [Ca 2+ ] cyto . In primary macrophage, translocon inhibition blocks Ca 2+ leak but does not influence resting [Ca 2+ ] cyto . We identify a role for translocon-mediated ER Ca 2+ leak in receptor-mediated Ca 2+ signalling in both model and primary human macrophage, whereby the Ca 2+ response to ADP (P2Y receptor agonist) is augmented following anisomycin treatment. In conclusion, we demonstrate a role of ER Ca 2+ leak via the translocon in controlling resting cytoplasmic Ca 2+ in model macrophage and receptor-mediated Ca 2+ signalling in model macrophage and primary macrophage. Copyright © 2017 Elsevier Inc. All rights reserved.

Importance of leptin signaling and signal transducer and activator of transcription-3 activation in mediating the cardiac hypertrophy associated with obesity.

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Leifheit-Nestler, Maren; Wagner, Nana-Maria; Gogiraju, Rajinikanth; Didié, Michael; Konstantinides, Stavros; Hasenfuss, Gerd; Schäfer, Katrin

2013-07-11

The adipokine leptin and its receptor are expressed in the heart, and leptin has been shown to promote cardiomyocyte hypertrophy in vitro. Obesity is associated with hyperleptinemia and hypothalamic leptin resistance as well as an increased risk to develop cardiac hypertrophy and heart failure. However, the role of cardiac leptin signaling in mediating the cardiomyopathy associated with increased body weight is unclear, in particular, whether it develops subsequently to cardiac leptin resistance or overactivation of hypertrophic signaling pathways via elevated leptin levels. The cardiac phenotype of high-fat diet (HFD)-induced obese wildtype (WT) mice was examined and compared to age-matched genetically obese leptin receptor (LepR)-deficient (LepRdb/db) or lean WT mice. To study the role of leptin-mediated STAT3 activation during obesity-induced cardiac remodeling, mice in which tyrosine residue 1138 within LepR had been replaced with a serine (LepRS1138) were also analyzed. Obesity was associated with hyperleptinemia and elevated cardiac leptin expression in both diet-induced and genetically obese mice. Enhanced LepR and STAT3 phosphorylation levels were detected in hearts of obese WT mice, but not in those with LepR mutations. Moreover, exogenous leptin continued to induce cardiac STAT3 activation in diet-induced obese mice. Although echocardiography revealed signs of cardiac hypertrophy in all obese mice, the increase in left ventricular (LV) mass and diameter was significantly more pronounced in LepRS1138 animals. LepRS1138 mice also exhibited an increased activation of signaling proteins downstream of LepR, including Jak2 (1.8-fold), Src kinase (1.7-fold), protein kinase B (1.3-fold) or C (1.6-fold). Histological analysis of hearts revealed that the inability of leptin to activate STAT3 in LepRdb/db and LepRS1138 mice was associated with reduced cardiac angiogenesis as well as increased apoptosis and fibrosis. Our findings suggest that hearts from obese mice

Canonical TGF-β Signaling Negatively Regulates Neuronal Morphogenesis through TGIF/Smad Complex-Mediated CRMP2 Suppression.

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Nakashima, Hideyuki; Tsujimura, Keita; Irie, Koichiro; Ishizu, Masataka; Pan, Miao; Kameda, Tomonori; Nakashima, Kinichi

2018-05-16

Functional neuronal connectivity requires proper neuronal morphogenesis and its dysregulation causes neurodevelopmental diseases. Transforming growth factor-β (TGF-β) family cytokines play pivotal roles in development, but little is known about their contribution to morphological development of neurons. Here we show that the Smad-dependent canonical signaling of TGF-β family cytokines negatively regulates neuronal morphogenesis during brain development. Mechanistically, activated Smads form a complex with transcriptional repressor TG-interacting factor (TGIF), and downregulate the expression of a neuronal polarity regulator, collapsin response mediator protein 2. We also demonstrate that TGF-β family signaling inhibits neurite elongation of human induced pluripotent stem cell-derived neurons. Furthermore, the expression of TGF-β receptor 1, Smad4, or TGIF, which have mutations found in patients with neurodevelopmental disorders, disrupted neuronal morphogenesis in both mouse (male and female) and human (female) neurons. Together, these findings suggest that the regulation of neuronal morphogenesis by an evolutionarily conserved function of TGF-β signaling is involved in the pathogenesis of neurodevelopmental diseases. SIGNIFICANCE STATEMENT Canonical transforming growth factor-β (TGF-β) signaling plays a crucial role in multiple organ development, including brain, and mutations in components of the signaling pathway associated with several human developmental disorders. In this study, we found that Smads/TG-interacting factor-dependent canonical TGF-β signaling regulates neuronal morphogenesis through the suppression of collapsin response mediator protein-2 (CRMP2) expression during brain development, and that function of this signaling is evolutionarily conserved in the mammalian brain. Mutations in canonical TGF-β signaling factors identified in patients with neurodevelopmental disorders disrupt the morphological development of neurons. Thus, our

The Role of ATP in the Regulation of NCAM Function

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Hübschmann, Martin; Skladchikova, Galina

2008-01-01

overlaps with the site of NCAM-FGFR interaction, and ATP is capable of disrupting NCAM-FGFR binding. This implies that NCAM signaling through FGFR can be regulated by ATP, which is supported by the observation that ATP can abrogate NCAM-induced neurite outgrowth. Finally, ATP can induce NCAM ectodomain...... shedding, possibly affecting the structural plasticity associated with learning and memory....

The impact of cow's milk-mediated mTORC1-signaling in the initiation and progression of prostate cancer

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2012-01-01

Prostate cancer (PCa) is dependent on androgen receptor signaling and aberrations of the PI3K-Akt-mTORC1 pathway mediating excessive and sustained growth signaling. The nutrient-sensitive kinase mTORC1 is upregulated in nearly 100% of advanced human PCas. Oncogenic mTORC1 signaling activates key subsets of mRNAs that cooperate in distinct steps of PCa initiation and progression. Epidemiological evidence points to increased dairy protein consumption as a major dietary risk factor for the development of PCa. mTORC1 is a master regulator of protein synthesis, lipid synthesis and autophagy pathways that couple nutrient sensing to cell growth and cancer. This review provides evidence that PCa initiation and progression are promoted by cow´s milk, but not human milk, stimulation of mTORC1 signaling. Mammalian milk is presented as an endocrine signaling system, which activates mTORC1, promotes cell growth and proliferation and suppresses autophagy. Naturally, milk-mediated mTORC1 signaling is restricted only to the postnatal growth phase of mammals. However, persistent consumption of cow´s milk proteins in humans provide highly insulinotropic branched-chain amino acids (BCAAs) provided by milk´s fast hydrolysable whey proteins, which elevate postprandial plasma insulin levels, and increase hepatic IGF-1 plasma concentrations by casein-derived amino acids. BCAAs, insulin and IGF-1 are pivotal activating signals of mTORC1. Increased cow´s milk protein-mediated mTORC1 signaling along with constant exposure to commercial cow´s milk estrogens derived from pregnant cows may explain the observed association between high dairy consumption and increased risk of PCa in Westernized societies. As well-balanced mTORC1-signaling plays an important role in appropriate prostate morphogenesis and differentiation, exaggerated mTORC1-signaling by high cow´s milk consumption predominantly during critical growth phases of prostate development and differentiation may exert long

The impact of cow's milk-mediated mTORC1-signaling in the initiation and progression of prostate cancer

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Melnik Bodo C

2012-08-01

Full Text Available Abstract Prostate cancer (PCa is dependent on androgen receptor signaling and aberrations of the PI3K-Akt-mTORC1 pathway mediating excessive and sustained growth signaling. The nutrient-sensitive kinase mTORC1 is upregulated in nearly 100% of advanced human PCas. Oncogenic mTORC1 signaling activates key subsets of mRNAs that cooperate in distinct steps of PCa initiation and progression. Epidemiological evidence points to increased dairy protein consumption as a major dietary risk factor for the development of PCa. mTORC1 is a master regulator of protein synthesis, lipid synthesis and autophagy pathways that couple nutrient sensing to cell growth and cancer. This review provides evidence that PCa initiation and progression are promoted by cow´s milk, but not human milk, stimulation of mTORC1 signaling. Mammalian milk is presented as an endocrine signaling system, which activates mTORC1, promotes cell growth and proliferation and suppresses autophagy. Naturally, milk-mediated mTORC1 signaling is restricted only to the postnatal growth phase of mammals. However, persistent consumption of cow´s milk proteins in humans provide highly insulinotropic branched-chain amino acids (BCAAs provided by milk´s fast hydrolysable whey proteins, which elevate postprandial plasma insulin levels, and increase hepatic IGF-1 plasma concentrations by casein-derived amino acids. BCAAs, insulin and IGF-1 are pivotal activating signals of mTORC1. Increased cow´s milk protein-mediated mTORC1 signaling along with constant exposure to commercial cow´s milk estrogens derived from pregnant cows may explain the observed association between high dairy consumption and increased risk of PCa in Westernized societies. As well-balanced mTORC1-signaling plays an important role in appropriate prostate morphogenesis and differentiation, exaggerated mTORC1-signaling by high cow´s milk consumption predominantly during critical growth phases of prostate development and

IFN-γ signaling to astrocytes protects from autoimmune mediated neurological disability.

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Claudia Hindinger

Full Text Available Demyelination and axonal degeneration are determinants of progressive neurological disability in patients with multiple sclerosis (MS. Cells resident within the central nervous system (CNS are active participants in development, progression and subsequent control of autoimmune disease; however, their individual contributions are not well understood. Astrocytes, the most abundant CNS cell type, are highly sensitive to environmental cues and are implicated in both detrimental and protective outcomes during autoimmune demyelination. Experimental autoimmune encephalomyelitis (EAE was induced in transgenic mice expressing signaling defective dominant-negative interferon gamma (IFN-γ receptors on astrocytes to determine the influence of inflammation on astrocyte activity. Inhibition of IFN-γ signaling to astrocytes did not influence disease incidence, onset, initial progression of symptoms, blood brain barrier (BBB integrity or the composition of the acute CNS inflammatory response. Nevertheless, increased demyelination at peak acute disease in the absence of IFN-γ signaling to astrocytes correlated with sustained clinical symptoms. Following peak disease, diminished clinical remission, increased mortality and sustained astrocyte activation within the gray matter demonstrate a critical role of IFN-γ signaling to astrocytes in neuroprotection. Diminished disease remission was associated with escalating demyelination, axonal degeneration and sustained inflammation. The CNS infiltrating leukocyte composition was not altered; however, decreased IL-10 and IL-27 correlated with sustained disease. These data indicate that astrocytes play a critical role in limiting CNS autoimmune disease dependent upon a neuroprotective signaling pathway mediated by engagement of IFN-γ receptors.

Alterations in Lipid Mediated Signaling and Wnt/β-Catenin Signaling in DMH Induced Colon Cancer on Supplementation of Fish Oil

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Shevali Kansal

2014-01-01

Full Text Available Ceramide mediates inhibition of cyclooxygenase-2 (COX-2 which catalyzes formation of prostaglandin further activating peroxisome proliferator-activated receptorγ (PPARγ and Wnt/β-catenin pathway; and hence plays a critical role in cancer. Therefore, in current study, ceramide, COX-2, 15-deoxy prostaglandin J2(15-deoxy PGJ2, PPARγ, and β-catenin were estimated to evaluate the effect of fish oil on lipid mediated and Wnt/β-catenin signaling in colon carcinoma. Male Wistar rats in Group I received purified diet while Groups II and III received modified diet supplemented with FO : CO(1 : 1 and FO : CO(2.5 : 1, respectively. These were further subdivided into controls receiving ethylenediaminetetraacetic acid and treated groups receiving dimethylhydrazine dihydrochloride (DMH/week for 4 weeks. Animals sacrificed 48 hours after last injection constituted initiation phase and those sacrificed after 16 weeks constituted postinitiation phase. Decreased ceramide and increased PPARγ were observed in postinitiation phase only. On receiving FO+CO(1 : 1+DMH and FO+CO(2.5 : 1+DMH in both phases, ceramide was augmented whereas COX-2, 15-deoxy PGJ2, and nuclear translocation of β-catenin were reduced with respect to cancerous animals. Decrease was more significant in postinitiation phase with FO+CO(2.5 : 1+DMH. Treatment with oils increased PPARγ in initiation phase but decreased it in postinitiation phase. Hence, fish oil altered lipid mediated signalling in a dose and time dependent manner so as to inhibit progression of colon cancer.

Characterization of murine melanocortin receptors mediating adipocyte lipolysis and examination of signalling pathways involved

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Møller, Cathrine Laustrup; Raun, Kirsten; Jacobsen, Marianne Lambert

2011-01-01

hormone (a-MSH) generated from proopiomelanocortin (POMC), as well as synthetic MSH analogues to stimulate lipolysis in murine 3T3-L1 adipocytes it is shown that MC2R and MC5R are lipolytic mediators in differentiated 3T3-L1 adipocytes. Involvement of cAMP, phosphorylated extracellular signal...

Non-Dioxin-Like Polychlorinated Biphenyls Inhibit G-Protein Coupled Receptor-Mediated Ca2+ Signaling by Blocking Store-Operated Ca2+ Entry.

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Se-Young Choi

Full Text Available Polychlorinated biphenyls (PCBs are ubiquitous pollutants which accumulate in the food chain. Recently, several molecular mechanisms by which non-dioxin-like (NDL PCBs mediate neurodevelopmental and neurobehavioral toxicity have been elucidated. However, although the G-protein coupled receptor (GPCR is a significant target for neurobehavioral disturbance, our understanding of the effects of PCBs on GPCR signaling remains unclear. In this study, we investigated the effects of NDL-PCBs on GPCR-mediated Ca2+ signaling in PC12 cells. We found that ortho-substituted 2,2',6-trichlorinated biphenyl (PCB19 caused a rapid decline in the Ca2+ signaling of bradykinin, a typical Gq- and phospholipase Cβ-coupled GPCR, without any effect on its inositol 1,4,5-trisphosphate production. PCB19 reduced thapsigargin-induced sustained cytosolic Ca2+ levels, suggesting that PCB19 inhibits SOCE. The abilities of other NDL-PCBs to inhibit store-operated Ca2+ entry (SOCE were also examined and found to be of similar potencies to that of PCB19. PCB19 also showed a manner equivalent to that of known SOCE inhibitors. PCB19-mediated SOCE inhibition was confirmed by demonstrating the ability of PCB19 to inhibit the SOCE current and thapsigargin-induced Mn2+ influx. These results imply that one of the molecular mechanism by which NDL-PCBs cause neurobehavioral disturbances involves NDL-PCB-mediated inhibition of SOCE, thereby interfering with GPCR-mediated Ca2+ signaling.

Global mapping of protein phosphorylation events identifies novel signalling hubs mediating fatty acid starvation responses in Saccharomyces cerevisiae

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Pultz, Dennis; Bennetzen, Martin; Rødkær, Steven Vestergaard

2011-01-01

Dietary restriction (DR) extends the life span of multiple species, ranging from single-celled organisms like yeast to mammals. This increase in longevity by dietary restriction is coupled to profound beneficial effects on age-related pathology. Despite the number of studies on DR...... and the physiological changes DR induces, only little is known about the genetics and signalling networks, which regulate the DR response. We have recently shown that inhibition of fatty acid synthesis in Saccharomyces cerevisiae induces autophagy mediated by TORC1 signalling and affects life span. In the present study...... in a temporal manner in response to inhibition of fatty acid synthesis by cerulenin. By in silico analysis of these phosphorylation events, we have identified the major downstream regulated processes and signalling networks mediating the cellular response to fatty acid starvation. The analysis further...

Distinct signalling properties of insulin receptor substrate (IRS)-1 and IRS-2 in mediating insulin/IGF-1 action

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Rabiee, Atefeh; Krüger, Marcus; Ardenkjær-Larsen, Jacob

2018-01-01

Insulin/IGF-1 action is driven by a complex and highly integrated signalling network. Loss-of-function studies indicate that the major insulin/IGF-1 receptor substrate (IRS) proteins, IRS-1 and IRS-2, mediate different biological functions in vitro and in vivo, suggesting specific signalling...... properties despite their high degree of homology. To identify mechanisms contributing to the differential signalling properties of IRS-1 and IRS-2 in the mediation of insulin/IGF-1 action, we performed comprehensive mass spectrometry (MS)-based phosphoproteomic profiling of brown preadipocytes from wild type......, IRS-1-/-and IRS-2-/-mice in the basal and IGF-1-stimulated states. We applied stable isotope labeling by amino acids in cell culture (SILAC) for the accurate quantitation of changes in protein phosphorylation. We found ~10% of the 6262 unique phosphorylation sites detected to be regulated by IGF-1...

Activation of Brain Somatostatin Signaling Suppresses CRF Receptor-Mediated Stress Response.

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Stengel, Andreas; Taché, Yvette F

2017-01-01

Corticotropin-releasing factor (CRF) is the hallmark brain peptide triggering the response to stress and mediates-in addition to the stimulation of the hypothalamus-pituitary-adrenal (HPA) axis-other hormonal, behavioral, autonomic and visceral components. Earlier reports indicate that somatostatin-28 injected intracerebroventricularly counteracts the acute stress-induced ACTH and catecholamine release. Mounting evidence now supports that activation of brain somatostatin signaling exerts a broader anti-stress effect by blunting the endocrine, autonomic, behavioral (with a focus on food intake) and visceral gastrointestinal motor responses through the involvement of distinct somatostatin receptor subtypes.

Interplay between TGF-β signaling and receptor tyrosine kinases in tumor development.

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Shi, Qiaoni; Chen, Ye-Guang

2017-10-01

Transforming growth factor-β (TGF-β) signaling regulates cell proliferation, differentiation, migration and death, and plays a critical role in embryogenesis and tissue homeostasis. Its deregulation results in various diseases including tumor formation. Receptor tyrosine kinases (RTKs), such as epidermal growth factor receptor (EGFR), fibroblast growth factor receptor (FGFR), vascular endothelial growth factor receptor (VEGFR) and platelet-derived growth factor receptor (PDGFR), also play key roles in the development and progression of many types of tumors. It has been realized that TGF-β signaling and RTK pathways interact with each other and their interplay is important for cancer development. They are mutually regulated and cooperatively modulate cell survival and migration, epithelial-mesenchymal transition, and tumor microenvironment to accelerate tumorigenesis and tumor metastasis. RTKs can modulate Smad-dependent transcription or cooperate with TGF-β to potentiate its oncogenic activity, while TGF-β signaling can in turn control RTK signaling by regulating their activities or expression. This review summarizes current understandings of the interplay between TGF-β signaling and RTKs and its influence on tumor development.

Tricyclic Antidepressant Amitriptyline-induced Glial Cell Line-derived Neurotrophic Factor Production Involves Pertussis Toxin-sensitive Gαi/o Activation in Astroglial Cells.

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Hisaoka-Nakashima, Kazue; Miyano, Kanako; Matsumoto, Chie; Kajitani, Naoto; Abe, Hiromi; Okada-Tsuchioka, Mami; Yokoyama, Akinobu; Uezono, Yasuhito; Morioka, Norimitsu; Nakata, Yoshihiro; Takebayashi, Minoru

2015-05-29

Further elaborating the mechanism of antidepressants, beyond modulation of monoaminergic neurotransmission, this study sought to elucidate the mechanism of amitriptyline-induced production of glial cell line-derived neurotrophic factor (GDNF) in astroglial cells. Previous studies demonstrated that an amitriptyline-evoked matrix metalloproteinase (MMP)/FGF receptor (FGFR)/FGFR substrate 2α (FRS2α)/ERK cascade is crucial for GDNF production, but how amitriptyline triggers this cascade remains unknown. MMP is activated by intracellular mediators such as G proteins, and this study sought to clarify the involvement of G protein signaling in amitriptyline-evoked GDNF production in rat C6 astroglial cells (C6 cells), primary cultured rat astrocytes, and normal human astrocytes. Amitriptyline-evoked GDNF mRNA expression and release were inhibited by pertussis toxin (PTX), a Gα(i/o) inhibitor, but not by NF449, a Gα(s) inhibitor, or YM-254890, a Gαq inhibitor. The activation of the GDNF production cascade (FGFR/FRS2α/ERK) was also inhibited by PTX. Deletion of Gα(ο1) and Gα(i3) by RNAi demonstrated that these G proteins play important roles in amitriptyline signaling. G protein activation was directly analyzed by electrical impedance-based biosensors (CellKey(TM) assay), using a label-free (without use of fluorescent proteins/probes or radioisotopes) and real time approach. Amitriptyline increased impedance, indicating Gα(i/o) activation that was suppressed by PTX treatment. The impedance evoked by amitriptyline was not affected by inhibitors of the GDNF production cascade. Furthermore, FGF2 treatment did not elicit any effect on impedance, indicating that amitriptyline targets PTX-sensitive Gα(i/o) upstream of the MMP/FGFR/FRS2α/ERK cascade. These results suggest novel targeting for the development of antidepressants. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

GPER1-mediated IGFBP-1 induction modulates IGF-1-dependent signaling in tamoxifen-treated breast cancer cells.

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Vaziri-Gohar, Ali; Houston, Kevin D

2016-02-15

Tamoxifen, a selective estrogen receptor modulator, is a commonly prescribed adjuvant therapy for estrogen receptor-α (ERα)-positive breast cancer patients. To determine if extracellular factors contribute to the modulation of IGF-1 signaling after tamoxifen treatment, MCF-7 cells were treated with IGF-1 in conditioned medium (CM) obtained from 4-OHT-treated MCF-7 cells and the accumulation of phospho-Akt (S473) was measured. CM inhibited IGF-1-dependent cell signaling and suggesting the involvement of extracellular factors (ie. IGFBPs). A significant increase in IGFBP-1 mRNA and extracellular IGFBP-1 protein was observed in 4-OHT-treated MCF-7 cells. Knockdown experiments demonstrated that both GPER1 and CREB mediate IGFBP-1 induction. Furthermore, experiments showed that 4-OHT-dependent IGFBP-1 transcription is downstream of GPER1-activation in breast cancer cells. Additionally, neutralization and knockdown experiments demonstrated a role for IGFBP-1 in the observed inhibition of IGF-1 signaling. These results suggested that 4-OHT inhibits IGF-1 signaling via GPER1 and CREB mediated extracellular IGFBP-1 accumulation in breast cancer cells. Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Intrinsic fluorescence of the clinically approved multikinase inhibitor nintedanib reveals lysosomal sequestration as resistance mechanism in FGFR-driven lung cancer.

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Englinger, Bernhard; Kallus, Sebastian; Senkiv, Julia; Heilos, Daniela; Gabler, Lisa; van Schoonhoven, Sushilla; Terenzi, Alessio; Moser, Patrick; Pirker, Christine; Timelthaler, Gerald; Jäger, Walter; Kowol, Christian R; Heffeter, Petra; Grusch, Michael; Berger, Walter

2017-09-07

Studying the intracellular distribution of pharmacological agents, including anticancer compounds, is of central importance in biomedical research. It constitutes a prerequisite for a better understanding of the molecular mechanisms underlying drug action and resistance development. Hyperactivated fibroblast growth factor receptors (FGFRs) constitute a promising therapy target in several types of malignancies including lung cancer. The clinically approved small-molecule FGFR inhibitor nintedanib exerts strong cytotoxicity in FGFR-driven lung cancer cells. However, subcellular pharmacokinetics of this compound and its impact on therapeutic efficacy remain obscure. 3-dimensional fluorescence spectroscopy was conducted to asses cell-free nintedanib fluorescence properties. MTT assay was used to determine the impact of the lysosome-targeting agents bafilomycin A1 and chloroquine combined with nintedanib on lung cancer cell viability. Flow cytometry and live cell as well as confocal microscopy were performed to analyze uptake kinetics as well as subcellular distribution of nintedanib. Western blot was conducted to investigate protein expression. Cryosections of subcutaneous tumor allografts were generated to detect intratumoral nintedanib in mice after oral drug administration. Here, we report for the first time drug-intrinsic fluorescence properties of nintedanib in living and fixed cancer cells as well as in cryosections derived from allograft tumors of orally treated mice. Using this feature in conjunction with flow cytometry and confocal microscopy allowed to determine cellular drug accumulation levels, impact of the ABCB1 efflux pump and to uncover nintedanib trapping into lysosomes. Lysosomal sequestration - resulting in an organelle-specific and pH-dependent nintedanib fluorescence - was identified as an intrinsic resistance mechanism in FGFR-driven lung cancer cells. Accordingly, combination of nintedanib with agents compromising lysosomal acidification

FGFR1OP tagSNP but not CCR6 polymorphisms are associated with Vogt-Koyanagi-Harada syndrome in Chinese Han.

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Xianglong Yi

Full Text Available BACKGROUND: Polymorphisms of the CC chemokine receptor 6 (CCR6 and FGFR10P tagSNP (locus close to CCR6 at 6q27 have recently been reported to be associated with the susceptibility to several immune-related diseases. This study was designed to determine the association of CCR6 and FGFR10P (tagSNPs with Vogt-Koyanagi-Harada (VKH syndrome, an autoimmune disease directed against melanocytes, in two independent Chinese Han populations. METHODOLOGY/PRINCIPAL FINDINGS: A total of 601 VKH patients and 725 healthy controls from two Chinese Han populations were genotyped by the polymerase chain reaction-restriction fragment length polymorphism method. Hardy-Weinberg equilibrium was tested using the χ(2 test. Genotype frequencies were estimated by direct counting. Allele and genotype frequencies were compared between patients and controls using the χ(2 test. The frequency of the A allele of rs2301436 was significantly higher both in Cohort 1 and Cohort 2 as compared with two separate controls (P = 0.044; P = 0.049, respectively. The significance was lost after Bonferroni correction in both cohorts (Pc = 0.516; Pc = 0.392, respectively. The frequency of the A allele was significantly higher in the combined patient group as compared with all controls before and after Bonferroni correction (P = 0.005, Pc = 0.025. The genotype and allele frequencies of rs3093024, rs6902119, rs3093023 and rs968334 were not different between patients with VKH and healthy controls based on analysis either for both cohorts or for the patients and controls in total. Analysis according to extra ocular clinical findings including headache, alopecia and poliosis, vitiligo and tinnitus did not show any association of the five polymorphisms with these parameters. CONCLUSION: These results suggest that the rs2301436 tagSNP of FGFR10P is positively associated with susceptibility to VKH syndrome in the tested Chinese Han populations. No association was found for

Influence of type I IFN signaling on anti-MOG antibody-mediated demyelination

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Berg, Carsten Tue; Khorooshi, Reza M. H.; Asgari, Nasrin

2017-01-01

Background Antibodies with specificity for myelin oligodendrocyte glycoprotein (MOG) are implicated in multiple sclerosis and related diseases. The pathogenic importance of anti-MOG antibody in primary demyelinating pathology remains poorly characterized. Objective The objective of this study...... is to investigate whether administration of anti-MOG antibody would be sufficient for demyelination and to determine if type I interferon (IFN) signaling plays a similar role in anti-MOG antibody-mediated pathology, as has been shown for neuromyelitis optica-like pathology. Methods Purified IgG2a monoclonal anti...... demyelination in wild-type and IFNAR1-KO mice. Conclusions Anti-MOG antibody and complement was sufficient to induce callosal demyelination, and pathology was dependent on type I IFN. Induction of EAE in IFNAR1-KO mice overcame the dependence on type I IFN for anti-MOG and complement-mediated demyelination....

