The CytR repressor antagonizes cyclic AMP-cyclic AMP receptor protein activation of the deoCp2 promoter of Escherichia coli K-12

DEFF Research Database (Denmark)

Søgaard-Andersen, L; Martinussen, J; Møllegaard, N E

1990-01-01

We have investigated the regulation of the Escherichia coli deoCp2 promoter by the CytR repressor and the cyclic AMP (cAMP) receptor protein (CRP) complexed to cAMP. Promoter regions controlled by these two proteins characteristically contain tandem cAMP-CRP binding sites. Here we show that (i) Cyt...

Screening of the transcriptional regulatory regions of vascular endothelial growth factor receptor 2 (VEGFR2 in amyotrophic lateral sclerosis

Directory of Open Access Journals (Sweden)

Hartley Judith

2007-04-01

Full Text Available Abstract Background Vascular endothelial growth factor (VEGF has neurotrophic activity which is mediated by its main agonist receptor, VEGFR2. Dysregulation of VEGF causes motor neurone degeneration in a mouse model of amyotrophic lateral sclerosis (ALS, and expression of VEGFR2 is reduced in motor neurones and spinal cord of patients with ALS. Methods We have screened the promoter region and 4 exonic regions of functional significance of the VEGFR2 gene in a UK population of patients with ALS, for mutations and polymorphisms that may affect expression or function of this VEGF receptor. Results No mutations were identified in the VEGFR2 gene. We found no association between polymorphisms in the regulatory regions of the VEGFR2 gene and ALS. Conclusion Mechanisms other than genetic variation may downregulate expression or function of the VEGFR2 receptor in patients with ALS.

Dual hypocretin receptor antagonism is more effective for sleep promotion than antagonism of either receptor alone.

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Stephen R Morairty

Full Text Available The hypocretin (orexin system is involved in sleep/wake regulation, and antagonists of both hypocretin receptor type 1 (HCRTR1 and/or HCRTR2 are considered to be potential hypnotic medications. It is currently unclear whether blockade of either or both receptors is more effective for promoting sleep with minimal side effects. Accordingly, we compared the properties of selective HCRTR1 (SB-408124 and SB-334867 and HCRTR2 (EMPA antagonists with that of the dual HCRTR1/R2 antagonist almorexant in the rat. All 4 antagonists bound to their respective receptors with high affinity and selectivity in vitro. Since in vivo pharmacokinetic experiments revealed poor brain penetration for SB-408124, SB-334867 was selected for subsequent in vivo studies. When injected in the mid-active phase, SB-334867 produced small increases in rapid-eye-movement (REM and non-REM (NR sleep. EMPA produced a significant increase in NR only at the highest dose studied. In contrast, almorexant decreased NR latency and increased both NR and REM proportionally throughout the subsequent 6 h without rebound wakefulness. The increased NR was due to a greater number of NR bouts; NR bout duration was unchanged. At the highest dose tested (100 mg/kg, almorexant fragmented sleep architecture by increasing the number of waking and REM bouts. No evidence of cataplexy was observed. HCRTR1 occupancy by almorexant declined 4-6 h post-administration while HCRTR2 occupancy was still elevated after 12 h, revealing a complex relationship between occupancy of HCRT receptors and sleep promotion. We conclude that dual HCRTR1/R2 blockade is more effective in promoting sleep than blockade of either HCRTR alone. In contrast to GABA receptor agonists which induce sleep by generalized inhibition, HCRTR antagonists seem to facilitate sleep by reducing waking "drive".

Functionality of promoter microsatellites of arginine vasopressin receptor 1A (AVPR1A): implications for autism

LENUS (Irish Health Repository)

Tansey, Katherine E

2011-03-31

Abstract Background Arginine vasopressin (AVP) has been hypothesized to play a role in aetiology of autism based on a demonstrated involvement in the regulation of social behaviours. The arginine vasopressin receptor 1A gene (AVPR1A) is widely expressed in the brain and is considered to be a key receptor for regulation of social behaviour. Moreover, genetic variation at AVPR1A has been reported to be associated with autism. Evidence from non-human mammals implicates variation in the 5\\'-flanking region of AVPR1A in variable gene expression and social behaviour. Methods We examined four tagging single nucleotide polymorphisms (SNPs) (rs3803107, rs1042615, rs3741865, rs11174815) and three microsatellites (RS3, RS1 and AVR) at the AVPR1A gene for association in an autism cohort from Ireland. Two 5\\'-flanking region polymorphisms in the human AVPR1A, RS3 and RS1, were also tested for their effect on relative promoter activity. Results The short alleles of RS1 and the SNP rs11174815 show weak association with autism in the Irish population (P = 0.036 and P = 0.008, respectively). Both RS1 and RS3 showed differences in relative promoter activity by length. Shorter repeat alleles of RS1 and RS3 decreased relative promoter activity in the human neuroblastoma cell line SH-SY5Y. Conclusions These aligning results can be interpreted as a functional route for this association, namely that shorter alleles of RS1 lead to decreased AVPR1A transcription, which may proffer increased susceptibility to the autism phenotype.

Opioid receptors in midbrain dopaminergic regions of the rat. 1. Mu receptor autoradiography

International Nuclear Information System (INIS)

German, D.C.; Speciale, S.G.; Manaye, K.F.; Sadeq, M.