Mapping glucose-mediated gut-to-brain signalling pathways in humans.

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Little, Tanya J; McKie, Shane; Jones, Richard B; D'Amato, Massimo; Smith, Craig; Kiss, Orsolya; Thompson, David G; McLaughlin, John T

2014-08-01

Previous fMRI studies have demonstrated that glucose decreases the hypothalamic BOLD response in humans. However, the mechanisms underlying the CNS response to glucose have not been defined. We recently demonstrated that the slowing of gastric emptying by glucose is dependent on activation of the gut peptide cholecystokinin (CCK1) receptor. Using physiological functional magnetic resonance imaging this study aimed to determine the whole brain response to glucose, and whether CCK plays a central role. Changes in blood oxygenation level-dependent (BOLD) signal were monitored using fMRI in 12 healthy subjects following intragastric infusion (250ml) of: 1M glucose+predosing with dexloxiglumide (CCK1 receptor antagonist), 1M glucose+placebo, or 0.9% saline (control)+placebo, in a single-blind, randomised fashion. Gallbladder volume, blood glucose, insulin, and GLP-1 and CCK concentrations were determined. Hunger, fullness and nausea scores were also recorded. Intragastric glucose elevated plasma glucose, insulin, and GLP-1, and reduced gall bladder volume (an in vivo assay for CCK secretion). Glucose decreased BOLD signal, relative to saline, in the brainstem and hypothalamus as well as the cerebellum, right occipital cortex, putamen and thalamus. The timing of the BOLD signal decrease was negatively correlated with the rise in blood glucose and insulin levels. The glucose+dex arm highlighted a CCK1-receptor dependent increase in BOLD signal only in the motor cortex. Glucose induces site-specific differences in BOLD response in the human brain; the brainstem and hypothalamus show a CCK1 receptor-independent reduction which is likely to be mediated by a circulatory effect of glucose and insulin, whereas the motor cortex shows an early dexloxiglumide-reversible increase in signal, suggesting a CCK1 receptor-dependent neural pathway. Copyright © 2014. Published by Elsevier Inc.

Mapping glucose-mediated gut-to-brain signalling pathways in humans☆

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Little, Tanya J.; McKie, Shane; Jones, Richard B.; D'Amato, Massimo; Smith, Craig; Kiss, Orsolya; Thompson, David G.; McLaughlin, John T.

2014-01-01

Objectives Previous fMRI studies have demonstrated that glucose decreases the hypothalamic BOLD response in humans. However, the mechanisms underlying the CNS response to glucose have not been defined. We recently demonstrated that the slowing of gastric emptying by glucose is dependent on activation of the gut peptide cholecystokinin (CCK1) receptor. Using physiological functional magnetic resonance imaging this study aimed to determine the whole brain response to glucose, and whether CCK plays a central role. Experimental design Changes in blood oxygenation level-dependent (BOLD) signal were monitored using fMRI in 12 healthy subjects following intragastric infusion (250 ml) of: 1 M glucose + predosing with dexloxiglumide (CCK1 receptor antagonist), 1 M glucose + placebo, or 0.9% saline (control) + placebo, in a single-blind, randomised fashion. Gallbladder volume, blood glucose, insulin, and GLP-1 and CCK concentrations were determined. Hunger, fullness and nausea scores were also recorded. Principal observations Intragastric glucose elevated plasma glucose, insulin, and GLP-1, and reduced gall bladder volume (an in vivo assay for CCK secretion). Glucose decreased BOLD signal, relative to saline, in the brainstem and hypothalamus as well as the cerebellum, right occipital cortex, putamen and thalamus. The timing of the BOLD signal decrease was negatively correlated with the rise in blood glucose and insulin levels. The glucose + dex arm highlighted a CCK1-receptor dependent increase in BOLD signal only in the motor cortex. Conclusions Glucose induces site-specific differences in BOLD response in the human brain; the brainstem and hypothalamus show a CCK1 receptor-independent reduction which is likely to be mediated by a circulatory effect of glucose and insulin, whereas the motor cortex shows an early dexloxiglumide-reversible increase in signal, suggesting a CCK1 receptor-dependent neural pathway. PMID:24685436

Importance of Mediator complex in the regulation and integration of diverse signaling pathways in plants.

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Samanta, Subhasis; Thakur, Jitendra K

2015-01-01

Basic transcriptional machinery in eukaryotes is assisted by a number of cofactors, which either increase or decrease the rate of transcription. Mediator complex is one such cofactor, and recently has drawn a lot of interest because of its integrative power to converge different signaling pathways before channeling the transcription instructions to the RNA polymerase II machinery. Like yeast and metazoans, plants do possess the Mediator complex across the kingdom, and its isolation and subunit analyses have been reported from the model plant, Arabidopsis. Genetic, and molecular analyses have unraveled important regulatory roles of Mediator subunits at every stage of plant life cycle starting from flowering to embryo and organ development, to even size determination. It also contributes immensely to the survival of plants against different environmental vagaries by the timely activation of its resistance mechanisms. Here, we have provided an overview of plant Mediator complex starting from its discovery to regulation of stoichiometry of its subunits. We have also reviewed involvement of different Mediator subunits in different processes and pathways including defense response pathways evoked by diverse biotic cues. Wherever possible, attempts have been made to provide mechanistic insight of Mediator's involvement in these processes.

Signal-off Electrochemiluminescence Biosensor Based on Phi29 DNA Polymerase Mediated Strand Displacement Amplification for MicroRNA Detection.

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Chen, Anyi; Gui, Guo-Feng; Zhuo, Ying; Chai, Ya-Qin; Xiang, Yun; Yuan, Ruo

2015-06-16

A target induced cycling strand displacement amplification (SDA) mediated by phi29 DNA polymerase (phi29) was first investigated and applied in a signal-off electrochemiluminescence (ECL) biosensor for microRNA (miRNA) detection. Herein, the target miRNA triggered the phi29-mediated SDA which could produce amounts of single-stranded DNA (assistant probe) with accurate and comprehensive nucleotide sequence. Then, the assistant probe hybridized with the capture probe and the ferrocene-labeled probe (Fc-probe) to form a ternary "Y" structure for ECL signal quenching by ferrocene. Therefore, the ECL intensity would decrease with increasing concentration of the target miRNA, and the sensitivity of biosensor would be promoted on account of the efficient signal amplification of the target induced cycling reaction. Besides, a self-enhanced Ru(II) ECL system was designed to obtain a stable and strong initial signal to further improve the sensitivity. The ECL assay for miRNA-21 detection is developed with excellent sensitivity of a concentration variation from 10 aM to 1.0 pM and limit of detection down to 3.3 aM.

A novel role of sesamol in inhibiting NF-κB-mediated signaling in platelet activation

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Chang Chao-Chien

2011-12-01

Full Text Available Abstract Background Platelet activation is relevant to a variety of coronary heart diseases. Our previous studies revealed that sesamol possesses potent antiplatelet activity through increasing cyclic AMP formation. Although platelets are anucleated cells, they also express the transcription factor, NF-κB, that may exert non-genomic functions in platelet activation. Therefore, we further investigated the inhibitory roles of sesamol in NF-κB-mediated platelet function. Methods Platelet aggregation, Fura 2-AM fluorescence, and immunoblotting analysis were used in this study. Results NF-κB signaling events, including IKKβ phosphorylation, IκBα degradation, and p65 phosphorylation, were markedly activated by collagen (1 μg/ml in washed human platelets, and these signaling events were attenuated by sesamol (2.5~25 μM. Furthermore, SQ22536 and ODQ, inhibitors of adenylate cyclase and guanylate cyclase, respectively, strongly reversed the sesamol (25 μM-mediated inhibitory effects of IKKβ phosphorylation, IκBα degradation, and p65 phosphorylation stimulated by collagen. The protein kinase A (PKA inhibitor, H89, also reversed sesamol-mediated inhibition of IκBα degradation. Moreover, BAY11-7082, an NF-κB inhibitor, abolished IκBα degradation, phospholipase C (PLCγ2 phosphorylation, protein kinase C (PKC activation, [Ca2+]i mobilization, and platelet aggregation stimulated by collagen. Preincubation of platelets with the inhibitors, SQ22536 and H89, both strongly reversed sesamol-mediated inhibition of platelet aggregation and [Ca2+]i mobilization. Conclusions Sesamol activates cAMP-PKA signaling, followed by inhibition of the NF-κB-PLC-PKC cascade, thereby leading to inhibition of [Ca2+]i mobilization and platelet aggregation. Because platelet activation is not only linked to hemostasis, but also has a relevant role in inflammation and metastasis, our data demonstrating that inhibition of NF-κB interferes with platelet function may

The MET/AXL/FGFR Inhibitor S49076 Impairs Aurora B Activity and Improves the Antitumor Efficacy of Radiotherapy.

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Clémenson, Céline; Chargari, Cyrus; Liu, Winchygn; Mondini, Michele; Ferté, Charles; Burbridge, Mike F; Cattan, Valérie; Jacquet-Bescond, Anne; Deutsch, Eric

2017-10-01

Several therapeutic agents targeting HGF/MET signaling are under clinical development as single agents or in combination, notably with anti-EGFR therapies in non-small cell lung cancer (NSCLC). However, despite increasing data supporting a link between MET, irradiation, and cancer progression, no data regarding the combination of MET-targeting agents and radiotherapy are available from the clinic. S49076 is an oral ATP-competitive inhibitor of MET, AXL, and FGFR1-3 receptors that is currently in phase I/II clinical trials in combination with gefitinib in NSCLC patients whose tumors show resistance to EGFR inhibitors. Here, we studied the impact of S49076 on MET signaling, cell proliferation, and clonogenic survival in MET-dependent (GTL16 and U87-MG) and MET-independent (H441, H460, and A549) cells. Our data show that S49076 exerts its cytotoxic activity at low doses on MET-dependent cells through MET inhibition, whereas it inhibits growth of MET-independent cells at higher but clinically relevant doses by targeting Aurora B. Furthermore, we found that S49076 improves the antitumor efficacy of radiotherapy in both MET-dependent and MET-independent cell lines in vitro and in subcutaneous and orthotopic tumor models in vivo In conclusion, our study demonstrates that S49076 has dual antitumor activity and can be used in combination with radiotherapy for the treatment of both MET-dependent and MET-independent tumors. These results support the evaluation of combined treatment of S49076 with radiation in clinical trials without patient selection based on the tumor MET dependency status. Mol Cancer Ther; 16(10); 2107-19. ©2017 AACR . ©2017 American Association for Cancer Research.

Expression of proteins FGFR3, PI3K, AKT, p21Waf1/Cip1 and cyclins D1 and D3 in patients with T1 bladder tumours: clinical implications and prognostic significance.

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Blanca Pedregosa, A M; Sánchez-González, Á; Carrasco Valiente, J; Ruiz García, J M; Gómez Gómez, E; López Beltrán, A; Requena Tapia, M J

2017-04-01

To determine the differential protein expression of biomarkers FGFR3, PI3K (subunits PI3Kp110α, PI3KClassIII, PI3Kp85), AKT, p21Waf1/Cip1 and cyclins D1 and D3 in T1 bladder cancer versus healthy tissue and to study their potential role as early recurrence markers. This is a prospective study that employed a total of 67 tissue samples (55 cases of T1 bladder tumours that underwent transurethral resection and 12 cases of adjacent healthy mucosa). The protein expression levels were assessed using Western blot, and the means and percentages were compared using Student's t-test and the chi-squared test. The survival analysis was conducted using the Kaplan-Meier method and the log-rank test. Greater protein expression was detected for FGFR3, PI3Kp110α, PI3KClassIII, cyclins D1 and D3 and p21Waf1/Cip1 in the tumour tissue than in the healthy mucosa. However, these differences were not significant for PI3Kp85 and AKT. We observed statistically significant correlations between early recurrence and PI3Kp110α, PI3KClassIII, PI3Kp85 and AKT (P=.003, P=.045, P=.050 and P=.028, respectively), between the tumour type (primary vs. recurrence) and cyclin D3 (P=.001), between the tumour size and FGFR3 (P=.035) and between multifocality and cyclin D1 (P=.039). The survival analysis selected FGFR3 (P=.024), PI3Kp110α (P=.014), PI3KClassIII (P=.042) and AKT (P=.008) as markers of early-recurrence-free survival. There is an increase in protein expression levels in bladder tumour tissue. The overexpression of FGFR3, PI3Kp110α, PI3KClassIII and AKT is associated with increased early-recurrence-free survival for patients with T1 bladder tumours. Copyright © 2016 AEU. Publicado por Elsevier España, S.L.U. All rights reserved.

Glucose delays the insulin-induced increase in thyroid hormone-mediated signaling in adipose of prolong-fasted elephant seal pups

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Soñanez-Organis, José G.; Viscarra, Jose A.; Jaques, John T.; MacKenzie, Duncan S.; Crocker, Daniel E.; Ortiz, Rudy M.

2016-01-01

Prolonged food deprivation in mammals typically reduces glucose, insulin, and thyroid hormone (TH) concentrations, as well as tissue deiodinase (DI) content and activity, which, collectively, suppress metabolism. However, in elephant seal pups, prolonged fasting does not suppress TH levels; it is associated with upregulation of adipose TH-mediated cellular mechanisms and adipose-specific insulin resistance. The functional relevance of this apparent paradox and the effects of glucose and insulin on TH-mediated signaling in an insulin-resistant tissue are not well defined. To address our hypothesis that insulin increases adipose TH signaling in pups during extended fasting, we assessed the changes in TH-associated genes in response to an insulin infusion in early- and late-fasted pups. In late fasting, insulin increased DI1, DI2, and THrβ-1 mRNA expression by 566%, 44%, and 267% at 60 min postinfusion, respectively, with levels decreasing by 120 min. Additionally, we performed a glucose challenge in late-fasted pups to differentiate between insulin- and glucose-mediated effects on TH signaling. In contrast to the insulin-induced effects, glucose infusion did not increase the expressions of DI1, DI2, and THrβ-1 until 120 min, suggesting that glucose delays the onset of the insulin-induced effects. The data also suggest that fasting duration increases the sensitivity of adipose TH-mediated mechanisms to insulin, some of which may be mediated by increased glucose. These responses appear to be unique among mammals and to have evolved in elephant seals to facilitate their adaptation to tolerate an extreme physiological condition. PMID:26739649

HIF-mediated innate immune responses: cell signaling and therapeutic implications

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Harris AJ

2014-05-01

Full Text Available Alison J Harris, AA Roger Thompson, Moira KB Whyte, Sarah R Walmsley Academic Unit of Respiratory Medicine, Department of Infection and Immunity, University of Sheffield, Sheffield, UK Abstract: Leukocytes recruited to infected, damaged, or inflamed tissues during an immune response must adapt to oxygen levels much lower than those in the circulation. Hypoxia inducible factors (HIFs are key mediators of cellular responses to hypoxia and, as in other cell types, HIFs are critical for the upregulation of glycolysis, which enables innate immune cells to produce adenosine triphosphate anaerobically. An increasing body of evidence demonstrates that hypoxia also regulates many other innate immunological functions, including cell migration, apoptosis, phagocytosis of pathogens, antigen presentation and production of cytokines, chemokines, and angiogenic and antimicrobial factors. Many of these functions are mediated by HIFs, which are not only stabilized posttranslationally by hypoxia, but also transcriptionally upregulated by inflammatory signals. Here, we review the role of HIFs in the responses of innate immune cells to hypoxia, both in vitro and in vivo, with a particular focus on myeloid cells, on which the majority of studies have so far been carried out. Keywords: hypoxia, neutrophils, monocytes, macrophages

Charge-signal multiplication mediated by urea wires inside Y-shaped carbon nanotubes

International Nuclear Information System (INIS)

Lv, Mei; Liu, Zengrong; He, Bing; Xiu, Peng; Tu, Yusong

2014-01-01

In previous studies, we reported molecular dynamics (MD) simulations showing that single-file water wires confined inside Y-shaped single-walled carbon nanotubes (Y-SWNTs) held strong and robust capability to convert and multiply charge signals [Y. S. Tu, P. Xiu, R. Z. Wan, J. Hu, R. H. Zhou, and H. P. Fang, Proc. Natl. Acad. Sci. U.S.A. 106, 18120 (2009); Y. Tu, H. Lu, Y. Zhang, T. Huynh, and R. Zhou, J. Chem. Phys. 138, 015104 (2013)]. It is fascinating to see whether the signal multiplication can be realized by other kinds of polar molecules with larger dipole moments (which make the experimental realization easier). In this article, we use MD simulations to study the urea-mediated signal conversion and multiplication with Y-SWNTs. We observe that when a Y-SWNT with an external charge of magnitude 1.0 e (the model of a signal at the single-electron level) is solvated in 1 M urea solutions, urea can induce drying of the Y-SWNT and fill its interiors in single-file, forming Y-shaped urea wires. The external charge can effectively control the dipole orientation of the urea wire inside the main channel (i.e., the signal can be readily converted), and this signal can further be multiplied into 2 (or more) output signals by modulating dipole orientations of urea wires in bifurcated branch channels of the Y-SWNT. This remarkable signal transduction capability arises from the strong dipole-induced ordering of urea wires under extreme confinement. We also discuss the advantage of urea as compared with water in the signal multiplication, as well as the robustness and biological implications of our findings. This study provides the possibility for multiplying signals by using urea molecules (or other polar organic molecules) with Y-shaped nanochannels and might also help understand the mechanism behind signal conduction in both physical and biological systems

Orexin/hypocretin receptor 1 signaling mediates Pavlovian cue-food conditioning and extinction.

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Keefer, Sara E; Cole, Sindy; Petrovich, Gorica D

2016-08-01

Learned food cues can drive feeding in the absence of hunger, and orexin/hypocretin signaling is necessary for this type of overeating. The current study examined whether orexin also mediates cue-food learning during the acquisition and extinction of these associations. In Experiment 1, rats underwent two sessions of Pavlovian appetitive conditioning, consisting of tone-food presentations. Prior to each session, rats received either the orexin 1 receptor antagonist SB-334867 (SB) or vehicle systemically. SB treatment did not affect conditioned responses during the first conditioning session, measured as food cup behavior during the tone and latency to approach the food cup after the tone onset, compared to the vehicle group. During the second conditioning session, SB treatment attenuated learning. All groups that received SB, prior to either the first or second conditioning session, displayed significantly less food cup behavior and had longer latencies to approach the food cup after tone onset compared to the vehicle group. These findings suggest orexin signaling at the 1 receptor mediates the consolidation and recall of cue-food acquisition. In Experiment 2, another group of rats underwent tone-food conditioning sessions (drug free), followed by two extinction sessions under either SB or vehicle treatment. Similar to Experiment 1, SB did not affect conditioned responses during the first session. During the second extinction session, the group that received SB prior to the first extinction session, but vehicle prior to the second, expressed conditioned food cup responses longer after tone offset, when the pellets were previously delivered during conditioning, and maintained shorter latencies to approach the food cup compared to the other groups. The persistence of these conditioned behaviors indicates impairment in extinction consolidation due to SB treatment during the first extinction session. Together, these results demonstrate an important role for orexin

Orexin/hypocretin receptor 1 signaling mediates Pavlovian cue-food conditioning and extinction

Science.gov (United States)

Keefer, Sara E.; Cole, Sindy; Petrovich, Gorica D.

2016-01-01

Learned food cues can drive feeding in the absence of hunger, and orexin/hypocretin signaling is necessary for this type of overeating. The current study examined whether orexin also mediates cue-food learning during the acquisition and extinction of these associations. In Experiment 1, rats underwent two sessions of Pavlovian appetitive conditioning, consisting of tone-food presentations. Prior to each session, rats received either the orexin 1 receptor antagonist SB-334867 (SB) or vehicle systemically. SB treatment did not affect conditioned responses during the first conditioning session, measured as food cup behavior during the tone and latency to approach the food cup after the tone onset, compared to the vehicle group. During the second conditioning session, SB treatment attenuated learning. All groups that received SB, prior to either the first or second conditioning session, displayed significantly less food cup behavior and had longer latencies to approach the food cup after tone onset compared to the vehicle group. These findings suggest orexin signaling at the 1 receptor mediates the consolidation and recall of cue-food acquisition. In Experiment 2, another group of rats underwent tone-food conditioning sessions (drug free), followed by two extinction sessions under either SB or vehicle treatment. Similar to Experiment 1, SB did not affect conditioned responses during the first session. During the second extinction session, the group that received SB prior to the first extinction session, but vehicle prior to the second, expressed conditioned food cup responses longer after tone offset, when the pellets were previously delivered during conditioning, and maintained shorter latencies to approach the food cup compared to the other groups. The persistence of these conditioned behaviors indicates impairment in extinction consolidation due to SB treatment during the first extinction session. Together, these results demonstrate an important role for orexin

Helicobacter pylori-derived Heat shock protein 60 enhances angiogenesis via a CXCR2-mediated signaling pathway

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Lin, Chen-Si [Department of Biological Science and Technology, National Chiao-Tung University, Hsin-Chu, Taiwan (China); School of Veterinary Medicine, National Taiwan University, Taipei, Taiwan (China); He, Pei-Juin; Hsu, Wei-Tung [Department of Biological Science and Technology, National Chiao-Tung University, Hsin-Chu, Taiwan (China); Wu, Ming-Shiang [Department of Internal Medicine, College of Medicine, National Taiwan University, Taipei, Taiwan (China); Wu, Chang-Jer [Department of Food Science, National Taiwan Ocean University, Keelung, Taiwan (China); Shen, Hsiao-Wei [Institute of Molecular Medicine and Bioengineering, National Chiao-Tung University, Hsin-Chu, Taiwan (China); Hwang, Chia-Hsiang [Yung-Shin Pharmaceutical Industry Co., Ltd., Tachia, Taichung, Taiwan (China); Lai, Yiu-Kay [Department of Life Science, Institute of Biotechnology, National Tsing Hua University, Hsin-Chu, Taiwan (China); Tsai, Nu-Man [School of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung, Taiwan (China); Liao, Kuang-Wen, E-mail: kitchhen@yahoo.com.tw [Institute of Molecular Medicine and Bioengineering, National Chiao-Tung University, Hsin-Chu, Taiwan (China)

2010-06-25

Helicobacter pylori is a potent carcinogen associated with gastric cancer malignancy. Recently, H. pylori Heat shock protein 60 (HpHSP60) has been reported to promote cancer development by inducing chronic inflammation and promoting tumor cell migration. This study demonstrates a role for HpHSP60 in angiogenesis, a necessary precursor to tumor growth. We showed that HpHSP60 enhanced cell migration and tube formation, but not cell proliferation, in human umbilical vein endothelial cells (HUVECs). HpHSP60 also indirectly promoted HUVEC proliferation when HUVECs were co-cultured with supernatants collected from HpHSP60-treated AGS or THP-1 cells. The angiogenic array showed that HpHSP60 dramatically induced THP-1 cells and HUVECs to produce the chemotactic factors IL-8 and GRO. Inhibition of CXCR2, the receptor for IL-8 and GRO, or downstream PLC{beta}2/Ca2+-mediated signaling, significantly abolished HpHSP60-induced tube formation. In contrast, suppression of MAP K or PI3 K signaling did not affect HpHSP60-mediated tubulogenesis. These data suggest that HpHSP60 enhances angiogenesis via CXCR2/PLC{beta}2/Ca2+ signal transduction in endothelial cells.

Helicobacter pylori-derived Heat shock protein 60 enhances angiogenesis via a CXCR2-mediated signaling pathway

International Nuclear Information System (INIS)

Lin, Chen-Si; He, Pei-Juin; Hsu, Wei-Tung; Wu, Ming-Shiang; Wu, Chang-Jer; Shen, Hsiao-Wei; Hwang, Chia-Hsiang; Lai, Yiu-Kay; Tsai, Nu-Man; Liao, Kuang-Wen

2010-01-01

Helicobacter pylori is a potent carcinogen associated with gastric cancer malignancy. Recently, H. pylori Heat shock protein 60 (HpHSP60) has been reported to promote cancer development by inducing chronic inflammation and promoting tumor cell migration. This study demonstrates a role for HpHSP60 in angiogenesis, a necessary precursor to tumor growth. We showed that HpHSP60 enhanced cell migration and tube formation, but not cell proliferation, in human umbilical vein endothelial cells (HUVECs). HpHSP60 also indirectly promoted HUVEC proliferation when HUVECs were co-cultured with supernatants collected from HpHSP60-treated AGS or THP-1 cells. The angiogenic array showed that HpHSP60 dramatically induced THP-1 cells and HUVECs to produce the chemotactic factors IL-8 and GRO. Inhibition of CXCR2, the receptor for IL-8 and GRO, or downstream PLCβ2/Ca2+-mediated signaling, significantly abolished HpHSP60-induced tube formation. In contrast, suppression of MAP K or PI3 K signaling did not affect HpHSP60-mediated tubulogenesis. These data suggest that HpHSP60 enhances angiogenesis via CXCR2/PLCβ2/Ca2+ signal transduction in endothelial cells.

c-Fms signaling mediates neurofibromatosis Type-1 osteoclast gain-in-functions.

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Yongzheng He

Full Text Available Skeletal abnormalities including osteoporosis and osteopenia occur frequently in both pediatric and adult neurofibromatosis type 1 (NF1 patients. NF1 (Nf1 haploinsufficient osteoclasts and osteoclast progenitors derived from both NF1 patients and Nf1(+/- mice exhibit increased differentiation, migration, and bone resorptive capacity in vitro, mediated by hyperactivation of p21(Ras in response to limiting concentrations of macrophage-colony stimulating factor (M-CSF. Here, we show that M-CSF binding to its receptor, c-Fms, results in increased c-Fms activation in Nf1(+/ (- osteoclast progenitors, mediating multiple gain-in-functions through the downstream effectors Erk1/2 and p90RSK. PLX3397, a potent and selective c-Fms inhibitor, attenuated M-CSF mediated Nf1(+/- osteoclast migration by 50%, adhesion by 70%, and pit formation by 60%. In vivo, we administered PLX3397 to Nf1(+/- osteoporotic mice induced by ovariectomy (OVX and evaluated changes in bone mass and skeletal architecture. We found that PLX3397 prevented bone loss in Nf1(+/--OVX mice by reducing osteoclast differentiation and bone resorptive activity in vivo. Collectively, these results implicate the M-CSF/c-Fms signaling axis as a critical pathway underlying the aberrant functioning of Nf1 haploinsufficient osteoclasts and may provide a potential therapeutic target for treating NF1 associated osteoporosis and osteopenia.

The BDNF/TrkB signaling pathway is involved in heat hyperalgesia mediated by Cdk5 in rats.

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Hong-Hai Zhang

Full Text Available Cyclin-dependent kinase 5 (Cdk5 has been shown to play an important role in mediating inflammation-induced heat hyperalgesia. However, the underlying mechanism remains unclear. The aim of this study was to determine whether roscovitine, an inhibitor of Cdk5, could reverse the heat hyperalgesia induced by peripheral injection of complete Freund's adjuvant (CFA via the brain-derived neurotrophic factor (BDNF-tyrosine kinase B (TrkB signaling pathway in the dorsal horn of the spinal cord in rats.Heat hyperalgesia induced by peripheral injection of CFA was significantly reversed by roscovitine, TrkB-IgG, and the TrkB inhibitor K252a, respectively. Furthermore, BDNF was significantly increased from 0.5 h to 24 h after CFA injection in the spinal cord dorsal horn. Intrathecal adminstration of the Cdk5 inhibitor roscovitine had no obvious effects on BDNF levels. Increased TrkB protein level was significantly reversed by roscovitine between 0.5 h and 6 h after CFA injection. Cdk5 and TrkB co-immunoprecipitation results suggested Cdk5 mediates the heat hyperalgesia induced by CFA injection by binding with TrkB, and the binding between Cdk5 and TrkB was markedly blocked by intrathecal adminstration of roscovitine.Our data suggested that the BDNF-TrkB signaling pathway was involved in CFA-induced heat hyperalgesia mediated by Cdk5. Roscovitine reversed the heat hyperalgesia induced by peripheral injection of CFA by blocking BDNF/TrkB signaling pathway, suggesting that severing the close crosstalk between Cdk5 and the BDNF/TrkB signaling cascade may present a potential target for anti-inflammatory pain.

Redox Signaling Mediated by Thioredoxin and Glutathione Systems in the Central Nervous System.

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Ren, Xiaoyuan; Zou, Lili; Zhang, Xu; Branco, Vasco; Wang, Jun; Carvalho, Cristina; Holmgren, Arne; Lu, Jun

2017-11-01

The thioredoxin (Trx) and glutathione (GSH) systems play important roles in maintaining the redox balance in the brain, a tissue that is prone to oxidative stress due to its high-energy demand. These two disulfide reductase systems are active in various areas of the brain and are considered to be critical antioxidant systems in the central nervous system (CNS). Various neuronal disorders have been characterized to have imbalanced redox homeostasis. Recent Advances: In addition to their detrimental effects, recent studies have highlighted that reactive oxygen species/reactive nitrogen species (ROS/RNS) act as critical signaling molecules by modifying thiols in proteins. The Trx and GSH systems, which reversibly regulate thiol modifications, regulate redox signaling involved in various biological events in the CNS. In this review, we focus on the following: (i) how ROS/RNS are produced and mediate signaling in CNS; (ii) how Trx and GSH systems regulate redox signaling by catalyzing reversible thiol modifications; (iii) how dysfunction of the Trx and GSH systems causes alterations of cellular redox signaling in human neuronal diseases; and (iv) the effects of certain small molecules that target thiol-based signaling pathways in the CNS. Further study on the roles of thiol-dependent redox systems in the CNS will improve our understanding of the pathogenesis of many human neuronal disorders and also help to develop novel protective and therapeutic strategies against neuronal diseases. Antioxid. Redox Signal. 27, 989-1010.

Interplay between Dioxin-Mediated Signaling and Circadian Clock: A Possible Determinant in Metabolic Homeostasis

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Chun Wang

2014-07-01

Full Text Available The rotation of the earth on its axis creates the environment of a 24 h solar day, which organisms on earth have used to their evolutionary advantage by integrating this timing information into their genetic make-up in the form of a circadian clock. This intrinsic molecular clock is pivotal for maintenance of synchronized homeostasis between the individual organism and the external environment to allow coordinated rhythmic physiological and behavioral function. Aryl hydrocarbon receptor (AhR is a master regulator of dioxin-mediated toxic effects, and is, therefore, critical in maintaining adaptive responses through regulating the expression of phase I/II drug metabolism enzymes. AhR expression is robustly rhythmic, and physiological cross-talk between AhR signaling and circadian rhythms has been established. Increasing evidence raises a compelling argument that disruption of endogenous circadian rhythms contributes to the development of disease, including sleep disorders, metabolic disorders and cancers. Similarly, exposure to environmental pollutants through air, water and food, is increasingly cited as contributory to these same problems. Thus, a better understanding of interactions between AhR signaling and the circadian clock regulatory network can provide critical new insights into environmentally regulated disease processes. This review highlights recent advances in the understanding of the reciprocal interactions between dioxin-mediated AhR signaling and the circadian clock including how these pathways relate to health and disease, with emphasis on the control of metabolic function.

C-Type Natriuretic Peptide Analog as Therapy for Achondroplasia.

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Legeai-Mallet, Laurence

2016-01-01

Fibroblast growth factor receptor 3 (FGFR3) is an important regulator of bone formation. Gain-of-function mutations in the FGFR3 gene result in chondrodysplasias which include achondroplasia (ACH), the most common form of dwarfism, in which skull, appendicular and axial skeletons are affected. The skeletal phenotype of patients with ACH showed defective proliferation and differentiation of the chondrocytes in the growth plate cartilage. Both endochondral and membranous ossification processes are disrupted during development. At cellular level, Fgfr3 mutations induce increased phosphorylation of the tyrosine kinase receptor FGFR3, which correlate with an enhanced activation of its downstream signaling pathways. Potential therapeutic strategies have emerged for ACH. Several preclinical studies have been conducted such as the C-type natriuretic peptide (CNP) analog (BMN111), intermittent parathyroid hormone injections, soluble FGFR3 therapy, and meclozine and statin treatments. Among the putative targets to antagonize FGFR3 signaling, CNP (or BMN111) is one of the most promising strategies. BMN111 acts as a key regulator of longitudinal bone growth by downregulating the mitogen-activated protein kinase pathway, which is activated as a result of a FGFR3 gain-of-function mutation. Preclinical studies showed that BMN111 treatment led to a large improvement in skeletal parameters in Fgfr3Y367C/+ mice mimicking ACH. In 2014, a clinical trial (phase 2) of BMN111 in pediatric patients with ACH has started. This first clinical trial marks the first big step towards real treatment for these patients. © 2016 S. Karger AG, Basel.

Inhibition of PTP1B Restores IRS1-Mediated Hepatic Insulin Signaling in IRS2-Deficient Mice

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González-Rodríguez, Águeda; Gutierrez, Jose A. Mas; Sanz-González, Silvia; Ros, Manuel; Burks, Deborah J.; Valverde, Ángela M.

2010-01-01

OBJECTIVE Mice with complete deletion of insulin receptor substrate 2 (IRS2) develop hyperglycemia, impaired hepatic insulin signaling, and elevated gluconeogenesis, whereas mice deficient for protein tyrosine phosphatase (PTP)1B display an opposing hepatic phenotype characterized by increased sensitivity to insulin. To define the relationship between these two signaling pathways in the regulation of liver metabolism, we used genetic and pharmacological approaches to study the effects of inhibiting PTP1B on hepatic insulin signaling and expression of gluconeogenic enzymes in IRS2−/− mice. RESEARCH DESIGN AND METHODS We analyzed glucose homeostasis and insulin signaling in liver and isolated hepatocytes from IRS2−/− and IRS2−/−/PTP1B−/− mice. Additionally, hepatic insulin signaling was assessed in control and IRS2−/− mice treated with resveratrol, an antioxidant present in red wine. RESULTS In livers of hyperglycemic IRS2−/− mice, the expression levels of PTP1B and its association with the insulin receptor (IR) were increased. The absence of PTP1B in the double-mutant mice restored hepatic IRS1-mediated phosphatidylinositol (PI) 3-kinase/Akt/Foxo1 signaling. Moreover, resveratrol treatment of hyperglycemic IRS2−/− mice decreased hepatic PTP1B mRNA and inhibited PTP1B activity, thereby restoring IRS1-mediated PI 3-kinase/Akt/Foxo1 signaling and peripheral insulin sensitivity. CONCLUSIONS By regulating the phosphorylation state of IR, PTB1B determines sensitivity to insulin in liver and exerts a unique role in the interplay between IRS1 and IRS2 in the modulation of hepatic insulin action. PMID:20028942

GLI1, a crucial mediator of sonic hedgehog signaling in prostate cancer, functions as a negative modulator for androgen receptor

International Nuclear Information System (INIS)

Chen, Guangchun; Goto, Yutaka; Sakamoto, Ryuichi; Tanaka, Kimitaka; Matsubara, Eri; Nakamura, Masafumi; Zheng, Hong; Lu, Jian; Takayanagi, Ryoichi; Nomura, Masatoshi

2011-01-01

Research highlights: → GLI1, which play a central role in sonic hedgehog signaling in prostate cancer, can act as a co-repressor to substantially block androgen receptor-mediated transactivation. → GLI1 directly interacts with AR. → SHH-GLI pathway might be one of determinants governing the transition of prostate cancer from an androgen-dependent to an androgen-independent state. -- Abstract: Sonic hedgehog (SHH) signaling, acting in a combinatorial manner with androgen signaling, is essential for prostate patterning and development. Recently, elevated activation of SHH signaling has been shown to play important roles in proliferation, progression and metastasis of prostate cancer. In this report, we demonstrate for the first time, that GLI1, which has been shown to play a central role in SHH signaling in prostate cancer, can act as a co-repressor to substantially block androgen receptor (AR)-mediated transactivation, at least in part, by directly interacting with AR. Our observations suggest that the SHH-GLI pathway might be one of determinants governing the transition of prostate cancer from an androgen-dependent to an androgen-independent state by compensating, or even superseding androgen signaling.

Importance of Mediator complex in the regulation and integration of diverse signaling pathways in plants

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Subhasis eSamanta

2015-09-01

Full Text Available Basic transcriptional machinery in eukaryotes is assisted by a number of cofactors, which either increase or decrease the rate of transcription. Mediator complex is one such cofactor, and recently has drawn a lot of interest because of its integrative power to converge different signaling pathways before channelling the transcription instructions to the RNA polymerase II machinery. Like yeast and metazoans, plants do possess the Mediator complex across the kingdom, and its isolation and subunit analyses have been reported from the model plant, Arabidopsis. Genetic and molecular analyses have unravelled important regulatory roles of Mediator subunits at every stage of plant life cycle starting from flowering to embryo and organ development, to even size determination. It also contributes immensely to the survival of plants against different environmental vagaries by the timely activation of its resistance mechanisms. Here, we have provided an overview of plant Mediator complex starting from its discovery to regulation of stoichiometry of its subunits. We have also reviewed involvement of different Mediator subunits in different processes and pathways including defense response pathways evoked by diverse biotic cues. Wherever possible, attempts have been made to provide mechanistic insight of Mediator’s involvement in these processes.

Caenorhabditis elegans fibroblast growth factor receptor signaling can occur independently of the multi-substrate adaptor FRS2.

Science.gov (United States)

Lo, Te-Wen; Bennett, Daniel C; Goodman, S Jay; Stern, Michael J

2010-06-01

The components of receptor tyrosine kinase signaling complexes help to define the specificity of the effects of their activation. The Caenorhabditis elegans fibroblast growth factor receptor (FGFR), EGL-15, regulates a number of processes, including sex myoblast (SM) migration guidance and fluid homeostasis, both of which require a Grb2/Sos/Ras cassette of signaling components. Here we show that SEM-5/Grb2 can bind directly to EGL-15 to mediate SM chemoattraction. A yeast two-hybrid screen identified SEM-5 as able to interact with the carboxy-terminal domain (CTD) of EGL-15, a domain that is specifically required for SM chemoattraction. This interaction requires the SEM-5 SH2-binding motifs present in the CTD (Y(1009) and Y(1087)), and these sites are required for the CTD role of EGL-15 in SM chemoattraction. SEM-5, but not the SEM-5 binding sites located in the CTD, is required for the fluid homeostasis function of EGL-15, indicating that SEM-5 can link to EGL-15 through an alternative mechanism. The multi-substrate adaptor protein FRS2 serves to link vertebrate FGFRs to Grb2. In C. elegans, an FRS2-like gene, rog-1, functions upstream of a Ras/MAPK pathway for oocyte maturation but is not required for EGL-15 function. Thus, unlike the vertebrate FGFRs, which require the multi-substrate adaptor FRS2 to recruit Grb2, EGL-15 can recruit SEM-5/Grb2 directly.

Progesterone receptor (PR) polyproline domain (PPD) mediates inhibition of epidermal growth factor receptor (EGFR) signaling in non-small cell lung cancer cells.

Science.gov (United States)

Kawprasertsri, Sornsawan; Pietras, Richard J; Marquez-Garban, Diana C; Boonyaratanakornkit, Viroj

2016-05-01

Recent evidence has suggested a possible role for progesterone receptor (PR) in the progression of non-small cell lung cancer (NSCLC). However, little is known concerning roles of PR in NSCLC. PR contains a polyproline domain (PPD), which directly binds to the SH3 domain of signaling molecules. Because PPD-SH3 interactions are essential for EGFR signaling, we hypothesized that the presence of PR-PPD interfered with EGFR-mediated signaling and cell proliferation. We examined the role of PR-PPD in cell proliferation and signaling by stably expressing PR-B, or PR-B with disrupting mutations in the PPD (PR-BΔSH3), from a tetracycline-regulated promoter in A549 NSCLC cells. PR-B dose-dependently inhibited cell growth in the absence of ligand, and progestin (R5020) treatment further suppressed the growth. Treatment with RU486 abolished PR-B- and R5020-mediated inhibition of cell proliferation. Expression of PR-BΔSH3 and treatment with R5020 or RU486 had no effect on cell proliferation. Furthermore, PR-B expression but not PR-BΔSH3 expression reduced EGF-induced A549 proliferation and activation of ERK1/2, in the absence of ligand. Taken together, our data demonstrated the significance of PR extranuclear signaling through PPD interactions in EGFR-mediated proliferation and signaling in NSCLC. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Characterization of the transport signals that mediate the nucleocytoplasmic traffic of low risk HPV11 E7

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McKee, Courtney H.; Onder, Zeynep; Ashok, Aditya; Cardoso, Rebeca; Moroianu, Junona, E-mail: moroianu@bc.edu

2013-08-15

We previously discovered that nuclear import of low risk HPV11 E7 is mediated by its zinc-binding domain via a pathway that is independent of karyopherins/importins (Piccioli et al., 2010. Virology 407, 100–109). In this study we mapped and characterized a leucine-rich nuclear export signal (NES), {sub 76}IRQLQDLLL{sub 84}, within the zinc-binding domain that mediates the nuclear export of HPV11 E7 in a CRM1-dependent manner. We also identified a mostly hydrophobic patch {sub 65}VRLVV{sub 69} within the zinc-binding domain that mediates nuclear import of HPV11 E7 via hydrophobic interactions with the FG-repeats domain of Nup62. Substitutions of hydrophobic residues to alanine within the {sub 65}VRLVV{sub 69} sequence disrupt the nuclear localization of 11E7, whereas the R66A mutation has no effect. Overall the data support a model of nuclear entry of HPV11 E7 protein via hydrophobic interactions with FG nucleoporins at the nuclear pore complex. - Highlights: • HPV11 E7 has a leucine-rich nuclear export signal that mediates its nuclear export via CRM1. • HPV11 E7 interacts via its unique cNLS with the FG domain of Nup62. • Identification of a hydrophobic patch essential for nuclear localization of HPV11 E7.

Characterization of the transport signals that mediate the nucleocytoplasmic traffic of low risk HPV11 E7

International Nuclear Information System (INIS)

McKee, Courtney H.; Onder, Zeynep; Ashok, Aditya; Cardoso, Rebeca; Moroianu, Junona

2013-01-01

We previously discovered that nuclear import of low risk HPV11 E7 is mediated by its zinc-binding domain via a pathway that is independent of karyopherins/importins (Piccioli et al., 2010. Virology 407, 100–109). In this study we mapped and characterized a leucine-rich nuclear export signal (NES), 76 IRQLQDLLL 84 , within the zinc-binding domain that mediates the nuclear export of HPV11 E7 in a CRM1-dependent manner. We also identified a mostly hydrophobic patch 65 VRLVV 69 within the zinc-binding domain that mediates nuclear import of HPV11 E7 via hydrophobic interactions with the FG-repeats domain of Nup62. Substitutions of hydrophobic residues to alanine within the 65 VRLVV 69 sequence disrupt the nuclear localization of 11E7, whereas the R66A mutation has no effect. Overall the data support a model of nuclear entry of HPV11 E7 protein via hydrophobic interactions with FG nucleoporins at the nuclear pore complex. - Highlights: • HPV11 E7 has a leucine-rich nuclear export signal that mediates its nuclear export via CRM1. • HPV11 E7 interacts via its unique cNLS with the FG domain of Nup62. • Identification of a hydrophobic patch essential for nuclear localization of HPV11 E7

Curcumin Induced Human Gastric Cancer BGC-823 Cells Apoptosis by ROS-Mediated ASK1-MKK4-JNK Stress Signaling Pathway

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Tao Liang

2014-09-01

Full Text Available The signaling mediated by stress-activated MAP kinases (MAPK, c-Jun N-terminal kinase (JNK has well-established importance in cancer. In the present report, we investigated the effects of curcumin on the signaling pathway in human gastric cancer BGC-823 cells. Curcumin induced reactive oxygen species (ROS production and BGC-823 cells apoptosis. Inhibition of ROS generation by antioxidant (NAC or Trion significantly prevented curcumin-mediated apoptosis. Notably, we observed that curcumin activated ASK1, a MAPKKK that is oxidative stress sensitive and responsible to phosphorylation of JNK via triggering cascades, up-regulated an upstream effector of the JNK, MKK4, and phosphorylated JNK protein expression in BGC-823 cells. However, curcumin induced ASK1-MKK4-JNK signaling was attenuated by NAC. All the findings confirm the possibility that oxidative stress-activated ASK1-MKK4-JNK signaling cascade promotes the apoptotic response in curcumin-treated BGC-823 cells.

Neuropilin-2 mediated β-catenin signaling and survival in human gastro-intestinal cancer cell lines.

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Shaija Samuel

Full Text Available NRP-2 is a high-affinity kinase-deficient receptor for ligands belonging to the class 3 semaphorin and vascular endothelial growth factor families. NRP-2 has been detected on the surface of several types of human cancer cells, but its expression and function in gastrointestinal (GI cancer cells remains to be determined. We sought to determine the function of NRP-2 in mediating downstream signals regulating the growth and survival of human gastrointestinal cancer cells. In human gastric cancer specimens, NRP-2 expression was detected in tumor tissues but not in adjacent normal mucosa. In CNDT 2.5 cells, shRNA mediated knockdown NRP-2 expression led to decreased migration and invasion in vitro (p<0.01. Focused gene-array analysis demonstrated that loss of NRP-2 reduced the expression of a critical metastasis mediator gene, S100A4. Steady-state levels and function of β-catenin, a known regulator of S100A4, were also decreased in the shNRP-2 clones. Furthermore, knockdown of NRP-2 sensitized CNDT 2.5 cells in vitro to 5FU toxicity. This effect was associated with activation of caspases 3 and 7, cleavage of PARP, and downregulation of Bcl-2. In vivo growth of CNDT 2.5 cells in the livers of nude mice was significantly decreased in the shNRP-2 group (p<0.05. Intraperitoneal administration of NRP-2 siRNA-DOPC decreased the tumor burden in mice (p = 0.01. Collectively, our results demonstrate that tumor cell-derived NRP-2 mediates critical survival signaling in gastrointestinal cancer cells.

Suppressor of cytokine signalling-3 expression inhibits cytokine-mediated destruction of primary mouse and rat pancreatic islets and delays allograft rejection

DEFF Research Database (Denmark)

Rønn, S G; Börjesson, A; Bruun, C

2008-01-01

The pro-inflammatory cytokines IL-1 and IFNgamma are critical molecules in immune-mediated beta cell destruction leading to type 1 diabetes mellitus. Suppressor of cytokine signalling (SOCS)-3 inhibits the cytokine-mediated destruction of insulinoma-1 cells. Here we investigate the effect of SOCS...

Insulin Signaling Augments eIF4E-Dependent Nonsense-Mediated mRNA Decay in Mammalian Cells.

Science.gov (United States)

Park, Jungyun; Ahn, Seyoung; Jayabalan, Aravinth K; Ohn, Takbum; Koh, Hyun Chul; Hwang, Jungwook

2016-07-01

Nonsense-mediated mRNA decay (NMD) modulates the level of mRNA harboring a premature termination codon (PTC) in a translation-dependent manner. Inhibition of translation is known to impair NMD; however, few studies have investigated the correlation between enhanced translation and increased NMD. Here, we demonstrate that insulin signaling events increase translation, leading to an increase in NMD of eIF4E-bound transcripts. We provide evidence that (i) insulin-mediated enhancement of translation augments NMD and rapamycin abrogates this enhancement; (ii) an increase in AKT phosphorylation due to inhibition of PTEN facilitates NMD; (iii) insulin stimulation increases the binding of up-frameshift factor 1 (UPF1), most likely to eIF4E-bound PTC-containing transcripts; and (iv) insulin stimulation induces the colocalization of UPF1 and eIF4E in processing bodies. These results illustrate how extracellular signaling promotes the removal of eIF4E-bound NMD targets. Copyright © 2015 Elsevier B.V. All rights reserved.

mTOR signaling promotes stem cell activation via counterbalancing BMP-mediated suppression during hair regeneration.

Science.gov (United States)

Deng, Zhili; Lei, Xiaohua; Zhang, Xudong; Zhang, Huishan; Liu, Shuang; Chen, Qi; Hu, Huimin; Wang, Xinyue; Ning, Lina; Cao, Yujing; Zhao, Tongbiao; Zhou, Jiaxi; Chen, Ting; Duan, Enkui

2015-02-01

Hair follicles (HFs) undergo cycles of degeneration (catagen), rest (telogen), and regeneration (anagen) phases. Anagen begins when the hair follicle stem cells (HFSCs) obtain sufficient activation cues to overcome suppressive signals, mainly the BMP pathway, from their niche cells. Here, we unveil that mTOR complex 1 (mTORC1) signaling is activated in HFSCs, which coincides with the HFSC activation at the telogen-to-anagen transition. By using both an inducible conditional gene targeting strategy and a pharmacological inhibition method to ablate or inhibit mTOR signaling in adult skin epithelium before anagen initiation, we demonstrate that HFs that cannot respond to mTOR signaling display significantly delayed HFSC activation and extended telogen. Unexpectedly, BMP signaling activity is dramatically prolonged in mTOR signaling-deficient HFs. Through both gain- and loss-of-function studies in vitro, we show that mTORC1 signaling negatively affects BMP signaling, which serves as a main mechanism whereby mTORC1 signaling facilitates HFSC activation. Indeed, in vivo suppression of BMP by its antagonist Noggin rescues the HFSC activation defect in mTORC1-null skin. Our findings reveal a critical role for mTOR signaling in regulating stem cell activation through counterbalancing BMP-mediated repression during hair regeneration. © The Author (2015). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS. All rights reserved.

Signaling dynamics of palmitate-induced ER stress responses mediated by ATF4 in HepG2 cells

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Cho Hyunju

2013-01-01

Full Text Available Abstract Background Palmitic acid, the most common saturated free fatty acid, has been implicated in ER (endoplasmic reticulum stress-mediated apoptosis. This lipoapotosis is dependent, in part, on the upregulation of the activating transcription factor-4 (ATF4. To better understand the mechanisms by which palmitate upregulates the expression level of ATF4, we integrated literature information on palmitate-induced ER stress signaling into a discrete dynamic model. The model provides an in silico framework that enables simulations and predictions. The model predictions were confirmed through further experiments in human hepatocellular carcinoma (HepG2 cells and the results were used to update the model and our current understanding of the signaling induced by palmitate. Results The three key things from the in silico simulation and experimental results are: 1 palmitate induces different signaling pathways (PKR (double-stranded RNA-activated protein kinase, PERK (PKR-like ER kinase, PKA (cyclic AMP (cAMP-dependent protein kinase A in a time dependent-manner, 2 both ATF4 and CREB1 (cAMP-responsive element-binding protein 1 interact with the Atf4 promoter to contribute to a prolonged accumulation of ATF4, and 3 CREB1 is involved in ER-stress induced apoptosis upon palmitate treatment, by regulating ATF4 expression and possibly Ca2+ dependent-CaM (calmodulin signaling pathway. Conclusion The in silico model helped to delineate the essential signaling pathways in palmitate-mediated apoptosis.

Fibroblast growth factor receptor 4 regulates tumor invasion by coupling fibroblast growth factor signaling to extracellular matrix degradation

DEFF Research Database (Denmark)

Sugiyama, Nami; Varjosalo, Markku; Meller, Pipsa

2010-01-01

/stroma border and tumor invasion front. The strongest overall coexpression was found in prostate carcinoma. Studies with cultured prostate carcinoma cell lines showed that the FGFR4-R388 variant, which has previously been associated with poor cancer prognosis, increased MT1-MMP-dependent collagen invasion......Aberrant expression and polymorphism of fibroblast growth factor receptor 4 (FGFR4) has been linked to tumor progression and anticancer drug resistance. We describe here a novel mechanism of tumor progression by matrix degradation involving epithelial-to-mesenchymal transition in response...... to membrane-type 1 matrix metalloproteinase (MT1-MMP, MMP-14) induction at the edge of tumors expressing the FGFR4-R388 risk variant. Both FGFR4 and MT1-MMP were upregulated in tissue biopsies from several human cancer types including breast adenocarcinomas, where they were partially coexpressed at the tumor...