1993-01-01

Several lines of evidence indicate that an interaction exists between opioid peptides and midbrain dopaminergic neurons. The purpose of this study was to map and quantify the density of the mu opioid receptor subtype relative to the location of the dopaminergic (DA) neurons in the retrorubral field (nucleus A8), substantia nigra (nucleus A9), and ventral tegmental area and related nuclei (nucleus A10) in the rat. Sections through the rostral-caudal extent of the midbrain were stained with an antibody against tyrosine hydroxylase, as a DA cell marker, and comparable sections were processed for in vitro receptor autoradiography using the mu-selective ligand, 3 H-Tyr-D-Ala-N-MePhe-Gyl-ol enkephalin. In the nucleus A8 region, there were low levels of mu binding. In the rostral portion of nucleus A9, there was prominent mu binding both in the ventral pars compacta, which contains numerous DA neurons, and in regions that correspond to the location of the DA dendrites which project ventrally into the underlying substantia nigra pars reticulata. In the caudal portion of nucleus A9, mu binding was greatest in the substantia nigra pars reticulata, but also in the same region that contains DA neurons. In nucleus A10, mu receptor densities differed depending upon the nucleus A10 subdivision, and the rostral-caudal position in the nucleus. Low receptor densities were observed in rostral portions of the ventral tegmental area and interfascicular nucleus, and there was negligible binding in the parabrachial pigmented nucleus and paranigral nucleus at the level of the interpeduncular nucleus; all regions where there are high densities of DA somata. Mu binding was relatively high in the central linear nucleus, and in the dorsal and medial divisions of the medial terminal nucleus of the accessory optic system, which has been shown to contain DA dendrites. These data indicate that mu opioid receptors are located in certain regions occupied by all three midbrain DA nuclei, but in a

Cell-Type-Specific Regulation of the Retinoic Acid Receptor Mediated by the Orphan Nuclear Receptor TLX†

Science.gov (United States)

Kobayashi, Mime; Yu, Ruth T.; Yasuda, Kunio; Umesono, Kazuhiko

2000-01-01

Malformations in the eye can be caused by either an excess or deficiency of retinoids. An early target gene of the retinoid metabolite, retinoic acid (RA), is that encoding one of its own receptors, the retinoic acid receptor β (RARβ). To better understand the mechanisms underlying this autologous regulation, we characterized the chick RARβ2 promoter. The region surrounding the transcription start site of the avian RARβ2 promoter is over 90% conserved with the corresponding region in mammals and confers strong RA-dependent transactivation in primary cultured embryonic retina cells. This response is selective for RAR but not retinoid X receptor-specific agonists, demonstrating a principal role for RAR(s) in retina cells. Retina cells exhibit a far higher sensitivity to RA than do fibroblasts or osteoblasts, a property we found likely due to expression of the orphan nuclear receptor TLX. Ectopic expression of TLX in fibroblasts resulted in increased sensitivity to RA induction, an effect that is conserved between chick and mammals. We have identified a cis element, the silencing element relieved by TLX (SET), within the RARβ2 promoter region which confers TLX- and RA-dependent transactivation. These results indicate an important role for TLX in autologous regulation of the RARβ gene in the eye. PMID:11073974

Cell-type-specific regulation of the retinoic acid receptor mediated by the orphan nuclear receptor TLX.

Science.gov (United States)

Kobayashi, M; Yu, R T; Yasuda, K; Umesono, K

2000-12-01

Malformations in the eye can be caused by either an excess or deficiency of retinoids. An early target gene of the retinoid metabolite, retinoic acid (RA), is that encoding one of its own receptors, the retinoic acid receptor beta (RARbeta). To better understand the mechanisms underlying this autologous regulation, we characterized the chick RARbeta2 promoter. The region surrounding the transcription start site of the avian RARbeta2 promoter is over 90% conserved with the corresponding region in mammals and confers strong RA-dependent transactivation in primary cultured embryonic retina cells. This response is selective for RAR but not retinoid X receptor-specific agonists, demonstrating a principal role for RAR(s) in retina cells. Retina cells exhibit a far higher sensitivity to RA than do fibroblasts or osteoblasts, a property we found likely due to expression of the orphan nuclear receptor TLX. Ectopic expression of TLX in fibroblasts resulted in increased sensitivity to RA induction, an effect that is conserved between chick and mammals. We have identified a cis element, the silencing element relieved by TLX (SET), within the RARbeta2 promoter region which confers TLX- and RA-dependent transactivation. These results indicate an important role for TLX in autologous regulation of the RARbeta gene in the eye.

Dopamine receptor D4 promoter hypermethylation increases the risk of drug addiction.

Science.gov (United States)

Ji, Huihui; Xu, Xuting; Liu, Guili; Liu, Huifen; Wang, Qinwen; Shen, Wenwen; Li, Longhui; Xie, Xiaohu; Hu, Haochang; Xu, Lei; Zhou, Wenhua; Duan, Shiwei

2018-02-01

Heroin and methylamphetamine (METH) are two addictive drugs that cause serious problems for society. Dopamine receptor D4 (DRD4), a key receptor in the dopaminergic system, may facilitate the development of drug addiction. The aim of the present study was to investigate the association between the promoter methylation level of DRD4 gene and drug addiction. Bisulfite pyrosequencing technology was used to measure the methylation levels of DRD4 promoter in 60 drug addicts and 52 matched controls. Significantly higher levels of DRD4 CpG1 and CpG4 methylation were detected in METH and heroin drug addicts compared with controls (Pdrug addiction.