MAPKs are essential upstream signaling pathways in proteolytic cartilage degradation--divergence in pathways leading to aggrecanase and MMP-mediated articular cartilage degradation

DEFF Research Database (Denmark)

Sondergaard, B-C; Schultz, N; Madsen, S H

2010-01-01

Matrix metalloproteinases (MMPs) and aggrecanases are essential players in cartilage degradation. However, the signaling pathways that results in MMP and/or aggrecanase synthesis and activation are not well understood. We investigated the molecular events leading to MMP- and aggrecanase-mediated ......Matrix metalloproteinases (MMPs) and aggrecanases are essential players in cartilage degradation. However, the signaling pathways that results in MMP and/or aggrecanase synthesis and activation are not well understood. We investigated the molecular events leading to MMP- and aggrecanase......-mediated cartilage degradation....

Atorvastatin restores arsenic-induced vascular dysfunction in rats: Modulation of nitric oxide signaling and inflammatory mediators

International Nuclear Information System (INIS)

Kesavan, Manickam; Sarath, Thengumpallil Sasindran; Kannan, Kandasamy; Suresh, Subramaniyam; Gupta, Priyanka; Vijayakaran, Karunakaran; Sankar, Palanisamy; Kurade, Nitin Pandurang; Mishra, Santosh Kumar; Sarkar, Souvendra Nath

2014-01-01

We evaluated whether atorvastatin, an extensively prescribed statin for reducing the risks of cardiovascular diseases, can reduce the risk of arsenic-induced vascular dysfunction and inflammation in rats and whether the modulation could be linked to improvement in vascular NO signaling. Rats were exposed to sodium arsenite (100 ppm) through drinking water for 90 consecutive days. Atorvastatin (10 mg/kg bw, orally) was administered once daily during the last 30 days of arsenic exposure. On the 91 st day, blood was collected for measuring serum C-reactive protein. Thoracic aorta was isolated for assessing reactivity to phenylephrine, sodium nitroprusside and acetylcholine; evaluating eNOS and iNOS mRNA expression and measuring NO production, while abdominal aorta was used for ELISA of cytokines, chemokine and vascular cell adhesion molecules. Histopathology was done in aortic arches. Arsenic did not alter phenylephrine-elicited contraction. Atorvastatin inhibited E max of phenylephrine, but it augmented the contractile response in aortic rings from arsenic-exposed animals. Sodium nitroprusside-induced relaxation was not altered with any treatment. However, arsenic reduced acetylcholine-induced relaxation and affected aortic eNOS at the levels of mRNA expression, protein concentration, phosphorylation and NO production. Further, it increased aortic iNOS mRNA expression, iNOS-derived NO synthesis, production of pro-inflammatory mediators (IL-1β, IL-6, MCP-1, VCAM, sICAM) and serum C-reactive protein and aortic vasculopathic lesions. Atorvastatin attenuated these arsenic-mediated functional, biochemical and structural alterations. Results show that atorvastatin has the potential to ameliorate arsenic-induced vascular dysfunction and inflammation by restoring endothelial function with improvement in NO signaling and attenuating production of pro-inflammatory mediators and cell adhesion molecules. - Highlights: • We evaluated if atorvastatin reduce arsenic

Atorvastatin restores arsenic-induced vascular dysfunction in rats: Modulation of nitric oxide signaling and inflammatory mediators

Energy Technology Data Exchange (ETDEWEB)

Kesavan, Manickam; Sarath, Thengumpallil Sasindran; Kannan, Kandasamy; Suresh, Subramaniyam; Gupta, Priyanka; Vijayakaran, Karunakaran; Sankar, Palanisamy; Kurade, Nitin Pandurang; Mishra, Santosh Kumar; Sarkar, Souvendra Nath, E-mail: snsarkar1911@rediffmail.com

2014-10-01

We evaluated whether atorvastatin, an extensively prescribed statin for reducing the risks of cardiovascular diseases, can reduce the risk of arsenic-induced vascular dysfunction and inflammation in rats and whether the modulation could be linked to improvement in vascular NO signaling. Rats were exposed to sodium arsenite (100 ppm) through drinking water for 90 consecutive days. Atorvastatin (10 mg/kg bw, orally) was administered once daily during the last 30 days of arsenic exposure. On the 91{sup st} day, blood was collected for measuring serum C-reactive protein. Thoracic aorta was isolated for assessing reactivity to phenylephrine, sodium nitroprusside and acetylcholine; evaluating eNOS and iNOS mRNA expression and measuring NO production, while abdominal aorta was used for ELISA of cytokines, chemokine and vascular cell adhesion molecules. Histopathology was done in aortic arches. Arsenic did not alter phenylephrine-elicited contraction. Atorvastatin inhibited E{sub max} of phenylephrine, but it augmented the contractile response in aortic rings from arsenic-exposed animals. Sodium nitroprusside-induced relaxation was not altered with any treatment. However, arsenic reduced acetylcholine-induced relaxation and affected aortic eNOS at the levels of mRNA expression, protein concentration, phosphorylation and NO production. Further, it increased aortic iNOS mRNA expression, iNOS-derived NO synthesis, production of pro-inflammatory mediators (IL-1β, IL-6, MCP-1, VCAM, sICAM) and serum C-reactive protein and aortic vasculopathic lesions. Atorvastatin attenuated these arsenic-mediated functional, biochemical and structural alterations. Results show that atorvastatin has the potential to ameliorate arsenic-induced vascular dysfunction and inflammation by restoring endothelial function with improvement in NO signaling and attenuating production of pro-inflammatory mediators and cell adhesion molecules. - Highlights: • We evaluated if atorvastatin reduce arsenic

Linear collider signal of anomaly mediated supersymmetry breaking model

International Nuclear Information System (INIS)

Ghosh Dilip Kumar; Kundu, Anirban; Roy, Probir; Roy, Sourov

2001-01-01

Though the minimal model of anomaly mediated supersymmetry breaking has been significantly constrained by recent experimental and theoretical work, there are still allowed regions of the parameter space for moderate to large values of tan β. We show that these regions will be comprehensively probed in a √s = 1 TeV e + e - linear collider. Diagnostic signals to this end are studied by zeroing in on a unique and distinct feature of a large class of models in this genre: a neutral winolike Lightest Supersymmetric Particle closely degenerate in mass with a winolike chargino. The pair production processes e + e - → e tilde L ± e tilde L ± , e tilde R ± e tilde R ± , e tilde L ± e tilde R ± , ν tilde anti ν tilde, χ tilde 1 0 χ tilde 2 0 , χ tilde 2 0 χ tilde 2 0 are all considered at √s = 1 TeV corresponding to the proposed TESLA linear collider in two natural categories of mass ordering in the sparticle spectra. The signals analysed comprise multiple combinations of fast charged leptons (any of which can act as the trigger) plus displaced vertices X D (any of which can be identified by a heavy ionizing track terminating in the detector) and/or associated soft pions with characteristic momentum distributions. (author)

Regulators of G-protein signaling 4: modulation of 5-HT1A-mediated neurotransmitter release in vivo.

Science.gov (United States)

Beyer, Chad E; Ghavami, Afshin; Lin, Qian; Sung, Amy; Rhodes, Kenneth J; Dawson, Lee A; Schechter, Lee E; Young, Kathleen H

2004-10-01

Regulators of G-protein signaling (RGS) play a key role in the signal transduction of G-protein-coupled receptors (GPCRs). Specifically, RGS proteins function as GTPase accelerating proteins (GAPs) to dampen or "negatively regulate" GPCR-mediated signaling. Our group recently showed that RGS4 effectively GAPs Galpha(i)-mediated signaling in CHO cells expressing the serotonin-1A (5-HT(1A)) receptor. However, whether a similar relationship exists in vivo has yet to be identified. In present studies, a replication-deficient herpes simplex virus (HSV) was used to elevate RGS4 mRNA in the rat dorsal raphe nuclei (DRN) while extracellular levels of 5-HT in the striatum were monitored by in vivo microdialysis. Initial experiments conducted with noninfected rats showed that acute administration of 8-OH-DPAT (0.01-0.3 mg/kg, subcutaneous [s.c.]) dose dependently decreased striatal levels of 5-HT, an effect postulated to result from activation of somatodendritic 5-HT(1A) autoreceptors in the DRN. In control rats receiving a single intra-DRN infusion of HSV-LacZ, 8-OH-DPAT (0.03 mg/kg, s.c.) decreased 5-HT levels to an extent similar to that observed in noninfected animals. Conversely, rats infected with HSV-RGS4 in the DRN showed a blunted neurochemical response to 8-OH-DPAT (0.03 mg/kg, s.c.); however, increasing the dose to 0.3 mg/kg reversed this effect. Together, these findings represent the first in vivo evidence demonstrating that RGS4 functions to GAP Galpha(i)-coupled receptors and suggest that drug discovery efforts targeting RGS proteins may represent a novel mechanism to manipulate 5-HT(1A)-mediated neurotransmitter release.

Inhibition of fibroblast growth factor receptor with AZD4547 mitigates juvenile nasopharyngeal angiofibroma.

Science.gov (United States)

Le, Tran; New, Jacob; Jones, Joel W; Usman, Shireen; Yalamanchali, Sreeya; Tawfik, Ossama; Hoover, Larry; Bruegger, Dan E; Thomas, Sufi Mary

2017-10-01

Juvenile nasopharyngeal angiofibroma (JNA) is a benign tumor that presents in adolescent males. Although surgical excision is the mainstay of treatment, recurrences complicate treatment. There is a need to develop less invasive approaches for management. JNA tumors are composed of fibroblasts and vascular endothelial cells. We identified fibroblast growth factor receptor (FGFR) and vascular endothelial growth factor (VEGF) expression in JNA-derived fibroblasts. FGFR influences fibroblast proliferation and VEGF is necessary for angiogenesis. We hypothesized that targeting FGFR would mitigate JNA fibroblast proliferation, invasion, and migration, and that targeting the VEGF receptor would attenuate endothelial tubule formation. After informed consent, fibroblasts from JNA explants of 3 patients were isolated. Fibroblasts were treated with FGFR inhibitor AZD4547, 0 to 25 μg/mL for 72 hours and proliferation was quantified using CyQuant assay. Migration and invasion of JNA were assessed using 24-hour transwell assays with subsequent fixation and quantification. Mitigation of FGFR and downstream signaling was evaluated by immunoblotting. Tubule formation was assessed in human umbilical vein endothelial cells (HUVECs) treated with vehicle control (dimethylsulfoxide [DMSO]) or semaxanib (SU5416) as well as in serum-free media (SFM) or JNA conditioned media (CM). Tubule length was compared between treatment groups. Compared to control, AZD4547 inhibited JNA fibroblast proliferation, migration, and invasion through inhibition of FGFR and downstream signaling, specifically phosphorylation of - p44/42 mitogen activated protein kinase (p44/42 MAPK). JNA fibroblast CM significantly increased HUVEC tubule formation (p = 0.0039). AZD4547 effectively mitigates FGFR signaling and decreases JNA fibroblast proliferation, migration, and invasion. SU5416 attenuated JNA fibroblast-induced tubule formation. AZD4547 may have therapeutic potential in the treatment of JNA. © 2017 ARS

Achondroplasia: pathogenesis and implications for future treatment.

Science.gov (United States)

Laederich, Melanie B; Horton, William A

2010-08-01

Although the genetic defect underlying achondroplasia has been known for over a decade, no effective therapies to stimulate bone growth have emerged. Here we review the recent literature and summarize the molecular mechanisms underlying disease pathology and examine their potential as therapeutic targets. Currently used preclinical models are discussed in the context of recent advances with a special focus on C-type natriuretic peptide. Research on the mutation in Fibroblast Growth Factor Receptor 3 (FGFR3) that causes achondroplasia suggests that disease results from increased signal transduction from the mutant receptor. Thus, current therapeutic strategies have focused on reducing signals emanating from FGFR3. First-generation therapies directly targeting FGFR3, such as kinase inhibitors and neutralizing antibodies, designed for targeting FGFR3 in cancer, are still in the preclinical phase and have yet to translate into the management of achondroplasia. Counteracting signal transduction pathways downstream of FGFR3 holds promise with the discovery that administration of C-type natriuretic peptide to achondroplastic mice ameliorates their clinical phenotype. However, more research into long-term effectiveness and safety of this strategy is needed. Direct targeting of therapeutic agents to growth plate cartilage may enhance efficacy and minimize side effects of these and future therapies. Current research into the pathogenesis of achondroplasia has expanded our understanding of the mechanisms of FGFR3-induced disease and has increased the number of approaches that we may use to potentially correct it. Further research is needed to validate these approaches in preclinical models of achondroplasia.

Antitumor Effect of AZD4547 in a Fibroblast Growth Factor Receptor 2–Amplified Gastric Cancer Patient–Derived Cell Model

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Jiryeon Jang

2017-08-01

Full Text Available BACKGROUND: FGFR2 amplification is associated with aggressive gastric cancer (GC, and targeted drugs have been developed for treatment of GC. We evaluated the antitumor activity of an FGFR inhibitor in FGFR2-amplified GC patients with peritoneal carcinomatosis. METHODS: Two GC patients with FGFR2 amplification confirmed by fluorescence in situ hybridization showed peritoneal seeding and malignant ascites. We used the patient-derived xenograft model; patient-derived cells (PDCs from malignant ascites were used to assess FGFR2 expression and its downstream pathway using immunofluorescence analysis and immunoblot assay in vitro. Apoptosis and cell cycle arrest after treatment of FGFR inhibitor were analyzed by Annexin V-FITC assay and cell cycle analysis. RESULTS: FGFR2 amplification was verified in both PDC lines. AZD4547 as an FGFR inhibitor decreased proliferation of PDCs, and the IC50 value was estimated to be 250 nM in PDC#1 and 210 nM in PDC#2. FGFR inhibitor also significantly decreased levels of phosphorylated FGFR2 and downstream signaling molecules in FGFR2-amplified PDC lines. In cell cycle analysis, apoptosis was significantly increased in AZD4547-treated cells compared with nontreated cells. The proportion of cells in the sub-G1 stage was significantly higher in AZD4547-treated PDCs than in control cells. CONCLUSION: Our findings suggest that FGFR2 amplification is a relevant therapeutic target in GC with peritoneal carcinomatosis.

p.Ser252Trp and p.Pro253Arg mutations in FGFR2 gene causing Apert syndrome: the first clinical and molecular report of Indonesian patients.

Science.gov (United States)

Mundhofir, Farmaditya E P; Sistermans, Erik A; Faradz, Sultana M H; Hamel, Ben C J

2013-03-01

Apert syndrome (AS) is a rare autosomal dominant disorder characterised by craniosynostosis and limb malformations, and is associated with congenital heart disease and other systemic malformations, including intellectual disability. We report two Indonesian patients with AS, in whom molecular analysis detected p.Ser252Trp (c.755C>G) and p.Pro253Arg (c.758C>G) mutations in the fibroblast growth factor receptor 2 (FGFR2) gene, respectively. Although the syndrome has been frequently described, this is the first clinical report of AS confirmed by molecular analysis in Indonesia. The difference in severity of clinical features in the two patients may be consistent with a genotype-phenotype correlation of the FGFR2mutation. The management of individuals with AS is best achieved within a multidisciplinary setting. However, in most developing countries, early intervention may be delayed due to late diagnosis, a lack of facilities and financial constraints. This report underpins the benefits of early diagnosis for AS management.

Intracellular Signaling Mediators in the Circulatory and Ventilatory Systems

CERN Document Server

Thiriet, Marc

2013-01-01

The volumes in this authoritative series present a multidisciplinary approach to modeling and simulation of flows in the cardiovascular and ventilatory systems, especially multiscale modeling and coupled simulations. The cardiovascular and respiratory systems are tightly coupled, as their primary function is to supply oxygen to and remove carbon dioxide from the body's cells. Because physiological conduits have deformable and reactive walls, macroscopic flow behavior and prediction must be coupled to phenomenological models of nano- and microscopic events in a corrector scheme of regulated mechanisms when the vessel lumen caliber varies markedly. Therefore, investigation of flows of blood and air in physiological conduits requires an understanding of the biology, chemistry, and physics of these systems together with the mathematical tools to describe their functioning. Volume 4 is devoted to major sets of intracellular mediators that transmit signals upon stimulation of cell-surface receptors.  Activation of...

IL1β-mediated Stromal COX-2 signaling mediates proliferation and invasiveness of colonic epithelial cancer cells

International Nuclear Information System (INIS)

Zhu, Yingting; Zhu, Min; Lance, Peter

2012-01-01

COX-2 is a major inflammatory mediator implicated in colorectal inflammation and cancer. However, the exact origin and role of COX-2 on colorectal inflammation and carcinogenesis are still not well defined. Recently, we reported that COX-2 and iNOS signalings interact in colonic CCD18Co fibroblasts. In this article, we investigated whether activation of COX-2 signaling by IL1β in primary colonic fibroblasts obtained from normal and cancer patients play a critical role in regulation of proliferation and invasiveness of human colonic epithelial cancer cells. Our results demonstrated that COX-2 level was significantly higher in cancer associated fibroblasts than that in normal fibroblasts with or without stimulation of IL-1β, a powerful stimulator of COX-2. Using in vitro assays for estimating proliferative and invasive potential, we discovered that the proliferation and invasiveness of the epithelial cancer cells were much greater when the cells were co-cultured with cancer associated fibroblasts than with normal fibroblasts, with or without stimulation of IL1β. Further analysis indicated that the major COX-2 product, prostaglandin E 2 , directly enhanced proliferation and invasiveness of the epithelial cancer cells in the absence of fibroblasts. Moreover, a selective COX-2 inhibitor, NS-398, blocked the proliferative and invasive effect of both normal and cancer associate fibroblasts on the epithelial cancer cells, with or without stimulation of IL-1β. Those results indicate that activation of COX-2 signaling in the fibroblasts plays a major role in promoting proliferation and invasiveness of the epithelial cancer cells. In this process, PKC is involved in the activation of COX-2 signaling induced by IL-1β in the fibroblasts.

A bipartite signal mediates the transfer of type IV secretion substrates of Bartonella henselae into human cells.

Science.gov (United States)

Schulein, Ralf; Guye, Patrick; Rhomberg, Thomas A; Schmid, Michael C; Schröder, Gunnar; Vergunst, Annette C; Carena, Ilaria; Dehio, Christoph

2005-01-18

Bacterial type IV secretion (T4S) systems mediate the transfer of macromolecular substrates into various target cells, e.g., the conjugative transfer of DNA into bacteria or the transfer of virulence proteins into eukaryotic host cells. The T4S apparatus VirB of the vascular tumor-inducing pathogen Bartonella henselae causes subversion of human endothelial cell (HEC) function. Here we report the identification of multiple protein substrates of VirB, which, upon translocation into HEC, mediate all known VirB-dependent cellular changes. These Bartonella-translocated effector proteins (Beps) A-G are encoded together with the VirB system and the T4S coupling protein VirD4 on a Bartonella-specific pathogenicity island. The Beps display a modular architecture, suggesting an evolution by extensive domain duplication and reshuffling. The C terminus of each Bep harbors at least one copy of the Bep-intracellular delivery domain and a short positively charged tail sequence. This biparte C terminus constitutes a transfer signal that is sufficient to mediate VirB/VirD4-dependent intracellular delivery of reporter protein fusions. The Bep-intracellular delivery domain is also present in conjugative relaxases of bacterial conjugation systems. We exemplarily show that the C terminus of such a conjugative relaxase mediates protein transfer through the Bartonella henselae VirB/VirD4 system into HEC. Conjugative relaxases may thus represent the evolutionary origin of the here defined T4S signal for protein transfer into human cells.

Mitochondria mediate tumor necrosis factor-alpha/NF-kappaB signaling in skeletal muscle myotubes

Science.gov (United States)

Li, Y. P.; Atkins, C. M.; Sweatt, J. D.; Reid, M. B.; Hamilton, S. L. (Principal Investigator)

1999-01-01

Tumor necrosis factor-alpha (TNF-alpha) is implicated in muscle atrophy and weakness associated with a variety of chronic diseases. Recently, we reported that TNF-alpha directly induces muscle protein degradation in differentiated skeletal muscle myotubes, where it rapidly activates nuclear factor kappaB (NF-kappaB). We also have found that protein loss induced by TNF-alpha is NF-kappaB dependent. In the present study, we analyzed the signaling pathway by which TNF-alpha activates NF-kappaB in myotubes differentiated from C2C12 and rat primary myoblasts. We found that activation of NF-kappaB by TNF-alpha was blocked by rotenone or amytal, inhibitors of complex I of the mitochondrial respiratory chain. On the other hand, antimycin A, an inhibitor of complex III, enhanced TNF-alpha activation of NK-kappaB. These results suggest a key role of mitochondria-derived reactive oxygen species (ROS) in mediating NF-kappaB activation in muscle. In addition, we found that TNF-alpha stimulated protein kinase C (PKC) activity. However, other signal transduction mediators including ceramide, Ca2+, phospholipase A2 (PLA2), and nitric oxide (NO) do not appear to be involved in the activation of NF-kappaB.

Multilepton signals of gauge mediated supersymmetry breaking at the LHC

Energy Technology Data Exchange (ETDEWEB)

D' Hondt, Jorgen [Theoretische Natuurkunde and IIHE/ELEM, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels (Belgium); International Solvay Institutes, Brussels (Belgium); De Causmaecker, Karen [Theoretische Natuurkunde and IIHE/ELEM, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels (Belgium); International Solvay Institutes, Brussels (Belgium); Theory Division, Physics Department, CERN, CH-1211 Geneva 23 (Switzerland); Fuks, Benjamin [Theory Division, Physics Department, CERN, CH-1211 Geneva 23 (Switzerland); Institut Pluridisciplinaire Hubert Curien/Département Recherches Subatomiques, Université de Strasbourg/CNRS-IN2P3, 23 Rue du Loess, F-67037 Strasbourg (France); Mariotti, Alberto [Institute for Particle Physics Phenomenology, Durham University, Durham DH1 3LE (United Kingdom); Mawatari, Kentarou [Theoretische Natuurkunde and IIHE/ELEM, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels (Belgium); International Solvay Institutes, Brussels (Belgium); Petersson, Christoffer [International Solvay Institutes, Brussels (Belgium); Physique Théorique et Mathématique, Université Libre de Bruxelles, C.P. 231, B-1050 Brussels (Belgium); Department of Fundamental Physics, Chalmers University of Technology, 412 96 Göteborg (Sweden); Redigolo, Diego [International Solvay Institutes, Brussels (Belgium); Physique Théorique et Mathématique, Université Libre de Bruxelles, C.P. 231, B-1050 Brussels (Belgium)

2014-04-04

We investigate multilepton LHC signals arising from electroweak processes involving sleptons. We consider the framework of general gauge mediated supersymmetry breaking, focusing on models where the low mass region of the superpartner spectrum consists of the three generations of charged sleptons and the nearly massless gravitino. We demonstrate how such models can provide an explanation for the anomalous four lepton events recently observed by the CMS Collaboration, while satisfying other existing experimental constraints. The best fit to the CMS data is obtained for a selectron/smuon mass of around 145 GeV and a stau mass of around 90 GeV. These models also give rise to final states with more than four leptons, offering alternative channels in which they can be probed and we estimate the corresponding production rates at the LHC.

Promoter hypermethylation of HS3ST2, SEPTIN9 and SLIT2 combined with FGFR3 mutations as a sensitive/specific urinary assay for diagnosis and surveillance in patients with low or high-risk non-muscle-invasive bladder cancer

KAUST Repository

Roperch, Jean-Pierre

2016-09-02

Background: Non-muscle-invasive bladder cancer (NMIBC) is a high incidence form of bladder cancer (BCa), where genetic and epigenetic alterations occur frequently. We assessed the performance of associating a FGFR3 mutation assay and a DNA methylation analysis to improve bladder cancer detection and to predict disease recurrence of NMIBC patients. Methods: We used allele specific PCR to determine the FGFR3 mutation status for R248C, S249C, G372C, and Y375C. We preselected 18 candidate genes reported in the literature as being hypermethylated in cancer and measured their methylation levels by quantitative multiplex-methylation specific PCR. We selected HS3ST2, SLIT2 and SEPTIN9 as the most discriminative between control and NMIBC patients and we assayed these markers on urine DNA from a diagnostic study consisting of 167 NMIBC and 105 controls and a follow-up study consisting of 158 NMIBC at diagnosis time\\'s and 425 at follow-up time. ROC analysis was performed to evaluate the diagnostic accuracy of each assay alone and in combination. Results: For Diagnosis: Using a logistic regression analysis with a model consisting of the 3 markers\\' methylation values, FGFR3 status, age and known smoker status at the diagnosis time we obtained sensitivity/specificity of 97.6 %/84.8 % and an optimism-corrected AUC of 0.96. With an estimated BCa prevalence of 12.1 % in a hematuria cohort, this corresponds to a negative predictive value (NPV) of 99.6 %. For Follow-up: Using a logistic regression with FGFR3 mutation and the CMI at two time points (beginning of the follow-up and current time point), we got sensitivity/specificity/NPV of 90.3 %/65.1 %/97.0 % and a corrected AUC of 0.84. We also tested a thresholding algorithm with FGFR3 mutation and the two time points as described above, obtaining sensitivity/specificity/NPV values of, respectively, 94.5 %/75.9 %/98.5 % and an AUC of 0.82. Conclusions: We showed that combined analysis of FGFR3 mutation and DNA methylation markers

Promoter hypermethylation of HS3ST2, SEPTIN9 and SLIT2 combined with FGFR3 mutations as a sensitive/specific urinary assay for diagnosis and surveillance in patients with low or high-risk non-muscle-invasive bladder cancer

KAUST Repository

Roperch, Jean-Pierre; Grandchamp, Bernard; Desgrandchamps, Franç ois; Mongiat-Artus, Pierre; Ravery, Vincent; Ouzaid, Idir; Roupret, Morgan; Phe, Vé ronique; Ciofu, Calin; Tubach, Florence; Cussenot, Olivier; Incitti, Roberto

2016-01-01

Background: Non-muscle-invasive bladder cancer (NMIBC) is a high incidence form of bladder cancer (BCa), where genetic and epigenetic alterations occur frequently. We assessed the performance of associating a FGFR3 mutation assay and a DNA methylation analysis to improve bladder cancer detection and to predict disease recurrence of NMIBC patients. Methods: We used allele specific PCR to determine the FGFR3 mutation status for R248C, S249C, G372C, and Y375C. We preselected 18 candidate genes reported in the literature as being hypermethylated in cancer and measured their methylation levels by quantitative multiplex-methylation specific PCR. We selected HS3ST2, SLIT2 and SEPTIN9 as the most discriminative between control and NMIBC patients and we assayed these markers on urine DNA from a diagnostic study consisting of 167 NMIBC and 105 controls and a follow-up study consisting of 158 NMIBC at diagnosis time's and 425 at follow-up time. ROC analysis was performed to evaluate the diagnostic accuracy of each assay alone and in combination. Results: For Diagnosis: Using a logistic regression analysis with a model consisting of the 3 markers' methylation values, FGFR3 status, age and known smoker status at the diagnosis time we obtained sensitivity/specificity of 97.6 %/84.8 % and an optimism-corrected AUC of 0.96. With an estimated BCa prevalence of 12.1 % in a hematuria cohort, this corresponds to a negative predictive value (NPV) of 99.6 %. For Follow-up: Using a logistic regression with FGFR3 mutation and the CMI at two time points (beginning of the follow-up and current time point), we got sensitivity/specificity/NPV of 90.3 %/65.1 %/97.0 % and a corrected AUC of 0.84. We also tested a thresholding algorithm with FGFR3 mutation and the two time points as described above, obtaining sensitivity/specificity/NPV values of, respectively, 94.5 %/75.9 %/98.5 % and an AUC of 0.82. Conclusions: We showed that combined analysis of FGFR3 mutation and DNA methylation markers on

Testin, a novel binding partner of the calcium-sensing receptor, enhances receptor-mediated Rho-kinase signalling

International Nuclear Information System (INIS)

Magno, Aaron L.; Ingley, Evan; Brown, Suzanne J.; Conigrave, Arthur D.; Ratajczak, Thomas; Ward, Bryan K.

2011-01-01

Highlights: → A yeast two-hybrid screen revealed testin bound to the calcium-sensing receptor. → The second zinc finger of LIM domain 1 of testin is critical for interaction. → Testin bound to a region of the receptor tail important for cell signalling. → Testin and receptor interaction was confirmed in mammalian (HEK293) cells. → Overexpression of testin enhanced receptor-mediated Rho signalling in HEK293 cells. -- Abstract: The calcium-sensing receptor (CaR) plays an integral role in calcium homeostasis and the regulation of other cellular functions including cell proliferation and cytoskeletal organisation. The multifunctional nature of the CaR is manifested through ligand-dependent stimulation of different signalling pathways that are also regulated by partner binding proteins. Following a yeast two-hybrid library screen using the intracellular tail of the CaR as bait, we identified several novel binding partners including the focal adhesion protein, testin. Testin has not previously been shown to interact with cell surface receptors. The sites of interaction between the CaR and testin were mapped to the membrane proximal region of the receptor tail and the second zinc-finger of LIM domain 1 of testin, the integrity of which was found to be critical for the CaR-testin interaction. The CaR-testin association was confirmed in HEK293 cells by coimmunoprecipitation and confocal microscopy studies. Ectopic expression of testin in HEK293 cells stably expressing the CaR enhanced CaR-stimulated Rho activity but had no effect on CaR-stimulated ERK signalling. These results suggest an interplay between the CaR and testin in the regulation of CaR-mediated Rho signalling with possible effects on the cytoskeleton.

P2X receptor-mediated ATP purinergic signaling in health and disease

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Jiang LH

2012-09-01

Full Text Available Lin-Hua JiangSchool of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, Leeds, United KingdomAbstract: Purinergic P2X receptors are plasma membrane proteins present in a wide range of mammalian cells where they act as a cellular sensor, enabling cells to detect and respond to extracellular adenosine triphosphate (ATP, an important signaling molecule. P2X receptors function as ligand-gated Ca2+-permeable cationic channels that open upon ATP binding to elevate intracellular Ca2+ concentrations and cause membrane depolarization. In response to sustained activation, P2X receptors induce formation of a pore permeable to large molecules. P2X receptors also interact with distinct functional proteins and membrane lipids to form specialized signaling complexes. Studies have provided compelling evidence to show that such P2X receptor-mediated ATP-signaling mechanisms determine and regulate a growing number and diversity of important physiological processes, including neurotransmission, muscle contraction, and cytokine release. There is accumulating evidence to support strong causative relationships of altered receptor expression and function with chronic pain, inflammatory diseases, cancers, and other pathologies or diseases. Numerous high throughput screening drug discovery programs and preclinical studies have thus far demonstrated the proof of concepts that the P2X receptors are druggable targets and selective receptor antagonism is a promising therapeutics approach. This review will discuss the recent progress in understanding the mammalian P2X receptors with respect to the ATP-signaling mechanisms, physiological and pathophysiological roles, and development and preclinical studies of receptor antagonists.Keywords: extracellular ATP, ion channel, large pore, signaling complex, chronic pain, inflammatory diseases

Reactive Oxygen Species-Mediated Loss of Synaptic Akt1 Signaling Leads to Deficient Activity-Dependent Protein Translation Early in Alzheimer's Disease.

Science.gov (United States)

Ahmad, Faraz; Singh, Kunal; Das, Debajyoti; Gowaikar, Ruturaj; Shaw, Eisha; Ramachandran, Arathy; Rupanagudi, Khader Valli; Kommaddi, Reddy Peera; Bennett, David A; Ravindranath, Vijayalakshmi

2017-12-01

Synaptic deficits are known to underlie the cognitive dysfunction seen in Alzheimer's disease (AD). Generation of reactive oxygen species (ROS) by β-amyloid has also been implicated in AD pathogenesis. However, it is unclear whether ROS contributes to synaptic dysfunction seen in AD pathogenesis and, therefore, we examined whether altered redox signaling could contribute to synaptic deficits in AD. Activity dependent but not basal translation was impaired in synaptoneurosomes from 1-month old presymptomatic APP Swe /PS1ΔE9 (APP/PS1) mice, and this deficit was sustained till middle age (MA, 9-10 months). ROS generation leads to oxidative modification of Akt1 in the synapse and consequent reduction in Akt1-mechanistic target of rapamycin (mTOR) signaling, leading to deficiency in activity-dependent protein translation. Moreover, we found a similar loss of activity-dependent protein translation in synaptoneurosomes from postmortem AD brains. Loss of activity-dependent protein translation occurs presymptomatically early in the pathogenesis of AD. This is caused by ROS-mediated loss of pAkt1, leading to reduced synaptic Akt1-mTOR signaling and is rescued by overexpression of Akt1. ROS-mediated damage is restricted to the synaptosomes, indicating selectivity. We demonstrate that ROS-mediated oxidative modification of Akt1 contributes to synaptic dysfunction in AD, seen as loss of activity-dependent protein translation that is essential for synaptic plasticity and maintenance. Therapeutic strategies promoting Akt1-mTOR signaling at synapses may provide novel target(s) for disease-modifying therapy in AD. Antioxid. Redox Signal. 27, 1269-1280.

SOCS2 mediates the cross talk between androgen and growth hormone signaling in prostate cancer

DEFF Research Database (Denmark)

Iglesias Gato, Diego; Chuan, Yin Choy; Wikström, Pernilla

2014-01-01

) as mediator of the cross talk between androgens and GH signals in the prostate and its potential role as tumor suppressor in prostate cancer (PCa). We observed that SOCS2 protein levels assayed by immunohistochemistry are elevated in hormone therapy-naive localized prostatic adenocarcinoma in comparison...... of transcription 5 protein (STAT5) and androgen receptor-dependent transcription. Consequentially, SOCS2 inhibits GH activation of Janus kinase 2, Src and STAT5 as well as both cell invasion and cell proliferation in vitro. In vivo, SOCS2 limits proliferation and production of IGF-1 in the prostate in response......Anabolic signals such as androgens and the growth hormone/insulin-like growth factor 1 (GH/IGF-1) axis play an essential role in the normal development of the prostate but also in its malignant transformation. In this study, we investigated the role of suppressor of cytokine signaling 2 (SOCS2...

Robo signaling regulates the production of cranial neural crest cells.

Science.gov (United States)

Li, Yan; Zhang, Xiao-Tan; Wang, Xiao-Yu; Wang, Guang; Chuai, Manli; Münsterberg, Andrea; Yang, Xuesong

2017-12-01

Slit/Robo signaling plays an important role in the guidance of developing neurons in developing embryos. However, it remains obscure whether and how Slit/Robo signaling is involved in the production of cranial neural crest cells. In this study, we examined Robo1 deficient mice to reveal developmental defects of mouse cranial frontal and parietal bones, which are derivatives of cranial neural crest cells. Therefore, we determined the production of HNK1 + cranial neural crest cells in early chick embryo development after knock-down (KD) of Robo1 expression. Detection of markers for pre-migratory and migratory neural crest cells, PAX7 and AP-2α, showed that production of both was affected by Robo1 KD. In addition, we found that the transcription factor slug is responsible for the aberrant delamination/EMT of cranial neural crest cells induced by Robo1 KD, which also led to elevated expression of E- and N-Cadherin. N-Cadherin expression was enhanced when blocking FGF signaling with dominant-negative FGFR1 in half of the neural tube. Taken together, we show that Slit/Robo signaling influences the delamination/EMT of cranial neural crest cells, which is required for cranial bone development. Copyright © 2017. Published by Elsevier Inc.

miR-148a-3p Mediates Notch Signaling to Promote the Differentiation and M1 Activation of Macrophages

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Fei Huang

2017-10-01

Full Text Available The Notch pathway plays critical roles in the differentiation and polarized activation of macrophages; however, the downstream molecular mechanisms underlying Notch activity in macrophages remain elusive. Our previous study has identified a group of microRNAs that mediate Notch signaling to regulate macrophage activation and tumor-associated macrophages (TAMs. In this study, we demonstrated that miR-148a-3p functions as a novel downstream molecule of Notch signaling to promote the differentiation of monocytes into macrophages in the presence of granulocyte macrophage colony-stimulating factor (GM-CSF. Meanwhile, miR-148a-3p promoted M1 and inhibited M2 polarization of macrophages upon Notch activation. Macrophages overexpressing miR-148a-3p exhibited enhanced ability to engulf and kill bacteria, which was mediated by excessive production of reactive oxygen species (ROS. Further studies using reporter assay and Western blotting identified Pten as a direct target gene of miR-148a-3p in macrophages. Macrophages overexpressing miR-148a-3p increased their ROS production through the PTEN/AKT pathway, likely to defend against bacterial invasion. Moreover, miR-148a-3p also enhanced M1 macrophage polarization and pro-inflammatory responses through PTEN/AKT-mediated upregulation of NF-κB signaling. In summary, our data establish a novel molecular mechanism by which Notch signaling promotes monocyte differentiation and M1 macrophage activation through miR-148a-3p, and suggest that miR-148a-3p-modified monocytes or macrophages are potential new tools for the treatment of inflammation-related diseases.

The E3 ubiquitin ligase NEDD4 mediates cell migration signaling of EGFR in lung cancer cells.

Science.gov (United States)

Shao, Genbao; Wang, Ranran; Sun, Aiqin; Wei, Jing; Peng, Ke; Dai, Qian; Yang, Wannian; Lin, Qiong

2018-02-19

EGFR-dependent cell migration plays an important role in lung cancer progression. Our previous study observed that the HECT E3 ubiquitin ligase NEDD4 is significantly correlated with tumor metastasis and required for migration and invasion signaling of EGFR in gastric cancer cells. However, how NEDD4 promotes the EGFR-dependent lung cancer cell migration is unknown. This study is to elucidate the mechanism by which NEDD4 mediates the EGFR lung cancer migration signaling. Lentiviral vector-loaded NEDD4 shRNA was used to deplete endogenous NEDD4 in lung cancer cell lines. Effects of the NEDD4 knockdown on the EGFR-dependent or independent lung cancer cell migration were determined using the wound-healing and transwell assays. Association of NEDD4 with activated EGFR was assayed by co-immunoprecipitation. Co-expression of NEDD4 with EGFR or PTEN was determined by immunohistochemical (IHC) staining in 63 lung adenocarcinoma tissue samples. Effects of NEDD4 ectopic expression or knockdown on PTEN ubiquitination and down-regulation, AKT activation and lysosomal secretion were examined using the GST-Uba pulldown assay, immunoblotting, immunofluorescent staining and a human cathepsin B ELISA assay respectively. The specific cathepsin B inhibitor CA-074Me was used for assessing the role of cathepsin B in lung cancer cell migration. Knockdown of NEDD4 significantly reduced EGF-stimulated cell migration in non-small cell lung carcinoma (NSCLC) cells. Co-immunoprecipitation assay found that NEDD4 is associated with EGFR complex upon EGF stimulation, and IHC staining indicates that NEDD4 is co-expressed with EGFR in lung adenocarcinoma tumor tissues, suggesting that NEDD4 might mediate lung cancer cell migration by interaction with the EGFR signaling complex. Interestingly, NEDD4 promotes the EGF-induced cathepsin B secretion, possibly through lysosomal exocytosis, as overexpression of the ligase-dead mutant of NEDD4 impedes lysosomal secretion, and knockdown of NEDD4

PINK1 positively regulates IL-1β-mediated signaling through Tollip and IRAK1 modulation

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Lee Hyun Jung

2012-12-01

Full Text Available Abstract Background Parkinson disease (PD is characterized by a slow, progressive degeneration of dopaminergic neurons in the substantianigra. The cause of neuronal loss in PD is not well understood, but several genetic loci, including PTEN-induced putative kinase 1 (PINK1, have been linked to early-onset autosomal recessive forms of familial PD. Neuroinflammation greatly contributes to PD neuronal degeneration and pathogenesis. IL-1 is one of the principal cytokines that regulates various immune and inflammatory responses via the activation of the transcription factors NF-κB and activating protein-1. Despite the close relationship between PD and neuroinflammation, the functional roles of PD-linked genes during inflammatory processes remain poorly understood. Methods To explore the functional roles of PINK1 in response to IL-1β stimulation, HEK293 cells, mouse embryonic fibroblasts derived from PINK1-null (PINK1−/− and control (PINK1+/+ mice, and 293 IL-1RI cells stably expressing type 1 IL-1 receptor were used. Immunoprecipitation and western blot analysis were performed to detect protein–protein interaction and protein ubiquitination. To confirm the effect of PINK1 on NF-κB activation, NF-κB-dependent firefly luciferase reporter assay was conducted. Results PINK1 specifically binds two components of the IL-1-mediated signaling cascade, Toll-interacting protein (Tollip and IL-1 receptor-associated kinase 1 (IRAK1. The association of PINK1 with Tollip, a negative regulator of IL-1β signaling, increases upon IL-1β stimulation, which then facilitates the dissociation of Tollip from IRAK1 as well as the assembly of the IRAK1–TNF receptor-associated factor 6 (TRAF6 complex. PINK1 also enhances Lys63-linked polyubiquitination of IRAK1, an essential modification of recruitment of NF-κB essential modulator and subsequent IκB kinase activation, and increases formation of the intermediate signalosome including IRAK1, TRAF6, and

GPCR-Mediated Signaling of Metabolites

DEFF Research Database (Denmark)

Husted, Anna Sofie; Trauelsen, Mette; Rudenko, Olga

2017-01-01

microbiota target primarily enteroendocrine, neuronal, and immune cells in the lamina propria of the gut mucosa and the liver and, through these tissues, the rest of the body. In contrast, metabolites from the intermediary metabolism act mainly as metabolic stress-induced autocrine and paracrine signals...... and obesity. The concept of key metabolites as ligands for specific GPCRs has broadened our understanding of metabolic signaling significantly and provides a number of novel potential drug targets....

Deficiency in LRP6-Mediated Wnt Signaling Contributes to Synaptic Abnormalities and Amyloid Pathology in Alzheimer’s Disease

OpenAIRE

Liu, Chia-Chen; Tsai, Chih-Wei; Deak, Ferenc; Rogers, Justin; Penuliar, Michael; Sung, You Me; Maher, James N.; Fu, Yuan; Li, Xia; Xu, Huaxi; Estus, Steven; Hoe, Hyang-Sook; Fryer, John D.; Kanekiyo, Takahisa; Bu, Guojun

2014-01-01

Alzheimer’s disease (AD) is an age-related neurological disorder characterized by synaptic loss and dementia. The low-density lipoprotein receptor-related protein 6 (LRP6) is an essential co-receptor for Wnt signaling and its genetic variants have been linked to AD risk. Here we report that neuronal LRP6-mediated Wnt signaling is critical for synaptic function and cognition. Conditional deletion of Lrp6 gene in mouse forebrain neurons leads to age-dependent deficits in synaptic integrity and ...

Nitric oxide agents impair insulin-mediated signal transduction in rat skeletal muscle

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Ragoobirsingh Dalip

2006-05-01

Full Text Available Abstract Background Evidence demonstrates that exogenously administered nitric oxide (NO can induce insulin resistance in skeletal muscle. We have investigated the modulatory effects of two NO donors, S-nitroso-N-acetyl-D, L-penicillamine (SNAP and S-nitrosoglutathione (GSNO on the early events in insulin signaling in rat skeletal myocytes. Results Skeletal muscle cells from 6–8 week old Sprague-Dawley rats were treated with SNAP or GSNO (25 ng/ml in the presence or absence of glucose (25 mM and insulin (100 nM. Cellular insulin receptor-β levels and tyrosine phosphorylation in IRS-1 were significantly reduced, while serine phosphorylation in IRS-1 was significantly increased in these cells, when compared to the insulin-stimulated control. Reversal to near normal levels was achieved using the NO scavenger, 2-(4-carboxyphenyl-4, 4, 5, 5-tetramethylimidazoline-1-oxyl 3-oxide (carboxy-PTIO. Conclusion These data suggest that NO is a potent modulator of insulin-mediated signal transduction and may play a significant role in the pathogenesis of type 2 diabetes mellitus.

Apert Syndrome: Molecularly Confirmed C.758C>G (P.Pro253Arg) in FGFR2

Energy Technology Data Exchange (ETDEWEB)

Cha Gon, Lee, E-mail: leechagon@eulji.ac.kr [Department of Pediatrics, Eulji General Hospital, College of Medicine, Eulji University, 68 Hangeulbiseok-ro, Nowon-gu, Seoul 139-711 (Korea, Republic of)

2016-03-21

A 5-day-old girl was referred to our clinic for evaluation of congenital malformations. She was identified with a pathogenic mutation c.758C>G (p.Pro253Arg) in FGFR2 gene using targeted exome sequencing. The de novo mutation was confirmed with Sanger sequencing in the patient and her parents. She showed occipital plagiocephaly with frontal bossing (Figure A and B). Skull frontal and lateral radiography revealed fusion of most of the sutures except coronal suture, with convolutional markings (Figure D and E). She had complete cleft palate (Figure C). Her fused bilateral hands showed type II syndactyly with complete syndactyly between the ring and the little fingers (Figure F1-F3). Both toes were simple syndactyly with side-to-side fusion of skin (Figure G1-)

Apert Syndrome: Molecularly Confirmed C.758C>G (P.Pro253Arg) in FGFR2

International Nuclear Information System (INIS)

Cha Gon, Lee

2016-01-01

A 5-day-old girl was referred to our clinic for evaluation of congenital malformations. She was identified with a pathogenic mutation c.758C>G (p.Pro253Arg) in FGFR2 gene using targeted exome sequencing. The de novo mutation was confirmed with Sanger sequencing in the patient and her parents. She showed occipital plagiocephaly with frontal bossing (Figure A and B). Skull frontal and lateral radiography revealed fusion of most of the sutures except coronal suture, with convolutional markings (Figure D and E). She had complete cleft palate (Figure C). Her fused bilateral hands showed type II syndactyly with complete syndactyly between the ring and the little fingers (Figure F1-F3). Both toes were simple syndactyly with side-to-side fusion of skin (Figure G1-)

Central mechanisms mediating the hypophagic effects of oleoylethanolamide and N-acylphosphatidylethanolamines: different lipid signals?

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Adele eRomano

2015-06-01

Full Text Available The spread of ‘obesity epidemic’ and the poor efficacy of many anti-obesity therapies in the long-term highlight the need to develop novel efficacious therapy. This necessity stimulates a large research effort to find novel mechanisms controlling feeding and energy balance. Among these mechanisms a great deal of attention has been attracted by a family of phospholipid-derived signaling molecules that play an important role in the regulation of food-intake. They include N-acylethanolamines (NAEs and N-acylphosphatidylethanolamines (NAPEs. NAPEs have been considered for a long time simply as phospholipid precursors of the lipid mediator NAEs, but increasing body of evidence suggest a role in many physiological processes including the regulation of feeding behavior. Several observations demonstrated that among NAEs, oleoylethanolamide (OEA acts as a satiety signal, which is generated in the intestine, upon the ingestion of fat, and signals to the central nervous system. At this level different neuronal pathways, including oxytocinergic, noradrenergic, and histaminergic neurons, seem to mediate its hypophagic action. Similarly to NAEs, NAPEs (with particular reference to the N16:0 species levels were shown to be regulated by the fed state and this finding was initially interpreted as fluctuations of NAE precursors. However, the observation that exogenously administered NAPEs are able to inhibit food intake, not only in normal rats and mice but also in mice lacking the enzyme that converts NAPEs into NAEs, supported the hypothesis of a role of NAPE in the regulation of feeding behavior. Indirect observations suggest that the hypophagic action of NAPEs might involve central mechanisms, although the molecular target remains unknown. The present paper reviews the role that OEA and NAPEs play in the mechanisms that control food intake, further supporting this group of phospholipids as optimal candidate for the development of novel anti

Specific blockade by CD54 and MHC II of CD40-mediated signaling for B cell proliferation and survival

DEFF Research Database (Denmark)

Doyle, I S; Hollmann, C A; Crispe, I N

2001-01-01

Regulation of B lymphocyte proliferation is critical to maintenance of self-tolerance, and intercellular interactions are likely to signal such regulation. Here, we show that coligation of either the adhesion molecule ICAM-1/CD54 or MHC II with CD40 inhibited cell cycle progression and promoted...... these effects. Addition of BCR or IL-4 signals did not overcome the effect of ICAM-1 or MHC II on CD40-induced proliferation. FasL expression was not detected in B cell populations. These results show that MHC II and ICAM-1 specifically modulate CD40-mediated signaling, so inhibiting proliferation...

Regulation of P450-mediated permethrin resistance in Culex quinquefasciatus by the GPCR/Gαs/AC/cAMP/PKA signaling cascade.

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Li, Ting; Liu, Nannan

2017-12-01

This study explores the role of G-protein-coupled receptor-intracellular signaling in the development of P450-mediated insecticide resistance in mosquitoes, Culex quinquefasciatus , focusing on the essential function of the GPCRs and their downstream effectors of Gs alpha subunit protein (Gαs) and adenylyl cyclase (ACs) in P450-mediated insecticide resistance of Culex mosquitoes. Our RNAi-mediated functional study showed that knockdown of Gαs caused the decreased expression of the downstream effectors of ACs and PKAs in the GPCR signaling pathway and resistance P450 genes, whereas knockdown of ACs decreased the expression of PKAs and resistance P450 genes. Knockdown of either Gαs or ACs resulted in an increased susceptibility of mosquitoes to permethrin. These results add significantly to our understanding of the molecular basis of resistance P450 gene regulation through GPCR/Gαs/AC/cAMP-PKA signaling pathways in the insecticide resistance of mosquitoes. The temporal and spatial dynamic analyses of GPCRs, Gαs, ACs, PKAs, and P450s in two insecticide resistant mosquito strains revealed that all the GPCR signaling pathway components tested, namely GPCRs, Gαs, ACs and PKAs, were most highly expressed in the brain for both resistant strains, suggesting the role played by these genes in signaling transduction and regulation. The resistance P450 genes were mainly expressed in the brain, midgut and malpighian tubules (MTs), suggesting their critical function in the central nervous system and importance for detoxification. The temporal dynamics analysis for the gene expression showed a diverse expression profile during mosquito development, indicating their initially functional importance in response to exposure to insecticides during their life stages.

A study on genetic variants of Fibroblast growth factor receptor 2 (FGFR2 and the risk of breast cancer from North India.

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Sarah Siddiqui

Full Text Available Genome-Wide Association Studies (GWAS have identified Fibroblast growth factor receptor 2 (FGFR2 as a candidate gene for breast cancer with single nucleotide polymorphisms (SNPs located in intron 2 region as the susceptibility loci strongly associated with the risk. However, replicate studies have often failed to extrapolate the association to diverse ethnic regions. This hints towards the existing heterogeneity among different populations, arising due to differential linkage disequilibrium (LD structures and frequencies of SNPs within the associated regions of the genome. It is therefore important to revisit the previously linked candidates in varied population groups to unravel the extent of heterogeneity. In an attempt to investigate the role of FGFR2 polymorphisms in susceptibility to the risk of breast cancer among North Indian women, we genotyped rs2981582, rs1219648, rs2981578 and rs7895676 polymorphisms in 368 breast cancer patients and 484 healthy controls by Polymerase chain reaction-Restriction fragment length polymorphism (PCR-RFLP assay. We observed a statistically significant association with breast cancer risk for all the four genetic variants (P<0.05. In per-allele model for rs2981582, rs1219648, rs7895676 and in dominant model for rs2981578, association remained significant after bonferroni correction (P<0.0125. On performing stratified analysis, significant correlations with various clinicopathological as well as environmental and lifestyle characteristics were observed. It was evident that rs1219648 and rs2981578 interacted with exogenous hormone use and advanced clinical stage III (after Bonferroni correction, P<0.000694, respectively. Furthermore, combined analysis on these four loci revealed that compared to women with 0-1 risk loci, those with 2-4 risk loci had increased risk (OR = 1.645, 95%CI = 1.152-2.347, P = 0.006. In haplotype analysis, for rs2981578, rs2981582 and rs1219648, risk haplotype (GTG was

Curcumin inhibits urothelial tumor development by suppressing IGF2 and IGF2-mediated PI3K/AKT/mTOR signaling pathway.