Nucleus Accumbens Shell and mPFC but not Insula Orexin-1 Receptors Promote Excessive Alcohol Drinking

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Kelly Lei

2016-08-01

Full Text Available Addiction to alcohol remains a major social and economic problem, in part because of the high motivation for alcohol that humans exhibit and the hazardous binge intake this promotes. Orexin-1-type receptors (OX1Rs promote reward intake under conditions of strong drives for reward, including excessive alcohol intake. While systemic modulation of OX1Rs can alter alcohol drinking, the brain regions that mediate this OX1R enhancement of excessive drinking remain unknown. Given the importance of the nucleus accumbens (NAc and anterior insular cortex (aINS in driving many addictive behaviors, including OX1Rs within these regions, we examined the importance of OX1Rs in these regions on excessive alcohol drinking in C57BL/6 mice during limited-access alcohol drinking in the dark cycle. Inhibition of OX1Rs with the widely used SB-334867 within the medial NAc Shell (mNAsh significantly reduced drinking of alcohol, with no effect on saccharin intake, and no effect on alcohol consumption when infused above the mNAsh. In contrast, intra-mNAsh infusion of the orexin-2 receptor TCS-OX2-29 had no impact on alcohol drinking. In addition, OX1R inhibition within the aINS had no effect on excessive drinking, which was surprising given the importance of aINS-NAc circuits in promoting alcohol consumption and the role for aINS OX1Rs in driving nicotine intake. However, OX1R inhibition within the mPFC did reduce alcohol drinking, indicating cortical OXR involvement in promoting intake. Also, in support of the critical role for mNAsh OX1Rs, SB within the mNAsh also significantly reduced operant alcohol self-administration in rats. Finally, orexin ex vivo enhanced firing in mNAsh neurons from alcohol-drinking mice, with no effect on evoked EPSCs or input resistance; a similar orexin increase in firing without a change in input resistance was observed in alcohol-naïve mice. Taken together, our results strongly suggest that OX1Rs within the mNAsh, but not the aINS, play a

[Health-Promoting Schools Regional Initiative of the Americas].

Science.gov (United States)

Ippolito-Shepherd, Josefa; Cerqueira, Maria Teresa; Ortega, Diana Patricia

2005-01-01

In Latin America, comprehensive health promotion programmes and activities are being implemented in the school setting, which take into account the conceptual framework of the Health-Promoting Schools Regional Initiative of the Pan American Health Organization, Regional office of the World Health Organization (PAHO/WHO). These programmes help to strengthen the working relationships between the health and education sectors. The Health-Promoting Schools Regional Initiative, officially launched by PAHO/WHO in 1995, aims to form future generations to have the knowledge, abilities, and skills necessary for promoting and caring for their health and that of their family and community, as well as to create and maintain healthy environments and communities. The Initiative focuses on three main components: comprehensive health education, the creation and maintenance of healthy physical and psychosocial environments, and the access to health and nutrition services, mental health, and active life. In 2001, PAHO conducted a survey in 19 Latin American countries to assess the status and trends of Health-Promoting Schools in the Region, for the appropriate regional, subregional, and national planning of pertinent health promotion and health education programmes and activities. The results of this survey provided information about policies and national plans, multisectoral coordination mechanisms for the support of health promotion in the school settings, the formation and participation in national and international networks of Health-Promoting Schools and about the level of dissemination of the strategy. For the successful development of Health-Promoting Schools is essential to involve the society as a whole, in order to mobilise human resources and materials necessary for implementing health promotion in the school settings. Thus, the constitution and consolidation of networks has been a facilitating mechanism for the exchange of ideas, resources and experiences to strengthen

Goat activin receptor type IIB knockdown by muscle specific promoter driven artificial microRNAs.

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Patel, Amrutlal K; Shah, Ravi K; Patel, Utsav A; Tripathi, Ajai K; Joshi, Chaitanya G

2014-10-10

Activin receptor type IIB (ACVR2B) is a transmembrane receptor which mediates signaling of TGF beta superfamily ligands known to function in regulation of muscle mass, embryonic development and reproduction. ACVR2B antagonism has shown to enhance the muscle growth in several disease and transgenic models. Here, we show ACVR2B knockdown by RNA interference using muscle creatine kinase (MCK) promoter driven artificial microRNAs (amiRNAs). Among the various promoter elements tested, the ∼1.26 kb MCK promoter region showed maximum transcriptional activity in goat myoblasts cells. We observed up to 20% silencing in non-myogenic 293T cells and up to 32% silencing in myogenic goat myoblasts by MCK directed amiRNAs by transient transfection. Goat myoblasts stably integrated with MCK directed amiRNAs showed merely 8% silencing in proliferating myoblasts which was increased to 34% upon induction of differentiation at transcript level whereas up to 57% silencing at protein level. Knockdown of ACVR2B by 5'-UTR derived amiRNAs resulted in decreased SMAD2/3 signaling, increased expression of myogenic regulatory factors (MRFs) and enhanced proliferation and differentiation of myoblasts. Unexpectedly, knockdown of ACVR2B by 3'-UTR derived amiRNAs resulted in increased SMAD2/3 signaling, reduced expression of MRFs and suppression of myogenesis. Our study offers muscle specific knockdown of ACVR2B as a potential strategy to enhance muscle mass in the farm animal species. Copyright © 2014 Elsevier B.V. All rights reserved.