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Tian, Binqiang; Zhao, Yingmei; Liang, Tao; Ye, Xuxiao; Li, Zuowei; Yan, Dongliang; Fu, Qiang; Li, Yonghui

2017-08-01

We have previously reported that curcumin inhibits urothelial tumor development in a rat bladder carcinogenesis model. In this study, we report that curcumin inhibits urothelial tumor development by suppressing IGF2 and IGF2-mediated PI3K/AKT/mTOR signaling pathway. Curcumin inhibits IGF2 expression at the transcriptional level and decreases the phosphorylation levels of IGF1R and IRS-1 in bladder cancer cells and N-methyl-N-nitrosourea (MNU)-induced urothelial tumor tissue. Ectopic expression of IGF2 and IGF1R, but not IGF1, in bladder cancer cells restored this process, suggesting that IGF2 is a target of curcumin. Moreover, introduction of constitutively active AKT1 abolished the inhibitory effect of curcumin on cell proliferation, migration, and restored the phosphorylation levels of 4E-BP1 and S6K1, suggesting that curcumin functions via suppressing IGF2-mediated AKT/mTOR signaling pathway. In summary, our results reveal that suppressing IGF2 and IGF2-mediated PI3K/AKT/mTOR signaling pathway is one of the mechanisms of action of curcumin. Our findings suggest a new therapeutic strategy against human bladder cancer caused by aberrant activation of IGF2, which are useful for translational application of curcumin.

Low dose gamma irradiation enhances defined signaling components of intercellular reactive oxygen-mediated apoptosis induction

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Bauer, G

2011-01-01

Transformed cells are selectively removed by intercellular ROS-mediated induction of apoptosis. Signaling is based on the HOCl and the NO/peroxynitrite pathway (major pathways) and the nitryl chloride and the metal-catalyzed Haber-Weiss pathway (minor pathways). During tumor progression, resistance against intercellular induction of apoptosis is acquired through expression of membrane-associated catalase. Low dose radiation of nontransformed cells has been shown to enhance intercellular induction of apoptosis. The present study was performed to define the signaling components which are modulated by low dose gamma irradiation. Low dose radiation induced the release of peroxidase from nontransformed, transformed and tumor cells. Extracellular superoxide anion generation was strongly enhanced in the case of transformed cells and tumor cells, but not in nontransformed cells. Enhancement of peroxidase release and superoxide anion generation either increased intercellular induction of apoptosis of transformed cells, or caused a partial protection under specific signaling conditions. In tumor cells, low dose radiation enhanced the production of major signaling components, but this had no effect on apoptosis induction, due to the strong resistance mechanism of tumor cells. Our data specify the nature of low dose radiation-induced effects on specific signaling components of intercellular induction of apoptosis at defined stages of multistep carcinogenesis.

Synthetic NCAM-derived Ligands of the Fibroblast Growth Factor Receptor

DEFF Research Database (Denmark)

Hansen, Stine; Li, Shizhong; Bock, Elisabeth

2008-01-01

The neural cell adhesion molecule (NCAM) responds to cues in the external environment and transmits signals to the cell through extracellular and intracellular interactions with a number of other signal transduction molecules. One such NCAM interaction partner is the fibroblast growth factor...... various FN3 module loop regions, have been identified as FGFR ligands. All four peptides activate FGFR and differentially modulate a number of neuronal functions, such as differentiation, survival, and synaptic changes that are important for learning, memory, and neuronal regeneration....

Wnt/β-catenin signalling pathway mediated aberrant hippocampal neurogenesis in kainic acid-induced epilepsy.

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Qu, Zhengyi; Su, Fang; Qi, Xueting; Sun, Jianbo; Wang, Hongcai; Qiao, Zhenkui; Zhao, Hong; Zhu, Yulan

2017-10-01

Temporal lobe epilepsy is a chronic disorder of nerve system, mainly characterized by hippocampal sclerosis with massive neuronal loss and severe gliosis. Aberrant neurogenesis has been shown in the epileptogenesis process of temporal lobe epilepsy. However, the molecular mechanisms underlying aberrant neurogenesis remain unclear. The roles of Wnt signalling cascade have been well established in neurogenesis during multiple aspects. Here, we used kainic acid-induced rat epilepsy model to investigate whether Wnt/β-catenin signalling pathway is involved in the aberrant neurogenesis in temporal lobe epilepsy. Immunostaining and western blotting results showed that the expression levels of β-catenin, Wnt3a, and cyclin D1, the key regulators in Wnt signalling pathway, were up-regulated during acute epilepsy induced by the injection of kainic acids, indicating that Wnt signalling pathway was activated in kainic acid-induced temporal lobe epilepsy. Moreover, BrdU labelling results showed that blockade of the Wnt signalling by knocking down β-catenin attenuated aberrant neurogenesis induced by kainic acids injection. Altogether, Wnt/β-catenin signalling pathway mediated hippocampal neurogenesis during epilepsy, which might provide new strategies for clinical treatment of temporal lobe epilepsy. Temporal lobe epilepsy is a chronic disorder of nerve system, mainly characterized by hippocampal sclerosis. Aberrant neurogenesis has been shown to involve in the epileptogenesis process of temporal lobe epilepsy. In the present study, we discovered that Wnt3a/β-catenin signalling pathway serves as a link between aberrant neurogenesis and underlying remodelling in the hippocampus, leading to temporal lobe epilepsy, which might provide new strategies for clinical treatment of temporal lobe epilepsy. Copyright © 2017 John Wiley & Sons, Ltd.

Multivalent Soluble Antigen Arrays Exhibit High Avidity Binding and Modulation of B Cell Receptor-Mediated Signaling to Drive Efficacy against Experimental Autoimmune Encephalomyelitis.

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Hartwell, Brittany L; Pickens, Chad J; Leon, Martin; Berkland, Cory

2017-06-12

A pressing need exists for antigen-specific immunotherapies (ASIT) that induce selective tolerance in autoimmune disease while avoiding deleterious global immunosuppression. Multivalent soluble antigen arrays (SAgA PLP:LABL ), consisting of a hyaluronic acid (HA) linear polymer backbone cografted with multiple copies of autoantigen (PLP) and cell adhesion inhibitor (LABL) peptides, are designed to induce tolerance to a specific multiple sclerosis (MS) autoantigen. Previous studies established that hydrolyzable SAgA PLP:LABL , employing a degradable linker to codeliver PLP and LABL, was therapeutic in experimental autoimmune encephalomyelitis (EAE) in vivo and exhibited antigen-specific binding with B cells, targeted the B cell receptor (BCR), and dampened BCR-mediated signaling in vitro. Our results pointed to sustained BCR engagement as the SAgA PLP:LABL therapeutic mechanism, so we developed a new version of the SAgA molecule using nonhydrolyzable conjugation chemistry, hypothesizing it would enhance and maintain the molecule's action at the cell surface to improve efficacy. "Click SAgA" (cSAgA PLP:LABL ) uses hydrolytically stable covalent conjugation chemistry (Copper-catalyzed Azide-Alkyne Cycloaddition (CuAAC)) rather than a hydrolyzable oxime bond to attach PLP and LABL to HA. We explored cSAgA PLP:LABL B cell engagement and modulation of BCR-mediated signaling in vitro through flow cytometry binding and calcium flux signaling assays. Indeed, cSAgA PLP:LABL exhibited higher avidity B cell binding and greater dampening of BCR-mediated signaling than hydrolyzable SAgA PLP:LABL . Furthermore, cSAgA PLP:LABL exhibited significantly enhanced in vivo efficacy compared to hydrolyzable SAgA PLP:LABL , achieving equivalent efficacy at one-quarter of the dose. These results indicate that nonhydrolyzable conjugation increased the avidity of cSAgA PLP:LABL to drive in vivo efficacy through modulated BCR-mediated signaling.

Protein tyrosine kinase and mitogen-activated protein kinase signalling pathways contribute to differences in heterophil-mediated innate immune responsiveness between two lines of broilers

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Protein tyrosine phosphorylation mediates signal transduction of cellular processes, with protein tyrosine kinases (PTKs) regulating virtually all signaling events. The mitogen-activated protein kinase (MAPK) super-family consists of three conserved pathways that convert receptor activation into ce...

Dopamine Receptor D3 Signaling on CD4+ T Cells Favors Th1- and Th17-Mediated Immunity.

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Contreras, Francisco; Prado, Carolina; González, Hugo; Franz, Dafne; Osorio-Barrios, Francisco; Osorio, Fabiola; Ugalde, Valentina; Lopez, Ernesto; Elgueta, Daniela; Figueroa, Alicia; Lladser, Alvaro; Pacheco, Rodrigo

2016-05-15

Dopamine receptor D3 (DRD3) expressed on CD4(+) T cells is required to promote neuroinflammation in a murine model of Parkinson's disease. However, how DRD3 signaling affects T cell-mediated immunity remains unknown. In this study, we report that TCR stimulation on mouse CD4(+) T cells induces DRD3 expression, regardless of the lineage specification. Importantly, functional analyses performed in vivo using adoptive transfer of OVA-specific OT-II cells into wild-type recipients show that DRD3 deficiency in CD4(+) T cells results in attenuated differentiation of naive CD4(+) T cells toward the Th1 phenotype, exacerbated generation of Th2 cells, and unaltered Th17 differentiation. The reciprocal regulatory effect of DRD3 signaling in CD4(+) T cells favoring Th1 generation and impairing the acquisition of Th2 phenotype was also reproduced using in vitro approaches. Mechanistic analysis indicates that DRD3 signaling evokes suppressor of cytokine signaling 5 expression, a negative regulator of Th2 development, which indirectly favors acquisition of Th1 phenotype. Accordingly, DRD3 deficiency results in exacerbated eosinophil infiltration into the airways of mice undergoing house dust mite-induced allergic response. Interestingly, our results show that, upon chronic inflammatory colitis induced by transfer of naive CD4(+) T cells into lymphopenic recipients, DRD3 deficiency not only affects Th1 response, but also the frequency of Th17 cells, suggesting that DRD3 signaling also contributes to Th17 expansion under chronic inflammatory conditions. In conclusion, our findings indicate that DRD3-mediated signaling in CD4(+) T cells plays a crucial role in the balance of effector lineages, favoring the inflammatory potential of CD4(+) T cells. Copyright © 2016 by The American Association of Immunologists, Inc.

Human SR-BII mediates SAA uptake and contributes to SAA pro-inflammatory signaling in vitro and in vivo.

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Irina N Baranova

Full Text Available Serum amyloid A (SAA is an acute phase protein with cytokine-like and chemotactic properties, that is markedly up-regulated during various inflammatory conditions. Several receptors, including FPRL-1, TLR2, TLR4, RAGE, class B scavenger receptors, SR-BI and CD36, have been identified as SAA receptors. This study provides new evidence that SR-BII, splice variant of SR-BI, could function as an SAA receptor mediating its uptake and pro-inflammatory signaling. The uptake of Alexa Fluor488 SAA was markedly (~3 fold increased in hSR-BII-expressing HeLa cells when compared with mock-transfected cells. The levels of SAA-induced interleukin-8 secretion by hSR-BII-expressing HEK293 cells were also significantly (~3-3.5 fold higher than those detected in control cells. Moderately enhanced levels of phosphorylation of all three mitogen-activated protein kinases, ERK1/2, and p38 and JNK, were observed in hSR-BII-expressing cells following SAA stimulation when compared with control wild type cells. Transgenic mice with pLiv-11-directed liver/kidney overexpression of hSR-BI or hSR-BII were used to assess the in vivo role of each receptor in SAA-induced pro-inflammatory response in these organs. Six hours after intraperitoneal SAA injection both groups of transgenic mice demonstrated markedly higher (~2-5-fold expression levels of inflammatory mediators in the liver and kidney compared to wild type mice. Histological examinations of hepatic and renal tissue from SAA-treated mice revealed moderate level of damage in the liver of both transgenic but not in the wild type mice. Activities of plasma transaminases, biomarkers of liver injury, were also moderately higher in hSR-B transgenic mice when compared to wild type mice. Our findings identify hSR-BII as a functional SAA receptor that mediates SAA uptake and contributes to its pro-inflammatory signaling via the MAPKs-mediated signaling pathways.

Regulator of G-protein signaling - 5 (RGS5 is a novel repressor of hedgehog signaling.

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William M Mahoney

Full Text Available Hedgehog (Hh signaling plays fundamental roles in morphogenesis, tissue repair, and human disease. Initiation of Hh signaling is controlled by the interaction of two multipass membrane proteins, patched (Ptc and smoothened (Smo. Recent studies identify Smo as a G-protein coupled receptor (GPCR-like protein that signals through large G-protein complexes which contain the Gαi subunit. We hypothesize Regulator of G-Protein Signaling (RGS proteins, and specifically RGS5, are endogenous repressors of Hh signaling via their ability to act as GTPase activating proteins (GAPs for GTP-bound Gαi, downstream of Smo. In support of this hypothesis, we demonstrate that RGS5 over-expression inhibits sonic hedgehog (Shh-mediated signaling and osteogenesis in C3H10T1/2 cells. Conversely, signaling is potentiated by siRNA-mediated knock-down of RGS5 expression, but not RGS4 expression. Furthermore, using immuohistochemical analysis and co-immunoprecipitation (Co-IP, we demonstrate that RGS5 is present with Smo in primary cilia. This organelle is required for canonical Hh signaling in mammalian cells, and RGS5 is found in a physical complex with Smo in these cells. We therefore conclude that RGS5 is an endogenous regulator of Hh-mediated signaling and that RGS proteins are potential targets for novel therapeutics in Hh-mediated diseases.

SAD-A kinase controls islet β-cell size and function as a mediator of mTORC1 signaling.

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Nie, Jia; Liu, Xiaolei; Lilley, Brendan N; Zhang, Hai; Pan, Y Albert; Kimball, Scot R; Zhang, Jun; Zhang, Weiping; Wang, Li; Jefferson, Leonard S; Sanes, Joshua R; Han, Xiao; Shi, Yuguang

2013-08-20

The mammalian target of rapamycin (mTOR) plays an important role in controlling islet β-cell function. However, the underlying molecular mechanisms remain poorly elucidated. Synapses of amphids defective kinase-A (SAD-A) is a 5' adenosine monophosphate-activated protein kinase-related protein kinase that is exclusively expressed in pancreas and brain. In this study, we investigated a role of the kinase in regulating pancreatic β-cell morphology and function as a mediator of mTOR complex 1 (mTORC1) signaling. We show that global SAD-A deletion leads to defective glucose-stimulated insulin secretion and petite islets, which are reminiscent of the defects in mice with global deletion of ribosomal protein S6 kinase 1, a downstream target of mTORC1. Consistent with these findings, selective deletion of SAD-A in pancreas decreased islet β-cell size, whereas SAD-A overexpression significantly increased the size of mouse insulinomas cell lines β-cells. In direct support of SAD-A as a unique mediator of mTORC1 signaling in islet β-cells, we demonstrate that glucose dramatically stimulated SAD-A protein translation in isolated mouse islets, which was potently inhibited by rapamycin, an inhibitor of mTORC1. Moreover, the 5'-untranslated region of SAD-A mRNA is highly structured and requires mTORC1 signaling for its translation initiation. Together, these findings identified SAD-A as a unique pancreas-specific effector protein of mTORC1 signaling.

Achondroplasia: Development, pathogenesis, and therapy.

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Ornitz, David M; Legeai-Mallet, Laurence

2017-04-01

Autosomal dominant mutations in fibroblast growth factor receptor 3 (FGFR3) cause achondroplasia (Ach), the most common form of dwarfism in humans, and related chondrodysplasia syndromes that include hypochondroplasia (Hch), severe achondroplasia with developmental delay and acanthosis nigricans (SADDAN), and thanatophoric dysplasia (TD). FGFR3 is expressed in chondrocytes and mature osteoblasts where it functions to regulate bone growth. Analysis of the mutations in FGFR3 revealed increased signaling through a combination of mechanisms that include stabilization of the receptor, enhanced dimerization, and enhanced tyrosine kinase activity. Paradoxically, increased FGFR3 signaling profoundly suppresses proliferation and maturation of growth plate chondrocytes resulting in decreased growth plate size, reduced trabecular bone volume, and resulting decreased bone elongation. In this review, we discuss the molecular mechanisms that regulate growth plate chondrocytes, the pathogenesis of Ach, and therapeutic approaches that are being evaluated to improve endochondral bone growth in people with Ach and related conditions. Developmental Dynamics 246:291-309, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

ZINC-INDUCED EGF RECEPTOR SIGNALING REQUIRES SRC-MEDIATED PHOSPHORYLATION OF THE EGF RECEPTOR ON TYROSINE 845 (Y845)

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ZINC-INDUCED EGF RECEPTOR SIGNALING REQUIRES Src-MEDIATED PHOSPHORYLATION OF THE EGF RECEPTOR ON TYROSINE 845 (Y845)Weidong Wu1, Lee M. Graves2, Gordon N. Gill3 and James M. Samet4 1Center for Environmental Medicine and Lung Biology; 2Department of Pharmacology, University o...

Disrupted epithelial/macrophage crosstalk via Spinster homologue 2-mediated S1P signaling may drive defective macrophage phagocytic function in COPD.

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Tran, Hai B; Jersmann, Hubertus; Truong, Tung Thanh; Hamon, Rhys; Roscioli, Eugene; Ween, Miranda; Pitman, Melissa R; Pitson, Stuart M; Hodge, Greg; Reynolds, Paul N; Hodge, Sandra

2017-01-01

We have previously established a link between impaired phagocytic capacity and deregulated S1P signaling in alveolar macrophages from COPD subjects. We hypothesize that this defect may include a disruption of epithelial-macrophage crosstalk via Spns2-mediated intercellular S1P signaling. Primary alveolar macrophages and bronchial epithelial cells from COPD subjects and controls, cell lines, and a mouse model of chronic cigarette smoke exposure were studied. Cells were exposed to 10% cigarette smoke extract, or vehicle control. Spns2 expression and subcellular localization was studied by immunofluorescence, confocal microscopy and RT-PCR. Phagocytosis was assessed by flow-cytometry. Levels of intra- and extracellular S1P were measured by S1P [3H]-labeling. Spns2 expression was significantly increased (pS1P in the airway and that there is a possible disruption of epithelial/macrophage cross talk via Spns2-mediated S1P signaling in COPD and in response to cigarette smoke exposure.

Dectin-1-mediated signaling leads to characteristic gene expressions and cytokine secretion via spleen tyrosine kinase (Syk) in rat mast cells.

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Kimura, Yukihiro; Chihara, Kazuyasu; Honjoh, Chisato; Takeuchi, Kenji; Yamauchi, Shota; Yoshiki, Hatsumi; Fujieda, Shigeharu; Sada, Kiyonao

2014-11-07

Dectin-1 recognizes β-glucan and plays important roles for the antifungal immunity through the activation of spleen tyrosine kinase (Syk) in dendritic cells or macrophages. Recently, expression of Dectin-1 was also identified in human and mouse mast cells, although its physiological roles were largely unknown. In this report, rat mast cell line RBL-2H3 was analyzed to investigate the molecular mechanism of Dectin-1-mediated activation and responses of mast cells. Treatment of cells with Dectin-1-specific agonist curdlan induced tyrosine phosphorylation of cellular proteins and the interaction of Dectin-1 with the Src homology 2 domain of Syk. These responses depended on tyrosine phosphorylation of the hemi-immunoreceptor tyrosine-based activation motif in the cytoplasmic tail of Dectin-1, whereas they were independent of the γ-subunit of high-affinity IgE receptor. DNA microarray and real-time PCR analyses showed that Dectin-1-mediated signaling stimulated gene expression of transcription factor Nfkbiz and inflammatory cytokines, such as monocyte chemoattractant protein-1, IL-3, IL-4, IL-13, and tumor necrosis factor (TNF)-α. The response was abrogated by pretreatment with Syk inhibitor R406. These results suggest that Syk is critical for Dectin-1-mediated activation of mast cells, although the signaling differs from that triggered by FcϵRI activation. In addition, these gene expressions induced by curdlan stimulation were specifically observed in mast cells, suggesting that Dectin-1-mediated signaling of mast cells offers new insight into the antifungal immunity. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

mTORC1 is a critical mediator of oncogenic Semaphorin3A signaling

Energy Technology Data Exchange (ETDEWEB)

Yamada, Daisuke; Kawahara, Kohichi; Maeda, Takehiko, E-mail: maeda@nupals.ac.jp

2016-08-05

Aberration of signaling pathways by genetic mutations or alterations in the surrounding tissue environments can result in tumor development or metastasis. However, signaling molecules responsible for these processes have not been completely elucidated. Here, we used mouse Lewis lung carcinoma cells (LLC) to explore the mechanism by which the oncogenic activity of Semaphorin3A (Sema3A) signaling is regulated. Sema3A knockdown by shRNA did not affect apoptosis, but decreased cell proliferation in LLCs; both the mammalian target of rapamycin complex 1 (mTORC1) level and glycolytic activity were also decreased. In addition, Sema3A knockdown sensitized cells to inhibition of oxidative phosphorylation by oligomycin, but conferred resistance to decreased cell viability induced by glucose starvation. Furthermore, recombinant SEMA3A rescued the attenuation of cell proliferation and glycolytic activity in LLCs after Sema3A knockdown, whereas mTORC1 inhibition by rapamycin completely counteracted this effect. These results demonstrate that Sema3A signaling exerts its oncogenic effect by promoting an mTORC1-mediated metabolic shift from oxidative phosphorylation to aerobic glycolysis. -- Highlights: •Sema3A knockdown decreased proliferation of Lewis lung carcinoma cells (LLCs). •Sema3A knockdown decreased mTORC1 levels and glycolytic activity in LLCs. •Sema3A knockdown sensitized cells to inhibition of oxidative phosphorylation. •Sema3A promotes shift from oxidative phosphorylation to aerobic glycolysis via mTORC1.

Polycomb-Mediated Repression and Sonic Hedgehog Signaling Interact to Regulate Merkel Cell Specification during Skin Development.

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Carolina N Perdigoto

2016-07-01

Full Text Available An increasing amount of evidence indicates that developmental programs are tightly regulated by the complex interplay between signaling pathways, as well as transcriptional and epigenetic processes. Here, we have uncovered coordination between transcriptional and morphogen cues to specify Merkel cells, poorly understood skin cells that mediate light touch sensations. In murine dorsal skin, Merkel cells are part of touch domes, which are skin structures consisting of specialized keratinocytes, Merkel cells, and afferent neurons, and are located exclusively around primary hair follicles. We show that the developing primary hair follicle functions as a niche required for Merkel cell specification. We find that intraepidermal Sonic hedgehog (Shh signaling, initiated by the production of Shh ligand in the developing hair follicles, is required for Merkel cell specification. The importance of Shh for Merkel cell formation is further reinforced by the fact that Shh overexpression in embryonic epidermal progenitors leads to ectopic Merkel cells. Interestingly, Shh signaling is common to primary, secondary, and tertiary hair follicles, raising the possibility that there are restrictive mechanisms that regulate Merkel cell specification exclusively around primary hair follicles. Indeed, we find that loss of Polycomb repressive complex 2 (PRC2 in the epidermis results in the formation of ectopic Merkel cells that are associated with all hair types. We show that PRC2 loss expands the field of epidermal cells competent to differentiate into Merkel cells through the upregulation of key Merkel-differentiation genes, which are known PRC2 targets. Importantly, PRC2-mediated repression of the Merkel cell differentiation program requires inductive Shh signaling to form mature Merkel cells. Our study exemplifies how the interplay between epigenetic and morphogen cues regulates the complex patterning and formation of the mammalian skin structures.

Polycomb-Mediated Repression and Sonic Hedgehog Signaling Interact to Regulate Merkel Cell Specification during Skin Development.

Science.gov (United States)

Perdigoto, Carolina N; Dauber, Katherine L; Bar, Carmit; Tsai, Pai-Chi; Valdes, Victor J; Cohen, Idan; Santoriello, Francis J; Zhao, Dejian; Zheng, Deyou; Hsu, Ya-Chieh; Ezhkova, Elena

2016-07-01

An increasing amount of evidence indicates that developmental programs are tightly regulated by the complex interplay between signaling pathways, as well as transcriptional and epigenetic processes. Here, we have uncovered coordination between transcriptional and morphogen cues to specify Merkel cells, poorly understood skin cells that mediate light touch sensations. In murine dorsal skin, Merkel cells are part of touch domes, which are skin structures consisting of specialized keratinocytes, Merkel cells, and afferent neurons, and are located exclusively around primary hair follicles. We show that the developing primary hair follicle functions as a niche required for Merkel cell specification. We find that intraepidermal Sonic hedgehog (Shh) signaling, initiated by the production of Shh ligand in the developing hair follicles, is required for Merkel cell specification. The importance of Shh for Merkel cell formation is further reinforced by the fact that Shh overexpression in embryonic epidermal progenitors leads to ectopic Merkel cells. Interestingly, Shh signaling is common to primary, secondary, and tertiary hair follicles, raising the possibility that there are restrictive mechanisms that regulate Merkel cell specification exclusively around primary hair follicles. Indeed, we find that loss of Polycomb repressive complex 2 (PRC2) in the epidermis results in the formation of ectopic Merkel cells that are associated with all hair types. We show that PRC2 loss expands the field of epidermal cells competent to differentiate into Merkel cells through the upregulation of key Merkel-differentiation genes, which are known PRC2 targets. Importantly, PRC2-mediated repression of the Merkel cell differentiation program requires inductive Shh signaling to form mature Merkel cells. Our study exemplifies how the interplay between epigenetic and morphogen cues regulates the complex patterning and formation of the mammalian skin structures.

Augmentation of Muscle Blood Flow by Ultrasound Cavitation Is Mediated by ATP and Purinergic Signaling.

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Belcik, J Todd; Davidson, Brian P; Xie, Aris; Wu, Melinda D; Yadava, Mrinal; Qi, Yue; Liang, Sherry; Chon, Chae Ryung; Ammi, Azzdine Y; Field, Joshua; Harmann, Leanne; Chilian, William M; Linden, Joel; Lindner, Jonathan R

2017-03-28

Augmentation of tissue blood flow by therapeutic ultrasound is thought to rely on convective shear. Microbubble contrast agents that undergo ultrasound-mediated cavitation markedly amplify these effects. We hypothesized that purinergic signaling is responsible for shear-dependent increases in muscle perfusion during therapeutic cavitation. Unilateral exposure of the proximal hindlimb of mice (with or without ischemia produced by iliac ligation) to therapeutic ultrasound (1.3 MHz, mechanical index 1.3) was performed for 10 minutes after intravenous injection of 2×10 8 lipid microbubbles. Microvascular perfusion was evaluated by low-power contrast ultrasound perfusion imaging. In vivo muscle ATP release and in vitro ATP release from endothelial cells or erythrocytes were assessed by a luciferin-luciferase assay. Purinergic signaling pathways were assessed by studying interventions that (1) accelerated ATP degradation; (2) inhibited P2Y receptors, adenosine receptors, or K ATP channels; or (3) inhibited downstream signaling pathways involving endothelial nitric oxide synthase or prostanoid production (indomethacin). Augmentation in muscle perfusion by ultrasound cavitation was assessed in a proof-of-concept clinical trial in 12 subjects with stable sickle cell disease. Therapeutic ultrasound cavitation increased muscle perfusion by 7-fold in normal mice, reversed tissue ischemia for up to 24 hours in the murine model of peripheral artery disease, and doubled muscle perfusion in patients with sickle cell disease. Augmentation in flow extended well beyond the region of ultrasound exposure. Ultrasound cavitation produced an ≈40-fold focal and sustained increase in ATP, the source of which included both endothelial cells and erythrocytes. Inhibitory studies indicated that ATP was a critical mediator of flow augmentation that acts primarily through either P2Y receptors or adenosine produced by ectonucleotidase activity. Combined indomethacin and inhibition of

A multiscale computational approach to dissect early events in the Erb family receptor mediated activation, differential signaling, and relevance to oncogenic transformations.

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Liu, Yingting; Purvis, Jeremy; Shih, Andrew; Weinstein, Joshua; Agrawal, Neeraj; Radhakrishnan, Ravi

2007-06-01

We describe a hierarchical multiscale computational approach based on molecular dynamics simulations, free energy-based molecular docking simulations, deterministic network-based kinetic modeling, and hybrid discrete/continuum stochastic dynamics protocols to study the dimer-mediated receptor activation characteristics of the Erb family receptors, specifically the epidermal growth factor receptor (EGFR). Through these modeling approaches, we are able to extend the prior modeling of EGF-mediated signal transduction by considering specific EGFR tyrosine kinase (EGFRTK) docking interactions mediated by differential binding and phosphorylation of different C-terminal peptide tyrosines on the RTK tail. By modeling signal flows through branching pathways of the EGFRTK resolved on a molecular basis, we are able to transcribe the effects of molecular alterations in the receptor (e.g., mutant forms of the receptor) to differing kinetic behavior and downstream signaling response. Our molecular dynamics simulations show that the drug sensitizing mutation (L834R) of EGFR stabilizes the active conformation to make the system constitutively active. Docking simulations show preferential characteristics (for wildtype vs. mutant receptors) in inhibitor binding as well as preferential enhancement of phosphorylation of particular substrate tyrosines over others. We find that in comparison to the wildtype system, the L834R mutant RTK preferentially binds the inhibitor erlotinib, as well as preferentially phosphorylates the substrate tyrosine Y1068 but not Y1173. We predict that these molecular level changes result in preferential activation of the Akt signaling pathway in comparison to the Erk signaling pathway for cells with normal EGFR expression. For cells with EGFR over expression, the mutant over activates both Erk and Akt pathways, in comparison to wildtype. These results are consistent with qualitative experimental measurements reported in the literature. We discuss these

Reactive oxygen species mediate Cr(VI)-induced carcinogenesis through PI3K/AKT-dependent activation of GSK-3β/β-catenin signaling

International Nuclear Information System (INIS)

Son, Young-Ok; Pratheeshkumar, Poyil; Wang, Lei; Wang, Xin; Fan, Jia; Kim, Dong-Hern; Lee, Ju-Yeon; Zhang, Zhuo; Lee, Jeong-Chae; Shi, Xianglin

2013-01-01

Cr(VI) compounds are known human carcinogens that primarily target the lungs. Cr(VI) produces reactive oxygen species (ROS), but the exact effects of ROS on the signaling molecules involved in Cr(VI)-induced carcinogenesis have not been extensively studied. Chronic exposure of human bronchial epithelial cells to Cr(VI) at nanomolar concentrations (10–100 nM) for 3 months not only induced cell transformation, but also increased the potential of these cells to invade and migrate. Injection of Cr(VI)-stimulated cells into nude mice resulted in the formation of tumors. Chronic exposure to Cr(VI) increased levels of intracellular ROS and antiapoptotic proteins. Transfection with catalase or superoxide dismutase (SOD) prevented Cr(VI)-mediated increases in colony formation, cell invasion, migration, and xenograft tumors. While chronic Cr(VI) exposure led to activation of signaling cascades involving PI3K/AKT/GSK-3β/β-catenin and PI3K/AKT/mTOR, transfection with catalase or SOD markedly inhibited Cr(VI)-mediated activation of these signaling proteins. Inhibitors specific for AKT or β-catenin almost completely suppressed the Cr(VI)-mediated increase in total and active β-catenin proteins and colony formation. In particular, Cr(VI) suppressed autophagy of epithelial cells under nutrition deprivation. Furthermore, there was a marked induction of AKT, GSK-3β, β-catenin, mTOR, and carcinogenic markers in tumor tissues formed in mice after injection with Cr(VI)-stimulated cells. Collectively, our findings suggest that ROS is a key mediator of Cr(VI)-induced carcinogenesis through the activation of PI3K/AKT-dependent GSK-3β/β-catenin signaling and the promotion of cell survival mechanisms via the inhibition of apoptosis and autophagy. - Highlights: • Chronic exposure to Cr(VI) induces carcinogenic properties in BEAS-2B cells. • ROS play an important role in Cr(VI)-induced tumorigenicity of BEAS-2B cells. • PI3K/AKT/GSK-3β/β-catenin signaling involved in Cr

Reactive oxygen species mediate Cr(VI)-induced carcinogenesis through PI3K/AKT-dependent activation of GSK-3β/β-catenin signaling

Energy Technology Data Exchange (ETDEWEB)

Son, Young-Ok; Pratheeshkumar, Poyil; Wang, Lei; Wang, Xin; Fan, Jia; Kim, Dong-Hern; Lee, Ju-Yeon; Zhang, Zhuo [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States); Lee, Jeong-Chae [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States); School of Dentistry and Institute of Oral Biosciences, Research Center of Bioactive Materials, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Shi, Xianglin, E-mail: xshi5@email.uky.edu [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States)

2013-09-01

Cr(VI) compounds are known human carcinogens that primarily target the lungs. Cr(VI) produces reactive oxygen species (ROS), but the exact effects of ROS on the signaling molecules involved in Cr(VI)-induced carcinogenesis have not been extensively studied. Chronic exposure of human bronchial epithelial cells to Cr(VI) at nanomolar concentrations (10–100 nM) for 3 months not only induced cell transformation, but also increased the potential of these cells to invade and migrate. Injection of Cr(VI)-stimulated cells into nude mice resulted in the formation of tumors. Chronic exposure to Cr(VI) increased levels of intracellular ROS and antiapoptotic proteins. Transfection with catalase or superoxide dismutase (SOD) prevented Cr(VI)-mediated increases in colony formation, cell invasion, migration, and xenograft tumors. While chronic Cr(VI) exposure led to activation of signaling cascades involving PI3K/AKT/GSK-3β/β-catenin and PI3K/AKT/mTOR, transfection with catalase or SOD markedly inhibited Cr(VI)-mediated activation of these signaling proteins. Inhibitors specific for AKT or β-catenin almost completely suppressed the Cr(VI)-mediated increase in total and active β-catenin proteins and colony formation. In particular, Cr(VI) suppressed autophagy of epithelial cells under nutrition deprivation. Furthermore, there was a marked induction of AKT, GSK-3β, β-catenin, mTOR, and carcinogenic markers in tumor tissues formed in mice after injection with Cr(VI)-stimulated cells. Collectively, our findings suggest that ROS is a key mediator of Cr(VI)-induced carcinogenesis through the activation of PI3K/AKT-dependent GSK-3β/β-catenin signaling and the promotion of cell survival mechanisms via the inhibition of apoptosis and autophagy. - Highlights: • Chronic exposure to Cr(VI) induces carcinogenic properties in BEAS-2B cells. • ROS play an important role in Cr(VI)-induced tumorigenicity of BEAS-2B cells. • PI3K/AKT/GSK-3β/β-catenin signaling involved in Cr

Evidence that the 5p12 Variant rs10941679 Confers Susceptibility to Estrogen-Receptor-Positive Breast Cancer through FGF10 and MRPS30 Regulation

DEFF Research Database (Denmark)

Ghoussaini, Maya; French, Juliet D; Michailidou, Kyriaki

2016-01-01

expression of FGF10 and MRPS30. Functional assays demonstrated that SNP rs10941679 maps to an enhancer element that physically interacts with the FGF10 and MRPS30 promoter regions in breast cancer cell lines. FGF10 is an oncogene that binds to FGFR2 and is overexpressed in ∼10% of human breast cancers......, whereas MRPS30 plays a key role in apoptosis. These data suggest that the strongest signal of association at 5p12 is mediated through coordinated activation of FGF10 and MRPS30, two candidate genes for breast cancer pathogenesis....

Vitamin K2 downregulates the expression of fibroblast growth factor receptor 3 in human hepatocellular carcinoma cells.

Science.gov (United States)

Cao, Ke; Liu, Weidong; Nakamura, Hideji; Enomoto, Hirayuki; Yamamoto, Teruhisa; Saito, Masaki; Imanishi, Hiroyasu; Shimomura, Soji; Cao, Peiguo; Nishiguchi, Shuhei

2009-11-01

Vitamin K2 exerts an antitumor activity on human hepatocellular carcinoma (HCC), however, its inhibitory mechanism has not yet been clarified. This study was designed to identify the attractive target molecule of vitamin K2 and shed some light on its effects on fibroblast growth factor receptor (FGFR)3 in HCC cells. The changes in the gene expression of HuH-7 after vitamin K2 treatment were evaluated by a DNA chip analysis. The mRNA and protein levels of FGFR were evaluated by semiquantitative reverse transcription polymerase chain reaction (RT-PCR), real-time PCR and western blot analysis. The promoter activity of the FGFR3 gene was measured by a dual-luciferase assay. The DNA chip analysis revealed different inhibitory rates of gene expression of FGFR3 (60.6%) and FGFR1 (19.4%) after vitamin K2 treatment. Vitamin K2 suppresses the proliferation of HuH-7 in a dose-dependent manner and its inhibitory rate reached approximately 61.8% at the dose of 30 microM. FGFR3 mRNA was significantly reduced based on semiquantitative RT-PCR and decreased 61.5% by a real-time PCR method after vitamin K2 treatment, but FGFR1 mRNA was not. The level of FGFR3 protein was also reduced by vitamin K2 treatment. The luciferase assay demonstrated that vitamin K2 significantly suppressed the promoter activity of FGFR3. Furthermore, the FGFR3-ERK1/2 signaling pathway was suppressed by vitamin K2 treatment. These findings suggest that vitamin K2 may suppress the proliferation of HCC cells through the downregulation of the FGFR3 expression. The transcriptional suppression of FGFR3 may be a novel mechanism of the vitamin K2 action for HCC cells.

Characterization of NAADP-mediated calcium signaling in human spermatozoa

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Sánchez-Tusie, A.A. [Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos (Mexico); Vasudevan, S.R.; Churchill, G.C. [Department of Pharmacology, University of Oxford, Oxford OX1 3QT, England (United Kingdom); Nishigaki, T. [Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos (Mexico); Treviño, C.L., E-mail: ctrevino@ibt.unam.mx [Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos (Mexico)

2014-01-10

Highlights: •Human sperm cells synthesize NAADP. •NAADP-AM mediates [Ca{sup 2+}]{sub i} increases in human sperm in the absence of [Ca{sup 2+}]{sub o}. •Human sperm have two acidic compartments located in the head and midpiece. -- Abstract: Ca{sup 2+} signaling in spermatozoa plays a crucial role during processes such as capacitation and release of the acrosome, but the underlying molecular mechanisms still remain unclear. Nicotinic acid adenine dinucleotide phosphate (NAADP) is a potent Ca{sup 2+}-releasing second messenger in a variety of cellular processes. The presence of a NAADP synthesizing enzyme in sea urchin sperm has been previously reported, suggesting a possible role of NAADP in sperm Ca{sup 2+} signaling. In this work we used in vitro enzyme assays to show the presence of a novel NAADP synthesizing enzyme in human sperm, and to characterize its sensitivity to Ca{sup 2+} and pH. Ca{sup 2+} fluorescence imaging studies demonstrated that the permeable form of NAADP (NAADP-AM) induces intracellular [Ca{sup 2+}] increases in human sperm even in the absence of extracellular Ca{sup 2+}. Using LysoTracker®, a fluorescent probe that selectively accumulates in acidic compartments, we identified two such stores in human sperm cells. Their acidic nature was further confirmed by the reduction in staining intensity observed upon inhibition of the endo-lysosomal proton pump with Bafilomycin, or after lysosomal bursting with glycyl-L-phenylalanine-2-naphthylamide. The selective fluorescent NAADP analog, Ned-19, stained the same subcellular regions as LysoTracker®, suggesting that these stores are the targets of NAADP action.

Bmp signaling mediates endoderm pouch morphogenesis by regulating Fgf signaling in zebrafish

Science.gov (United States)

Swartz, Mary E.; McCarthy, Neil; Norrie, Jacqueline L.; Eberhart, Johann K.

2016-01-01

The endodermal pouches are a series of reiterated structures that segment the pharyngeal arches and help pattern the vertebrate face. Multiple pathways regulate the complex process of endodermal development, including the Bone morphogenetic protein (Bmp) pathway. However, the role of Bmp signaling in pouch morphogenesis is poorly understood. Using genetic and chemical inhibitor approaches, we show that pouch morphogenesis requires Bmp signaling from 10-18 h post-fertilization, immediately following gastrulation. Blocking Bmp signaling during this window results in morphological defects to the pouches and craniofacial skeleton. Using genetic chimeras we show that Bmp signals directly to the endoderm for proper morphogenesis. Time-lapse imaging and analysis of reporter transgenics show that Bmp signaling is necessary for pouch outpocketing via the Fibroblast growth factor (Fgf) pathway. Double loss-of-function analyses demonstrate that Bmp and Fgf signaling interact synergistically in craniofacial development. Collectively, our analyses shed light on the tissue and signaling interactions that regulate development of the vertebrate face. PMID:27122171

Mycobacterium tuberculosis Controls Phagosomal Acidification by Targeting CISH-Mediated Signaling

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Christophe J. Queval

2017-09-01

Full Text Available Pathogens have evolved a range of mechanisms to counteract host defenses, notably to survive harsh acidic conditions in phagosomes. In the case of Mycobacterium tuberculosis, it has been shown that regulation of phagosome acidification could be achieved by interfering with the retention of the V-ATPase complexes at the vacuole. Here, we present evidence that M. tuberculosis resorts to yet another strategy to control phagosomal acidification, interfering with host suppressor of cytokine signaling (SOCS protein functions. More precisely, we show that infection of macrophages with M. tuberculosis leads to granulocyte-macrophage colony-stimulating factor (GM-CSF secretion, inducing STAT5-mediated expression of cytokine-inducible SH2-containing protein (CISH, which selectively targets the V-ATPase catalytic subunit A for ubiquitination and degradation by the proteasome. Consistently, we show that inhibition of CISH expression leads to reduced replication of M. tuberculosis in macrophages. Our findings further broaden the molecular understanding of mechanisms deployed by bacteria to survive.

TSLP signaling pathway map: a platform for analysis of TSLP-mediated signaling.

Science.gov (United States)

Zhong, Jun; Sharma, Jyoti; Raju, Rajesh; Palapetta, Shyam Mohan; Prasad, T S Keshava; Huang, Tai-Chung; Yoda, Akinori; Tyner, Jeffrey W; van Bodegom, Diederik; Weinstock, David M; Ziegler, Steven F; Pandey, Akhilesh

2014-01-01

Thymic stromal lymphopoietin (TSLP) is a four-helix bundle cytokine that plays a critical role in the regulation of immune responses and in the differentiation of hematopoietic cells. TSLP signals through a heterodimeric receptor complex consisting of an interleukin-7 receptor α chain and a unique TSLP receptor (TSLPR) [also known as cytokine receptor-like factor 2 (CRLF2)]. Cellular targets of TSLP include dendritic cells, B cells, mast cells, regulatory T (Treg) cells and CD4+ and CD8+ T cells. The TSLP/TSLPR axis can activate multiple signaling transduction pathways including the JAK/STAT pathway and the PI-3 kinase pathway. Aberrant TSLP/TSLPR signaling has been associated with a variety of human diseases including asthma, atopic dermatitis, nasal polyposis, inflammatory bowel disease, eosinophilic eosophagitis and, most recently, acute lymphoblastic leukemia. A centralized resource of the TSLP signaling pathway cataloging signaling events is not yet available. In this study, we present a literature-annotated resource of reactions in the TSLP signaling pathway. This pathway map is publicly available through NetPath (http://www.netpath.org/), an open access signal transduction pathway resource developed previously by our group. This map includes 236 molecules and 252 reactions that are involved in TSLP/TSLPR signaling pathway. We expect that the TSLP signaling pathway map will provide a rich resource to study the biology of this important cytokine as well as to identify novel therapeutic targets for diseases associated with dysregulated TSLP/TSLPR signaling. Database URL: http://www.netpath.org/pathways?path_id=NetPath_24.

Increased sensitivity of thyroid hormone-mediated signaling despite prolonged fasting.

Science.gov (United States)

Martinez, Bridget; Scheibner, Michael; Soñanez-Organis, José G; Jaques, John T; Crocker, Daniel E; Ortiz, Rudy M

2017-10-01

Thyroid hormones (TH) can increase cellular metabolism. Food deprivation in mammals is typically associated with reduced thyroid gland responsiveness, in an effort to suppress cellular metabolism and abate starvation. However, in prolonged-fasted, elephant seal pups, cellular TH-mediated proteins are up-regulated and TH levels are maintained with fasting duration. The function and contribution of the thyroid gland to this apparent paradox is unknown and physiologically perplexing. Here we show that the thyroid gland remains responsive during prolonged food deprivation, and that its function and production of TH increase with fasting duration in elephant seals. We discovered that our modeled plasma TH data in response to exogenous thyroid stimulating hormone predicted cellular signaling, which was corroborated independently by the enzyme expression data. The data suggest that the regulation and function of the thyroid gland in the northern elephant seal is atypical for a fasted animal, and can be better described as, "adaptive fasting". Furthermore, the modeling data help substantiate the in vivo responses measured, providing unique insight on hormone clearance, production rates, and thyroid gland responsiveness. Because these unique endocrine responses occur simultaneously with a nearly strict reliance on the oxidation of lipid, these findings provide an intriguing model to better understand the TH-mediated reliance on lipid metabolism that is not otherwise present in morbidly obese humans. When coupled with cellular, tissue-specific responses, these data provide a more integrated assessment of thyroidal status that can be extrapolated for many fasting/food deprived mammals. Copyright © 2017 Elsevier Inc. All rights reserved.

Enhanced NMDA receptor-mediated intracellular calcium signaling in magnocellular neurosecretory neurons in heart failure rats.

Science.gov (United States)

Stern, Javier E; Potapenko, Evgeniy S

2013-08-15

An enhanced glutamate excitatory function within the hypothalamic supraoptic and paraventricluar nuclei is known to contribute to increased neurosecretory and presympathetic neuronal activity, and hence, neurohumoral activation, during heart failure (HF). Still, the precise mechanisms underlying enhanced glutamate-driven neuronal activity in HF remain to be elucidated. Here, we performed simultaneous electrophysiology and fast confocal Ca²⁺ imaging to determine whether altered N-methyl-d-aspartate (NMDA) receptor-mediated changes in intracellular Ca²⁺ levels (NMDA-ΔCa²⁺) occurred in hypothalamic magnocellular neurosecretory cells (MNCs) in HF rats. We found that activation of NMDA receptors resulted in a larger ΔCa²⁺ in MNCs from HF when compared with sham rats. The enhanced NMDA-ΔCa²⁺ was neither dependent on the magnitude of the NMDA-mediated current (voltage clamp) nor on the degree of membrane depolarization or firing activity evoked by NMDA (current clamp). Differently from NMDA receptor activation, firing activity evoked by direct membrane depolarization resulted in similar changes in intracellular Ca²⁺ in sham and HF rats. Taken together, our results support a relatively selective alteration of intracellular Ca²⁺ homeostasis and signaling following activation of NMDA receptors in MNCs during HF. The downstream functional consequences of such altered ΔCa²⁺ signaling during HF are discussed.

Phytomelatonin receptor PMTR1-mediated signaling regulates stomatal closure in Arabidopsis thaliana.

Science.gov (United States)

Wei, Jian; Li, Dong-Xu; Zhang, Jia-Rong; Shan, Chi; Rengel, Zed; Song, Zhong-Bang; Chen, Qi

2018-04-27

Melatonin has been detected in plants in 1995; however, the function and signaling pathway of this putative phytohormone are largely undetermined due to a lack of knowledge about its receptor. Here, we discovered the first phytomelatonin receptor (CAND2/PMTR1) in Arabidopsis thaliana and found that melatonin governs the receptor-dependent stomatal closure. The application of melatonin induced stomatal closure through the heterotrimeric G protein α subunit-regulated H 2 O 2 and Ca 2+ signals. The Arabidopsis mutant lines lacking AtCand2 that encodes a candidate G protein-coupled receptor were insensitive to melatonin-induced stomatal closure. Accordingly, the melatonin-induced H 2 O 2 production and Ca 2+ influx were completely abolished in cand2. CAND2 is a membrane protein that interacts with GPA1 and the expression of AtCand2 was tightly regulated by melatonin in various organs and guard cells. CAND2 showed saturable and specific 125 I-melatonin binding, with apparent K d (dissociation constant) of 0.73 ± 0.10 nmol/L (r 2  = .99), demonstrating this protein is a phytomelatonin receptor (PMTR1). Our results suggest that the phytomelatonin regulation of stomatal closure is dependent on its receptor CAND2/PMTR1-mediated H 2 O 2 and Ca 2+ signaling transduction cascade. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Cancer-related marketing centrality motifs acting as pivot units in the human signaling network and mediating cross-talk between biological pathways.

Science.gov (United States)

Li, Wan; Chen, Lina; Li, Xia; Jia, Xu; Feng, Chenchen; Zhang, Liangcai; He, Weiming; Lv, Junjie; He, Yuehan; Li, Weiguo; Qu, Xiaoli; Zhou, Yanyan; Shi, Yuchen

2013-12-01

Network motifs in central positions are considered to not only have more in-coming and out-going connections but are also localized in an area where more paths reach the networks. These central motifs have been extensively investigated to determine their consistent functions or associations with specific function categories. However, their functional potentials in the maintenance of cross-talk between different functional communities are unclear. In this paper, we constructed an integrated human signaling network from the Pathway Interaction Database. We identified 39 essential cancer-related motifs in central roles, which we called cancer-related marketing centrality motifs, using combined centrality indices on the system level. Our results demonstrated that these cancer-related marketing centrality motifs were pivotal units in the signaling network, and could mediate cross-talk between 61 biological pathways (25 could be mediated by one motif on average), most of which were cancer-related pathways. Further analysis showed that molecules of most marketing centrality motifs were in the same or adjacent subcellular localizations, such as the motif containing PI3K, PDK1 and AKT1 in the plasma membrane, to mediate signal transduction between 32 cancer-related pathways. Finally, we analyzed the pivotal roles of cancer genes in these marketing centrality motifs in the pathogenesis of cancers, and found that non-cancer genes were potential cancer-related genes.

Novel water-soluble curcumin derivative mediating erectile signaling.

Science.gov (United States)

Abdel Aziz, Mohamed Talaat; El Asmer, Mohammed F; Rezq, Ameen; Kumosani, Taha Abdullah; Mostafa, Samya; Mostafa, Taymour; Atta, Hazem; Abdel Aziz Wassef, Mohamed; Fouad, Hanan H; Rashed, Laila; Sabry, Dina; Hassouna, Amira A; Senbel, Amira; Abdel Aziz, Ahmed

2010-08-01

Curcumin is an inducer of heme oxygenase enzyme-1 (HO-1) that is involved in erectile signaling via elevating cyclic guanosine monophosphate (cGMP)levels. To assess the effect of oral administration of a water-soluble long-acting curcumin derivative on erectile signaling. Two hundred and thirty six male white albino rats were divided into four groups; group 1 (N = 20) includes control. Group 2 (N = 72) was equally divided into four subgroups; subgroup 1 received pure curcumin (10 mg/kg), subgroup 2 received the long-acting curcumin derivative (2 mg/kg), subgroup 3 received the long-acting curcumin derivative (10 mg/kg), and subgroup 4 received sildenafil (4 mg/kg). Subgroups were sacrificed after the first, second, and third hour. Group 3 (N = 72) was equally divided into the same four subgroups already mentioned and were sacrificed after 24 hours, 48 hours, and 1 week. Group 4 (N = 72) was subjected to intracavernosal pressure (ICP) measurements 1 hour following oral administration of the same previous doses in the same rat subgroups. Cavernous tissue HO enzyme activity, cGMP, and ICP. In group 2, there was a significant progressive maintained elevation of HO activity and cGMP tissue levels starting from the first hour in subgroups 3 and 4, whereas, the rise in HO activity and cGMP started from second hour regarding the other rat subgroups. Sildenafil effect decreased after 3 hours. In group 3, there was a significant maintained elevation of HO activity and cGMP tissue levels extended to 1 week as compared to controls for all rat subgroups that received both forms of curcumin. In group 4, long-acting curcumin derivative exhibited more significant potentiation of intracavernosal pressure as compared to control and to the pure curcumin. Water-soluble long-acting curcumin derivative could mediate erectile function via upregulating cavernous tissue cGMP. © 2009 International Society for Sexual Medicine.

Hypoxia induces cancer-associated cAMP/PKA signalling through HIF-mediated transcriptional control of adenylyl cyclases VI and VII.