Knock-In Mice with NOP-eGFP Receptors Identify Receptor Cellular and Regional Localization.

Science.gov (United States)

Ozawa, Akihiko; Brunori, Gloria; Mercatelli, Daniela; Wu, Jinhua; Cippitelli, Andrea; Zou, Bende; Xie, Xinmin Simon; Williams, Melissa; Zaveri, Nurulain T; Low, Sarah; Scherrer, Grégory; Kieffer, Brigitte L; Toll, Lawrence

2015-08-19

The nociceptin/orphanin FQ (NOP) receptor, the fourth member of the opioid receptor family, is involved in many processes common to the opioid receptors including pain and drug abuse. To better characterize receptor location and trafficking, knock-in mice were created by inserting the gene encoding enhanced green fluorescent protein (eGFP) into the NOP receptor gene (Oprl1) and producing mice expressing a functional NOP-eGFP C-terminal fusion in place of the native NOP receptor. The NOP-eGFP receptor was present in brain of homozygous knock-in animals in concentrations somewhat higher than in wild-type mice and was functional when tested for stimulation of [(35)S]GTPγS binding in vitro and in patch-clamp electrophysiology in dorsal root ganglia (DRG) neurons and hippocampal slices. Inhibition of morphine analgesia was equivalent when tested in knock-in and wild-type mice. Imaging revealed detailed neuroanatomy in brain, spinal cord, and DRG and was generally consistent with in vitro autoradiographic imaging of receptor location. Multicolor immunohistochemistry identified cells coexpressing various spinal cord and DRG cellular markers, as well as coexpression with μ-opioid receptors in DRG and brain regions. Both in tissue slices and primary cultures, the NOP-eGFP receptors appear throughout the cell body and in processes. These knock-in mice have NOP receptors that function both in vitro and in vivo and appear to be an exceptional tool to study receptor neuroanatomy and correlate with NOP receptor function. The NOP receptor, the fourth member of the opioid receptor family, is involved in pain, drug abuse, and a number of other CNS processes. The regional and cellular distribution has been difficult to determine due to lack of validated antibodies for immunohistochemical analysis. To provide a new tool for the investigation of receptor localization, we have produced knock-in mice with a fluorescent-tagged NOP receptor in place of the native NOP receptor. These

The retinoid X receptor response element in the human aldehyde dehydrogenase 2 promoter is antagonized by the chicken ovalbumin upstream promoter family of orphan receptors

NARCIS (Netherlands)

Pinaire, J; Hasanadka, R; Fang, M; Chou, WY; Stewart, MJ; Kruijer, W; Crabb, D

2000-01-01

Two tandem sites in the aldehyde dehydrogenase 2 promoter (designated FP330-5' and FP330-3') that bind members of the nuclear receptor superfamily mere recently identified. Antibodies against apolipoprotein regulatory protein (ARP-1) altered DNA-protein interactions in electrophoretic mobility shift

Activation of dopamine receptors in the nucleus accumbens promotes sucrose-reinforced cued approach behavior

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Saleem M. Nicola

2016-07-01

Full Text Available Dopamine receptor activation in the nucleus accumbens (NAc promotes vigorous environmentally-cued food-seeking in hungry rats. Rats fed ad libitum, however, respond to fewer food-predictive cues, particularly when the value of food reward is low. Here, we investigated whether this difference could be due to differences in the degree of dopamine receptor activation in the NAc. First, we observed that although rats given ad libitum access to chow in their home cages approached a food receptacle in response to reward-predictive cues, the number of such approaches declined as animals accumulated food rewards. Intriguingly, cued approach to food occurred in clusters, with several cued responses followed by successive non-responses. This pattern suggested that behavior was dictated by transitions between two states, responsive and non-responsive. Injection of D1 or D2 dopamine receptor agonists into the NAc dose-dependently increased cue responding by promoting transitions to the responsive state and by preventing transitions to the non-responsive state. In contrast, antagonists of either D1 or D2 receptors promoted long bouts of non-responding by inducing transitions to the non-responsive state and by preventing transitions to the responsive state. Moreover, locomotor behavior during the inter-trial interval was correlated with the responsive state, and was also increased by dopamine receptor agonists. These results suggest that activation of NAc dopamine receptors plays an important role in regulating the probability of approach to food under conditions of normative satiety.

Activation of GLP-1 Receptor Promotes Bone Marrow Stromal Cell Osteogenic Differentiation through β-Catenin

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Jingru Meng

2016-04-01

Full Text Available Glucagon-like peptide 1 (GLP-1 plays an important role in regulating bone remodeling, and GLP-1 receptor agonist shows a positive relationship with osteoblast activity. However, GLP-1 receptor is not found in osteoblast, and the mechanism of GLP-1 receptor agonist on regulating bone remodeling is unclear. Here, we show that the GLP-1 receptor agonist exendin-4 (Ex-4 promoted bone formation and increased bone mass and quality in a rat unloading-induced bone loss model. These functions were accompanied by an increase in osteoblast number and serum bone formation markers, while the adipocyte number was decreased. Furthermore, GLP-1 receptor was detected in bone marrow stromal cells (BMSCs, but not in osteoblast. Activation of GLP-1 receptor by Ex-4 promoted the osteogenic differentiation and inhibited BMSC adipogenic differentiation through regulating PKA/β-catenin and PKA/PI3K/AKT/GSK3β signaling. These findings reveal that GLP-1 receptor regulates BMSC osteogenic differentiation and provide a molecular basis for therapeutic potential of GLP-1 against osteoporosis.

A growth hormone receptor SNP promotes lung cancer by impairment of SOCS2-mediated degradation

DEFF Research Database (Denmark)

Chhabra, Y.; Wong, H. Y.; Nikolajsen, Louise Fletcher

2018-01-01

Both humans and mice lacking functional growth hormone (GH) receptors are known to be resistant to cancer. Further, autocrine GH has been reported to act as a cancer promoter. Here we present the first example of a variant of the GH receptor (GHR) associated with cancer promotion, in this case lu......-mesenchymal transition and metastases (TWIST1, SNAI2, EGFR, MYC and CCND1) at 2 h after a GH pulse. This is consistent with prolonged GH signalling acting to promote cancer progression in lung cancer.Oncogene advance online publication, 2 October 2017; doi:10.1038/onc.2017.352....

Liver Tumor Promotion by 2,3,7,8-Tetrachlorodibenzo-p-dioxin Is Dependent on the Aryl Hydrocarbon Receptor and TNF/IL-1 Receptors

Science.gov (United States)

Kennedy, Gregory D.; Nukaya, Manabu; Moran, Susan M.; Glover, Edward; Weinberg, Samuel; Balbo, Silvia; Hecht, Stephen S.; Pitot, Henry C.; Drinkwater, Norman R.; Bradfield, Christopher A.

2014-01-01

We set out to better understand the signal transduction pathways that mediate liver tumor promotion by 2,3,7,8-tetrachlorodibenzo-p-dioxn (“dioxin”). To this end, we first employed congenic mice homozygous for either the Ahrb1 or Ahrd alleles (encoding an aryl hydrocarbon receptor (AHR) with high or low binding affinity for dioxin, respectively) and demonstrated that hepatocellular tumor promotion in response to dioxin segregated with the Ahr locus. Once we had genetic evidence for the importance of AHR signaling, we then asked if tumor promotion by dioxin was influenced by “interleukin-1 (IL-1)-like” inflammatory cytokines. The importance of this question arose from our earlier observation that aspects of the acute hepatocellular toxicity of dioxin are dependent upon IL1-like cytokine signaling. To address this issue, we employed a triple knock-out (TKO) mouse model with null alleles at the loci encoding the three relevant receptors for tumor necrosis factors α and β and IL-1α and IL-1β (i.e., null alleles at the Tnfrsf1a, Tnfrsf1b, and Il-1r1 loci). The observation that TKO mice were resistant to the tumor promoting effects of dioxin in liver suggests that inflammatory cytokines play an important step in dioxin mediated liver tumor promotion in the mouse. Collectively, these data support the idea that the mechanism of dioxin acute hepatotoxicity and its activity as a promoter in a mouse two stage liver cancer model may be similar, i.e., tumor promotion by dioxin, like acute hepatotoxicity, are mediated by the linked action of two receptor systems, the AHR and the receptors for the “IL-1-like” cytokines. PMID:24718703

Mutational analysis of the promoter and the coding region of the 5-HT1A gene

Energy Technology Data Exchange (ETDEWEB)

Erdmann, J.; Noethen, M.M.; Shimron-Abarbanell, D. [Univ. of Bonn (Germany)] [and others

1994-09-01

Disturbances of serotonergic pathways have been implicated in many neuropsychiatric disorders. Serotonin (5HT) receptors can be subdivided into at least three major families (5HT1, 5HT2, and 5HT3). Five human 5HT1 receptor subtypes have been cloned, namely 1A, 1D{alpha}, 1D{beta}, 1E, and 1F. Of these, the 5HT1A receptor is the best characterized subtype. In the present study we sought to identify genetic variation in the 5HT1A receptor gene which through alteration of protein function or level of expression might contribute to the genetics of neuropsychiatric diseases. The coding region and the 5{prime} promoter region of the 5HT1A gene from 159 unrelated subjects (45 schizophrenic, 46 bipolar affective, and 43 patients with Tourette`s syndrome, as well as 25 controls) were analyzed using SSCA. SSCA revealed the presence of two mutations both located in the coding region of the 5HT1A receptor gene. The first mutation is a rare silent C{r_arrow}T substitution at nucleotide position 549. The second mutation is characterized by a base pair substitution (A{r_arrow}G) at the first position of codon 28 and results in an amino acid exchange (Ile{r_arrow}Val). Since Val28 was found only in a single schizophrenic patient and in none of the other patients or controls, we decided to extend our samples and to use a restriction assay for screening a further 74 schizophrenic, 95 bipolar affective, and 49 patients with Tourette`s syndrome, as well as 185 controls, for the presence of the mutation. In total, the mutation was found in 2 schizophrenic patients, in 3 bipolars, in 1 Tourette patient, and in 5 controls. To our knowledge the Ile-28-Val substitution reported here is the first natural occuring molecular variant which has been identified for a serotonin receptor so far.