Science.gov (United States)

Simko, Veronika; Iuliano, Filippo; Sevcikova, Andrea; Labudova, Martina; Barathova, Monika; Radvak, Peter; Pastorekova, Silvia; Pastorek, Jaromir; Csaderova, Lucia

2017-08-31

Hypoxia is a phenomenon often arising in solid tumours, linked to aggressive malignancy, bad prognosis and resistance to therapy. Hypoxia-inducible factor-1 has been identified as a key mediator of cell and tissue adaptation to hypoxic conditions through transcriptional activation of many genes involved in glucose metabolism and other cancer-related processes, such as angiogenesis, cell survival and cell invasion. Cyclic adenosine 3'5'-monophosphate is one of the most ancient and evolutionarily conserved signalling molecules and the cAMP/PKA signalling pathway plays an important role in cellular adaptation to hypoxia. We have investigated possible new mechanisms behind hypoxic activation of the cAMP/PKA pathway. For the first time, we have shown that hypoxia induces transcriptional up-regulation of the system of adenylyl cyclases, enzymes responsible for cAMP production, in a panel of carcinoma cell lines of various origin. Our data prove functional relevance of the hypoxic increase of adenylyl cyclases VI and VII at least partially mediated by HIF-1 transcription factor. We have identified adenylyl cyclase VI and VII isoforms as mediators of cellular response to hypoxia, which led to the elevation of cAMP levels and enhanced PKA activity, with an impact on cell migration and pH regulation.

The role of Ca(2+) mediated signaling pathways on the effect of taurine against Streptococcus uberis infection.

Science.gov (United States)

Dai, Bin; Zhang, Jinqiu; Liu, Ming; Lu, Jinye; Zhang, Yuanshu; Xu, Yuanyuan; Miao, Jinfeng; Yin, Yulong

2016-08-30

To provide insight into the mechanisms of taurine attenuation of pro-inflammatory response in mouse mammary epithelial cell line (EpH4-Ev, purchased by ATCC, USA) after Streptococcus uberis (S. uberis, 0140J) challenge, we infected MECs with S. uberis (2.5×10(7)cfumL(-1), MOI=10) for 3h and quantified changes in TLR-2 and calcium (Ca(2+)) mediated signaling pathways. The results indicate that S. uberis infection significantly increases the expression of TLR-2, intracellular Ca(2+) levels, PLC-γ1 and PKC-α, the activities of transcription factors NF-κB and NFAT, and related cytokines (TNF-α, IL-1β, IL-6, G-CSF, IL-2, KC, IL-15, FasL, MCP-1, and LIX) in culture supernatants. Taurine administration downregulated all these indices, the activities of NF-κB and NFAT. Cytokine secretions were similar using special PKC inhibitor Go 6983 and NFAT inhibitor VIVIT. Our data indicate that S. uberis infection induces pro-inflammatory response of MECs through a TLR-2 mediated signaling pathway. In addition, taurine can prevent MEC damage by affecting both PLC-γ1-Ca(2+)-PKC-α-NF-κB and PLC-γ1-Ca(2+)-NFATs signaling pathways. This is the first report to demonstrate the mechanisms of taurine attenuated pro-inflammatory response in MECs after S. uberis challenge. Copyright © 2016 Elsevier B.V. All rights reserved.

Inhibition of fibroblast growth by Notch1 signaling is mediated by induction of Wnt11-dependent WISP-1.

Directory of Open Access Journals (Sweden)

Zhao-Jun Liu

Full Text Available Fibroblasts are an integral component of stroma and important source of growth factors and extracellular matrix (ECM. They play a prominent role in maintaining tissue homeostasis and in wound healing and tumor growth. Notch signaling regulates biological function in a variety of cells. To elucidate the physiological function of Notch signaling in fibroblasts, we ablated Notch1 in mouse (Notch1(Flox/Flox embryonic fibroblasts (MEFs. Notch1-deficient (Notch1(-/- MEFs displayed faster growth and motility rate compared to Notch1(Flox/Flox MEFs. Such phenotypic changes, however, were reversible by reconstitution of Notch1 activation via overexpression of the intracellular domain of Notch1 (NICD1 in Notch1-deficient MEFs. In contrast, constitutive activation of Notch1 signaling by introducing NICD1 into primary human dermal fibroblasts (FF2441, which caused pan-Notch activation, inhibited cell growth and motility, whereas cellular inhibition was relievable when the Notch activation was countered with dominant-negative mutant of Master-mind like 1 (DN-MAML-1. Functionally, "Notch-activated" stromal fibroblasts could inhibit tumor cell growth/invasion. Moreover, Notch activation induced expression of Wnt-induced secreted proteins-1 (WISP-1/CCN4 in FF2441 cells while deletion of Notch1 in MEFs resulted in an opposite effect. Notably, WISP-1 suppressed fibroblast proliferation, and was responsible for mediating Notch1's inhibitory effect since siRNA-mediated blockade of WISP-1 expression could relieve cell growth inhibition. Notch1-induced WISP-1 expression appeared to be Wnt11-dependent, but Wnt1-independent. Blockade of Wnt11 expression resulted in decreased WISP-1 expression and liberated Notch-induced cell growth inhibition. These findings indicated that inhibition of fibroblast proliferation by Notch pathway activation is mediated, at least in part, through regulating Wnt1-independent, but Wnt11-dependent WISP-1 expression.

Phospholipase D1 mediates AMP-activated protein kinase signaling for glucose uptake.

Directory of Open Access Journals (Sweden)

Jong Hyun Kim

2010-03-01

Full Text Available Glucose homeostasis is maintained by a balance between hepatic glucose production and peripheral glucose utilization. In skeletal muscle cells, glucose utilization is primarily regulated by glucose uptake. Deprivation of cellular energy induces the activation of regulatory proteins and thus glucose uptake. AMP-activated protein kinase (AMPK is known to play a significant role in the regulation of energy balances. However, the mechanisms related to the AMPK-mediated control of glucose uptake have yet to be elucidated.Here, we found that AMPK-induced phospholipase D1 (PLD1 activation is required for (14C-glucose uptake in muscle cells under glucose deprivation conditions. PLD1 activity rather than PLD2 activity is significantly enhanced by glucose deprivation. AMPK-wild type (WT stimulates PLD activity, while AMPK-dominant negative (DN inhibits it. AMPK regulates PLD1 activity through phosphorylation of the Ser-505 and this phosphorylation is increased by the presence of AMP. Furthermore, PLD1-S505Q, a phosphorylation-deficient mutant, shows no changes in activity in response to glucose deprivation and does not show a significant increase in (14C-glucose uptake when compared to PLD1-WT. Taken together, these results suggest that phosphorylation of PLD1 is important for the regulation of (14C-glucose uptake. In addition, extracellular signal-regulated kinase (ERK is stimulated by AMPK-induced PLD1 activation through the formation of phosphatidic acid (PA, which is a product of PLD. An ERK pharmacological inhibitor, PD98059, and the PLD inhibitor, 1-BtOH, both attenuate (14C-glucose uptake in muscle cells. Finally, the extracellular stresses caused by glucose deprivation or aminoimidazole carboxamide ribonucleotide (AICAR; AMPK activator regulate (14C-glucose uptake and cell surface glucose transport (GLUT 4 through ERK stimulation by AMPK-mediated PLD1 activation.These results suggest that AMPK-mediated PLD1 activation is required for (14C

The orphan receptor ALK7 and the Activin receptor ALK4 mediate signaling by Nodal proteins during vertebrate development

Science.gov (United States)

Reissmann, Eva; Jörnvall, Henrik; Blokzijl, Andries; Andersson, Olov; Chang, Chenbei; Minchiotti, Gabriella; Persico, M. Graziella; Ibáñez, Carlos F.; Brivanlou, Ali H.

2001-01-01

Nodal proteins have crucial roles in mesendoderm formation and left–right patterning during vertebrate development. The molecular mechanisms of signal transduction by Nodal and related ligands, however, are not fully understood. In this paper, we present biochemical and functional evidence that the orphan type I serine/threonine kinase receptor ALK7 acts as a receptor for mouse Nodal and Xenopus Nodal-related 1 (Xnr1). Receptor reconstitution experiments indicate that ALK7 collaborates with ActRIIB to confer responsiveness to Xnr1 and Nodal. Both receptors can independently bind Xnr1. In addition, Cripto, an extracellular protein genetically implicated in Nodal signaling, can independently interact with both Xnr1 and ALK7, and its expression greatly enhances the ability of ALK7 and ActRIIB to respond to Nodal ligands. The Activin receptor ALK4 is also able to mediate Nodal signaling but only in the presence of Cripto, with which it can also interact directly. A constitutively activated form of ALK7 mimics the mesendoderm-inducing activity of Xnr1 in Xenopus embryos, whereas a dominant-negative ALK7 specifically blocks the activities of Nodal and Xnr1 but has little effect on other related ligands. In contrast, a dominant-negative ALK4 blocks all mesoderm-inducing ligands tested, including Nodal, Xnr1, Xnr2, Xnr4, and Activin. In agreement with a role in Nodal signaling, ALK7 mRNA is localized to the ectodermal and organizer regions of Xenopus gastrula embryos and is expressed during early stages of mouse embryonic development. Therefore, our results indicate that both ALK4 and ALK7 can mediate signal transduction by Nodal proteins, although ALK7 appears to be a receptor more specifically dedicated to Nodal signaling. PMID:11485994

Nuclear translocation of glutathione S-transferase {pi} is mediated by a non-classical localization signal

Energy Technology Data Exchange (ETDEWEB)

Kawakatsu, Miho [Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan); Goto, Shinji, E-mail: sgoto@nagasaki-u.ac.jp [Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan); Yoshida, Takako; Urata, Yoshishige; Li, Tao-Sheng [Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan)

2011-08-12

Highlights: {yields} Nuclear translocation of GST{pi} is abrogated by the deletion of the last 16 amino acid residues in the carboxy-terminal region, indicating that residues 195-208 of GST{pi} are required for nuclear translocation. {yields} The lack of a contiguous stretch of positively charged amino acid residues within the carboxy-terminal region of GST{pi}, suggests that the nuclear translocation of GST{pi} is mediated by a non-classical nuclear localization signal. {yields} An in vitro transport assay shows that the nuclear translocation of GST{pi} is dependent on cytosolic factors and ATP. -- Abstract: Glutathione S-transferase {pi} (GST{pi}), a member of the GST family of multifunctional enzymes, is highly expressed in human placenta and involved in the protection of cellular components against electrophilic compounds or oxidative stress. We have recently found that GST{pi} is expressed in the cytoplasm, mitochondria, and nucleus in some cancer cells, and that the nuclear expression of GST{pi} appears to correlate with resistance to anti-cancer drugs. Although the mitochondrial targeting signal of GST{pi} was previously identified in the amino-terminal region, the mechanism of nuclear translocation remains completely unknown. In this study, we find that the region of GST{pi}195-208 is critical for nuclear translocation, which is mediated by a novel and non-classical nuclear localization signal. In addition, using an in vitro transport assay, we demonstrate that the nuclear translocation of GST{pi} depends on the cytosolic extract and ATP. Although further experiments are needed to understand in depth the precise mechanism of nuclear translocation of GST{pi}, our results may help to establish more efficient anti-cancer therapy, especially with respect to resistance to anti-cancer drugs.

Reactive oxygen species mediate nitric oxide production through ERK/JNK MAPK signaling in HAPI microglia after PFOS exposure

Energy Technology Data Exchange (ETDEWEB)

Wang, Cheng; Nie, Xiaoke; Zhang, Yan [Department of Nutrition and Food Hygiene, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Li, Ting; Mao, Jiamin [Department of Labor and Environmental Hygiene, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Liu, Xinhang [Department of Occupational Medicine and Environmental Toxicology, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Gu, Yiyang; Shi, Jiyun [Department of Labor and Environmental Hygiene, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Xiao, Jing [Department of Occupational Medicine and Environmental Toxicology, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Wan, Chunhua [Department of Nutrition and Food Hygiene, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Wu, Qiyun, E-mail: wqy@ntu.edu.cn [Department of Nutrition and Food Hygiene, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China)

2015-10-15

Perfluorooctane sulfonate (PFOS), an emerging persistent contaminant that is commonly encountered during daily life, has been shown to exert toxic effects on the central nervous system (CNS). However, the molecular mechanisms underlying the neurotoxicity of PFOS remain largely unknown. It has been widely acknowledged that the inflammatory mediators released by hyper-activated microglia play vital roles in the pathogenesis of various neurological diseases. In the present study, we examined the impact of PFOS exposure on microglial activation and the release of proinflammatory mediators, including nitric oxide (NO) and reactive oxidative species (ROS). We found that PFOS exposure led to concentration-dependent NO and ROS production by rat HAPI microglia. We also discovered that there was rapid activation of the ERK/JNK MAPK signaling pathway in the HAPI microglia following PFOS treatment. Moreover, the PFOS-induced iNOS expression and NO production were attenuated after the inhibition of ERK or JNK MAPK by their corresponding inhibitors, PD98059 and SP600125. Interestingly, NAC, a ROS inhibitor, blocked iNOS expression, NO production, and activation of ERK and JNK MAPKs, which suggested that PFOS-mediated microglial NO production occurs via a ROS/ERK/JNK MAPK signaling pathway. Finally, by exposing SH-SY5Y cells to PFOS-treated microglia-conditioned medium, we demonstrated that NO was responsible for PFOS-mediated neuronal apoptosis. - Highlights: • PFOS exposure induced expression of iNOS and production of NO in HAPI microglia. • PFOS induced the production of ROS in HAPI microglia. • ERK/JNK MAPK pathways were activated following PFOS exposure in HAPI microglia. • NO released by HAPI microglia participated in the apoptosis of SH-SY5Y cells.

Reactive oxygen species mediate nitric oxide production through ERK/JNK MAPK signaling in HAPI microglia after PFOS exposure

International Nuclear Information System (INIS)

Wang, Cheng; Nie, Xiaoke; Zhang, Yan; Li, Ting; Mao, Jiamin; Liu, Xinhang; Gu, Yiyang; Shi, Jiyun; Xiao, Jing; Wan, Chunhua; Wu, Qiyun

2015-01-01

Perfluorooctane sulfonate (PFOS), an emerging persistent contaminant that is commonly encountered during daily life, has been shown to exert toxic effects on the central nervous system (CNS). However, the molecular mechanisms underlying the neurotoxicity of PFOS remain largely unknown. It has been widely acknowledged that the inflammatory mediators released by hyper-activated microglia play vital roles in the pathogenesis of various neurological diseases. In the present study, we examined the impact of PFOS exposure on microglial activation and the release of proinflammatory mediators, including nitric oxide (NO) and reactive oxidative species (ROS). We found that PFOS exposure led to concentration-dependent NO and ROS production by rat HAPI microglia. We also discovered that there was rapid activation of the ERK/JNK MAPK signaling pathway in the HAPI microglia following PFOS treatment. Moreover, the PFOS-induced iNOS expression and NO production were attenuated after the inhibition of ERK or JNK MAPK by their corresponding inhibitors, PD98059 and SP600125. Interestingly, NAC, a ROS inhibitor, blocked iNOS expression, NO production, and activation of ERK and JNK MAPKs, which suggested that PFOS-mediated microglial NO production occurs via a ROS/ERK/JNK MAPK signaling pathway. Finally, by exposing SH-SY5Y cells to PFOS-treated microglia-conditioned medium, we demonstrated that NO was responsible for PFOS-mediated neuronal apoptosis. - Highlights: • PFOS exposure induced expression of iNOS and production of NO in HAPI microglia. • PFOS induced the production of ROS in HAPI microglia. • ERK/JNK MAPK pathways were activated following PFOS exposure in HAPI microglia. • NO released by HAPI microglia participated in the apoptosis of SH-SY5Y cells.

Supplementary Material for: Promoter hypermethylation of HS3ST2, SEPTIN9 and SLIT2 combined with FGFR3 mutations as a sensitive/specific urinary assay for diagnosis and surveillance in patients with low or high-risk non-muscle-invasive bladder cancer

KAUST Repository

Roperch, Jean-Pierre; Grandchamp, Bernard; Desgrandchamps, Franç ois; Mongiat-Artus, Pierre; Ravery, Vincent; Ouzaid, Idir; Roupret, Morgan; Phe, Vé ronique; Ciofu, Calin; Tubach, Florence; Cussenot, Olivier; Incitti, Roberto

2016-01-01

Abstract Background Non-muscle-invasive bladder cancer (NMIBC) is a high incidence form of bladder cancer (BCa), where genetic and epigenetic alterations occur frequently. We assessed the performance of associating a FGFR3 mutation assay and a DNA methylation analysis to improve bladder cancer detection and to predict disease recurrence of NMIBC patients. Methods We used allele specific PCR to determine the FGFR3 mutation status for R248C, S249C, G372C, and Y375C. We preselected 18 candidate genes reported in the literature as being hypermethylated in cancer and measured their methylation levels by quantitative multiplex-methylation specific PCR. We selected HS3ST2, SLIT2 and SEPTIN9 as the most discriminative between control and NMIBC patients and we assayed these markers on urine DNA from a diagnostic study consisting of 167 NMIBC and 105 controls and a follow-up study consisting of 158 NMIBC at diagnosis time’s and 425 at follow-up time. ROC analysis was performed to evaluate the diagnostic accuracy of each assay alone and in combination. Results For Diagnosis: Using a logistic regression analysis with a model consisting of the 3 markers’ methylation values, FGFR3 status, age and known smoker status at the diagnosis time we obtained sensitivity/specificity of 97.6 %/84.8 % and an optimism-corrected AUC of 0.96. With an estimated BCa prevalence of 12.1 % in a hematuria cohort, this corresponds to a negative predictive value (NPV) of 99.6 %. For Follow-up: Using a logistic regression with FGFR3 mutation and the CMI at two time points (beginning of the follow-up and current time point), we got sensitivity/specificity/NPV of 90.3 %/65.1 %/97.0 % and a corrected AUC of 0.84. We also tested a thresholding algorithm with FGFR3 mutation and the two time points as described above, obtaining sensitivity/specificity/NPV values of, respectively, 94.5 %/75.9 %/98.5 % and an AUC of 0.82. Conclusions We showed that combined analysis of FGFR3 mutation and DNA

Mixed Signals: Co-Stimulation in Invariant Natural Killer T Cell-Mediated Cancer Immunotherapy

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Susannah C. Shissler

2017-11-01

Full Text Available Invariant natural killer T (iNKT cells are an integral component of the immune system and play an important role in antitumor immunity. Upon activation, iNKT cells can directly kill malignant cells as well as rapidly produce cytokines that stimulate other immune cells, making them a front line defense against tumorigenesis. Unfortunately, iNKT cell number and activity are reduced in multiple cancer types. This anergy is often associated with upregulation of co-inhibitory markers such as programmed death-1. Similar to conventional T cells, iNKT cells are influenced by the conditions of their activation. Conventional T cells receive signals through the following three types of receptors: (1 T cell receptor (TCR, (2 co-stimulation molecules, and (3 cytokine receptors. Unlike conventional T cells, which recognize peptide antigen presented by MHC class I or II, the TCRs of iNKT cells recognize lipid antigen in the context of the antigen presentation molecule CD1d (Signal 1. Co-stimulatory molecules can positively and negatively influence iNKT cell activation and function and skew the immune response (Signal 2. This study will review the background of iNKT cells and their co-stimulatory requirements for general function and in antitumor immunity. We will explore the impact of monoclonal antibody administration for both blocking inhibitory pathways and engaging stimulatory pathways on iNKT cell-mediated antitumor immunity. This review will highlight the incorporation of co-stimulatory molecules in antitumor dendritic cell vaccine strategies. The use of co-stimulatory intracellular signaling domains in chimeric antigen receptor-iNKT therapy will be assessed. Finally, we will explore the influence of innate-like receptors and modification of immunosuppressive cytokines (Signal 3 on cancer immunotherapy.

Rapid detection of K650E mutation in FGFR3 using uncultured amniocytes in a pregnancy affected with fetal cloverleaf skull, occipital pseudoencephalocele, ventriculomegaly, straight short femurs, and thanatophoric dysplasia type II

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Chih-Ping Chen

2013-09-01

Conclusion: A prenatal diagnosis of cloverleaf skull, short limbs, straight femurs, and occipital pseudoencephalocele should include a differential diagnosis of TD2. A molecular analysis of FGFR3 using uncultured amniocytes is useful for the rapid confirmation of TD2 at prenatal diagnosis.

Genetic interaction of two abscisic acid signaling regulators, HY5 and FIERY1, in mediating lateral root formation

KAUST Repository

Chen, Hao

2011-01-01

Root architecture is continuously shaped in a manner that helps plants to better adapt to the environment. Gene regulation at the transcriptional or post-transcriptional levels largely controls this environmental response. Recently, RNA silencing has emerged as an important player in gene regulation and is involved in many aspects of plant development, including lateral root formation. In a recent study, we found that FIERY1, a bifunctional abiotic stress and abscisic acid (ABA) signaling regulator and an endogenous RNA silencing suppressor, mediates auxin response during lateral root formation in Arabidopsis. We proposed that FRY1 regulates lateral root development through its activity on adenosine 3,5-bisphosphate (PAP), a strong inhibitor of exoribonucleases (XRNs). Interestingly, some of the phenotypes of fry1, such as enhanced response to light in repressing hypocotyl elongation and hypersensitivity to ABA in lateral root growth, are opposite to those of another light- and ABA-signaling mutant, hy5. Here we analyzed the hy5 fry1 double mutant for root and hypocotyl growth. We found that the hy5 mutation can suppress the enhanced light sensitivity in fry1 hypocotyl elongation and restore the lateral root formation. The genetic interaction between HY5 and FRY1 indicates that HY5 and FRY1 may act in overlapping pathways that mediate light signaling and lateral root development. © 2011 Landes Bioscience.

Agonist-mediated activation of Bombyx mori diapause hormone receptor signals to extracellular signal-regulated kinases 1 and 2 through Gq-PLC-PKC-dependent cascade.

Science.gov (United States)

Jiang, Xue; Yang, Jingwen; Shen, Zhangfei; Chen, Yajie; Shi, Liangen; Zhou, Naiming

2016-08-01

Diapause is a developmental strategy adopted by insects to survive in challenging environments such as the low temperatures of a winter. This unique process is regulated by diapause hormone (DH), which is a neuropeptide hormone that induces egg diapause in Bombyx mori and is involved in terminating pupal diapause in heliothis moths. An G protein-coupled receptor from the silkworm, B. mori, has been identified as a specific cell surface receptor for DH. However, the detailed information on the DH-DHR system and its mechanism(s) involved in the induction of embryonic diapause remains unknown. Here, we combined functional assays with various specific inhibitors to elucidate the DHR-mediated signaling pathways. Upon activation by DH, B. mori DHR is coupled to the Gq protein, leading to a significant increase of intracellular Ca(2+) and cAMP response element-driven luciferase activity in an UBO-QIC, a specific Gq inhibitor, sensitive manner. B. mori DHR elicited ERK1/2 phosphorylation in a dose- and time-dependent manner in response to DH. This effect was almost completely inhibited by co-incubation with UBO-QIC and was also significantly suppressed by PLC inhibitor U73122, PKC inhibitors Gö6983 and the Ca(2+) chelator EGTA. Moreover, DHR-induced activation of ERK1/2 was significantly attenuated by treatment with the Gβγ specific inhibitors gallein and M119K and the PI3K specific inhibitor Wortmannin, but not by the Src specific inhibitor PP2. Our data also demonstrates that the EGFR-transactivation pathway is not involved in the DHR-mediated ERK1/2 phosphorylation. Future efforts are needed to clarify the role of the ERK1/2 signaling pathway in the DH-mediated induction of B. mori embryonic diapause. Copyright © 2016 Elsevier Ltd. All rights reserved.

Blockade of IP[subscript 3]-Mediated SK Channel Signaling in the Rat Medial Prefrontal Cortex Improves Spatial Working Memory

Science.gov (United States)

Brennan, Avis R.; Dolinsky, Beth; Vu, Mai-Anh T.; Stanley, Marion; Yeckel, Mark F.; Arnsten, Amy F. T.

2008-01-01

Planning and directing thought and behavior require the working memory (WM) functions of prefrontal cortex. WM is compromised by stress, which activates phosphatidylinositol (PI)-mediated IP[subscript 3]-PKC intracellular signaling. PKC overactivation impairs WM operations and in vitro studies indicate that IP[subscript 3] receptor (IP[subscript…

MEDIATOR18 and MEDIATOR20 confer susceptibility to Fusarium oxysporum in Arabidopsis thaliana

Science.gov (United States)

Stiller, Jiri; Davoine, Celine; Björklund, Stefan; Manners, John M.; Kazan, Kemal; Schenk, Peer M.

2017-01-01

The conserved protein complex known as Mediator conveys transcriptional signals by acting as an intermediary between transcription factors and RNA polymerase II. As a result, Mediator subunits play multiple roles in regulating developmental as well as abiotic and biotic stress pathways. In this report we identify the head domain subunits MEDIATOR18 and MEDIATOR20 as important susceptibility factors for Fusarium oxysporum infection in Arabidopsis thaliana. Mutants of MED18 and MED20 display down-regulation of genes associated with jasmonate signaling and biosynthesis while up-regulation of salicylic acid associated pathogenesis related genes and reactive oxygen producing and scavenging genes. We propose that MED18 and MED20 form a sub-domain within Mediator that controls the balance of salicylic acid and jasmonate associated defense pathways. PMID:28441405

Transmitters and pathways mediating inhibition of spinal itch-signaling neurons by scratching and other counterstimuli.

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Tasuku Akiyama

Full Text Available Scratching relieves itch, but the underlying neural mechanisms are poorly understood. We presently investigated a role for the inhibitory neurotransmitters GABA and glycine in scratch-evoked inhibition of spinal itch-signaling neurons in a mouse model of chronic dry skin itch. Superficial dorsal horn neurons ipsilateral to hindpaw dry skin treatment exhibited a high level of spontaneous firing that was significantly attenuated by cutaneous scratching, pinch and noxious heat. Scratch-evoked inhibition was nearly abolished by spinal delivery of the glycine antagonist, strychnine, and was markedly attenuated by respective GABA(A and GABA(B antagonists bicuculline and saclofen. Scratch-evoked inhibition was also significantly attenuated (but not abolished by interruption of the upper cervical spinal cord, indicating the involvement of both segmental and suprasegmental circuits that engage glycine- and GABA-mediated inhibition of spinal itch-signaling neurons by noxious counterstimuli.

Heterotrimeric G proteins-mediated resistance to necrotrophic pathogens includes mechanisms independent of salicylic acid-, jasmonic acid/ethylene- and abscisic acid-mediated defense signaling.