Polymorphism in the oxytocin promoter region in patients with lactase non-persistence is not related to symptoms

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Simrén Magnus

2009-11-01

Full Text Available Abstract Background Oxytocin and the oxytocin receptor have been demonstrated in the gastrointestinal (GI tract and have been shown to exert physiological effects on gut motility. The role for oxytocin in the pathophysiology of GI complaints is unknown. The aim of this study was to examine genetic variations or polymorphism of oxytocin (OXT and its receptor (OXTR genes in patients with GI complaints without visible organic abnormalities. Methods Genetic variants in the OXT promoter region, and in the OXTR gene in DNA samples from 131 rigorously evaluated patients with Irritable Bowel Syndrome (IBS, 408 homozygous subjects referred for lactase (LCT-13910 C>T, rs4988235 genotyping, and 299 asymptomatic blood donors were compared. One polymorphism related to the OXT gene (rs6133010 A>G and 4 related to the OXTR gene (rs1465386 G>T, rs3806675 G>A, rs968389 A>G, rs1042778 G>T were selected for genotyping using Applied Biosystems 7900 HT allele discrimination assays. Results There were no statistically significant differences in the genotype or allele frequencies in any of the SNPs when IBS patients were compared to healthy controls. Among subjects referred for lactase genotyping, the rs6133010 A>G OXT promoter A/G genotype tended to be more common in the 154 non-persistent (27.3% subjects than in the 254 lactase persistant (18.1% subjects and in the healthy controls (19.4% (p = 0.08. When direct comparing, the A/G genotype was less common in the OXT promoter region in controls (p = 0.09 and in subjects with lactase persistence (p = 0.03 compared to subjects with lactase non-persistence. When healthy controls were viewed according to their own LCT-13910 genotypes, the C/C lactase non-persistent controls had a higher frequency for the OXT promoter A/G genotype than LCT-13910 T/T lactase persistent controls (41.2% vs 13.1%. No significant differences in frequencies of the investigated OXTR SNPs were noted in this study. Conclusion The results suggest

EPO-independent functional EPO receptor in breast cancer enhances estrogen receptor activity and promotes cell proliferation

International Nuclear Information System (INIS)

Reinbothe, Susann; Larsson, Anna-Maria; Vaapil, Marica; Wigerup, Caroline; Sun, Jianmin; Jögi, Annika; Neumann, Drorit; Rönnstrand, Lars; Påhlman, Sven

2014-01-01

Highlights: • New anti-human EPOR antibody confirms full-length EPOR expression in breast cancer cells. • Proliferation of breast cancer cells is not affected by rhEPO treatment in vitro. • EPOR knockdown impairs proliferation of ERa positive breast cancer cells. • EPOR knockdown reduces AKT phosphorylation and ERa activity. - Abstract: The main function of Erythropoietin (EPO) and its receptor (EPOR) is the stimulation of erythropoiesis. Recombinant human EPO (rhEPO) is therefore used to treat anemia in cancer patients. However, clinical trials have indicated that rhEPO treatment might promote tumor progression and has a negative effect on patient survival. In addition, EPOR expression has been detected in several cancer forms. Using a newly produced anti-EPOR antibody that reliably detects the full-length isoform of the EPOR we show that breast cancer tissue and cells express the EPOR protein. rhEPO stimulation of cultured EPOR expressing breast cancer cells did not result in increased proliferation, overt activation of EPOR (receptor phosphorylation) or a consistent activation of canonical EPOR signaling pathway mediators such as JAK2, STAT3, STAT5, or AKT. However, EPOR knockdown experiments suggested functional EPO receptors in estrogen receptor positive (ERα + ) breast cancer cells, as reduced EPOR expression resulted in decreased proliferation. This effect on proliferation was not seen in ERα negative cells. EPOR knockdown decreased ERα activity further supports a mechanism by which EPOR affects proliferation via ERα-mediated mechanisms. We show that EPOR protein is expressed in breast cancer cells, where it appears to promote proliferation by an EPO-independent mechanism in ERα expressing breast cancer cells

EPO-independent functional EPO receptor in breast cancer enhances estrogen receptor activity and promotes cell proliferation

Energy Technology Data Exchange (ETDEWEB)

Reinbothe, Susann; Larsson, Anna-Maria; Vaapil, Marica; Wigerup, Caroline [Department of Laboratory Medicine, Translational Cancer Research, Medicon Village, Lund University, SE-223 81 Lund (Sweden); CREATE Health, Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv (Israel); Sun, Jianmin [Department of Laboratory Medicine, Translational Cancer Research, Medicon Village, Lund University, SE-223 81 Lund (Sweden); Jögi, Annika [Department of Laboratory Medicine, Translational Cancer Research, Medicon Village, Lund University, SE-223 81 Lund (Sweden); CREATE Health, Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv (Israel); Neumann, Drorit [Department of Cell and Developmental Biology, Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv (Israel); Rönnstrand, Lars [Department of Laboratory Medicine, Translational Cancer Research, Medicon Village, Lund University, SE-223 81 Lund (Sweden); Påhlman, Sven, E-mail: sven.pahlman@med.lu.se [Department of Laboratory Medicine, Translational Cancer Research, Medicon Village, Lund University, SE-223 81 Lund (Sweden); CREATE Health, Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv (Israel)

2014-02-28

Highlights: • New anti-human EPOR antibody confirms full-length EPOR expression in breast cancer cells. • Proliferation of breast cancer cells is not affected by rhEPO treatment in vitro. • EPOR knockdown impairs proliferation of ERa positive breast cancer cells. • EPOR knockdown reduces AKT phosphorylation and ERa activity. - Abstract: The main function of Erythropoietin (EPO) and its receptor (EPOR) is the stimulation of erythropoiesis. Recombinant human EPO (rhEPO) is therefore used to treat anemia in cancer patients. However, clinical trials have indicated that rhEPO treatment might promote tumor progression and has a negative effect on patient survival. In addition, EPOR expression has been detected in several cancer forms. Using a newly produced anti-EPOR antibody that reliably detects the full-length isoform of the EPOR we show that breast cancer tissue and cells express the EPOR protein. rhEPO stimulation of cultured EPOR expressing breast cancer cells did not result in increased proliferation, overt activation of EPOR (receptor phosphorylation) or a consistent activation of canonical EPOR signaling pathway mediators such as JAK2, STAT3, STAT5, or AKT. However, EPOR knockdown experiments suggested functional EPO receptors in estrogen receptor positive (ERα{sup +}) breast cancer cells, as reduced EPOR expression resulted in decreased proliferation. This effect on proliferation was not seen in ERα negative cells. EPOR knockdown decreased ERα activity further supports a mechanism by which EPOR affects proliferation via ERα-mediated mechanisms. We show that EPOR protein is expressed in breast cancer cells, where it appears to promote proliferation by an EPO-independent mechanism in ERα expressing breast cancer cells.

Glycine Receptor α2 Subunit Activation Promotes Cortical Interneuron Migration

Directory of Open Access Journals (Sweden)

Ariel Avila

2013-08-01

Full Text Available Glycine receptors (GlyRs are detected in the developing CNS before synaptogenesis, but their function remains elusive. This study demonstrates that functional GlyRs are expressed by embryonic cortical interneurons in vivo. Furthermore, genetic disruption of these receptors leads to interneuron migration defects. We discovered that extrasynaptic activation of GlyRs containing the α2 subunit in cortical interneurons by endogenous glycine activates voltage-gated calcium channels and promotes calcium influx, which further modulates actomyosin contractility to fine-tune nuclear translocation during migration. Taken together, our data highlight the molecular events triggered by GlyR α2 activation that control cortical tangential migration during embryogenesis.

Two distinct promoters drive transcription of the human D1A dopamine receptor gene.

Science.gov (United States)

Lee, S H; Minowa, M T; Mouradian, M M

1996-10-11

The human D1A dopamine receptor gene has a GC-rich, TATA-less promoter located upstream of a small, noncoding exon 1, which is separated from the coding exon 2 by a 116-base pair (bp)-long intron. Serial 3'-deletions of the 5'-noncoding region of this gene, including the intron and 5'-end of exon 2, resulted in 80 and 40% decrease in transcriptional activity of the upstream promoter in two D1A-expressing neuroblastoma cell lines, SK-N-MC and NS20Y, respectively. To investigate the function of this region, the intron and 245 bp at the 5'-end of exon 2 were investigated. Transient expression analyses using various chloramphenicol acetyltransferase constructs showed that the transcriptional activity of the intron is higher than that of the upstream promoter by 12-fold in SK-N-MC cells and by 5.5-fold in NS20Y cells in an orientation-dependent manner, indicating that the D1A intron is a strong promoter. Primer extension and ribonuclease protection assays revealed that transcription driven by the intron promoter is initiated at the junction of intron and exon 2 and at a cluster of nucleotides located 50 bp downstream from this junction. The same transcription start sites are utilized by the chloramphenicol acetyltransferase constructs employed in transfections as well as by the D1A gene expressed within the human caudate. The relative abundance of D1A transcripts originating from the upstream promoter compared with those transcribed from the intron promoter is 1.5-2.9 times in SK-N-MC cells and 2 times in the human caudate. Transcript stability studies in SK-N-MC cells revealed that longer D1A mRNA molecules containing exon 1 are degraded 1.8 times faster than shorter transcripts lacking exon 1. Although gel mobility shift assay could not detect DNA-protein interaction at the D1A intron, competitive co-transfection using the intron as competitor confirmed the presence of trans-acting factors at the intron. These data taken together indicate that the human D1A gene has

The human luteinizing hormone receptor gene promoter: activation by Sp1 and Sp3 and inhibitory regulation.

Science.gov (United States)

Geng, Y; Tsai-Morris, C H; Zhang, Y; Dufau, M L

1999-09-24

To understand the transcriptional mechanism(s) of human LH receptor (LHR) gene expression, we have identified the dominant functional cis-elements that regulate the activity of the promoter domain (-1 to -176 bp from ATG). Mutagenesis demonstrated that the promoter activity was dependent on two Sp1 domains (-79 bp, -120 bp) in a transformed normal placental cell (PLC) and the choriocarcinoma JAR cell. Both elements interacted with endogenous Sp1 and Sp3 factors but not with Sp2 or Sp4. In Drosophila SL2 cells, the promoter was activated by either Sp1 or Sp3. An ERE half-site (EREhs) at -174 bp was inhibitory (by 100%), but was unresponsive to estradiol and did not bind the estrogen receptor or orphan receptors ERR1 and SF-1. The 5' upstream sequence (-177 to -2056 bp) inhibited promoter activity in PLC by 60%, but only minimally in JAR cells. Activation of the human LHR promoter through Sp1/3 factors is negatively regulated through EREhs and upstream sequences to exert control of gene expression. Copyright 1999 Academic Press.

Variants in the dopamine-4-receptor gene promoter are not associated with sensation seeking in skiers.

Science.gov (United States)

Thomson, Cynthia J; Rajala, Amelia K; Carlson, Scott R; Rupert, Jim L

2014-01-01

Sensation seeking is a personality trait that has been associated with disinhibited behaviours including substance use and gambling, but also with high-risk sport practices including skydiving, paragliding, and downhill skiing. Twin studies have shown that sensation seeking is moderately heritable, and candidate genes encoding components involved in dopaminergic transmission have been investigated as contributing to this type of behaviour. To determine whether variants in the regulatory regions of the dopamine-4-receptor gene (DRD4) influenced sport-specific sensation seeking, we analyzed five polymorphisms (-1106T/C, -906T/C, -809G/A, -291C/T, 120-bp duplication) in the promoter region of the gene in a cohort of skiers and snowboarders (n = 599) that represented a broad range of sensation seeking behaviours. We grouped subjects by genotype at each of the five loci and compared impulsive sensation seeking and domain-specific (skiing) sensation seeking between groups. There were no significant associations between genotype(s) and general or domain-specific sensation seeking in the skiers and snowboarders, suggesting that while DRD4 has previously been implicated in sensation seeking, the promoter variants investigated in this study do not contribute to sensation seeking in this athlete population.

Antihypertensive drug Valsartan promotes dendritic spine density by altering AMPA receptor trafficking

Science.gov (United States)

Sohn, Young In; Lee, Nathanael J.; Chung, Andrew; Saavedra, Juan M.; Turner, R. Scott; Pak, Daniel T. S.; Hoe, Hyang-Sook

2013-01-01

Recent studies demonstrated that the antihypertensive drug Valsartan improved spatial and episodic memory in mouse models of Alzheimer’s Disease (AD) and human subjects with hypertension. However, the molecular mechanism by which Valsartan can regulate cognitive function is still unknown. Here, we investigated the effect of Valsartan on dendritic spine formation in primary hippocampal neurons, which is correlated with learning and memory. Interestingly, we found that Valsartan promotes spinogenesis in developing and mature neurons. In addition, we found that Valsartan increases the puncta number of PSD-95 and trends toward an increase in the puncta number of synaptophysin. Moreover, Valsartan increased the cell surface levels of AMPA receptors and selectively altered the levels of spinogenesis-related proteins, including CaMKIIα and phospho-CDK5. These data suggest that Valsartan may promote spinogenesis by enhancing AMPA receptor trafficking and synaptic plasticity signaling. PMID:24012668

Evaluation of methylation pattern in promoter region of E-cadherin ...

African Journals Online (AJOL)

user

2011-03-07

Mar 7, 2011 ... promoter methylation in CDH1 gene inactivation in breast cancer, the CpG methylation status of E- ..... 5'CpG island of CDH1 in prostate, lung, liver, bladder, .... and estrogen receptor alpha from Sp1 sites to induce cell cycle.

Genome-wide analysis of regions similar to promoters of histone genes

KAUST Repository

Chowdhary, Rajesh

2010-05-28

Background: The purpose of this study is to: i) develop a computational model of promoters of human histone-encoding genes (shortly histone genes), an important class of genes that participate in various critical cellular processes, ii) use the model so developed to identify regions across the human genome that have similar structure as promoters of histone genes; such regions could represent potential genomic regulatory regions, e.g. promoters, of genes that may be coregulated with histone genes, and iii/ identify in this way genes that have high likelihood of being coregulated with the histone genes.Results: We successfully developed a histone promoter model using a comprehensive collection of histone genes. Based on leave-one-out cross-validation test, the model produced good prediction accuracy (94.1% sensitivity, 92.6% specificity, and 92.8% positive predictive value). We used this model to predict across the genome a number of genes that shared similar promoter structures with the histone gene promoters. We thus hypothesize that these predicted genes could be coregulated with histone genes. This hypothesis matches well with the available gene expression, gene ontology, and pathways data. Jointly with promoters of the above-mentioned genes, we found a large number of intergenic regions with similar structure as histone promoters.Conclusions: This study represents one of the most comprehensive computational analyses conducted thus far on a genome-wide scale of promoters of human histone genes. Our analysis suggests a number of other human genes that share a high similarity of promoter structure with the histone genes and thus are highly likely to be coregulated, and consequently coexpressed, with the histone genes. We also found that there are a large number of intergenic regions across the genome with their structures similar to promoters of histone genes. These regions may be promoters of yet unidentified genes, or may represent remote control regions that

Transactivation of the proximal promoter of human oxytocin gene by TR4 orphan receptor

International Nuclear Information System (INIS)

Wang, C.-P.; Lee, Y.-F.; Chang, C.; Lee, H.-J.

2006-01-01

The human testicular receptor 4 (TR4) shares structural homology with members of the nuclear receptor superfamily. Some other members of this superfamily were able to regulate the transcriptional activity of the human oxytocin (OXT) promoter by binding to the first DR0 regulatory site. However, little investigation was conducted systematically in the study of the second dDR4 site of OXT proximal promoter, and the relationship between the first and the second sites of OXT promoter. Here, we demonstrated for the first time that TR4 could increase the proximal promoter activity of the human OXT gene via DR0, dDR4, and OXT (both DR0 and dDR4) elements, respectively. TR4 might induce OXT gene expression through the OXT element in a dose-dependent manner. However, there is no synergistic effect between DR0 and dDR4 elements during TR4 transactivation. Taken together, these results suggested that TR4 should be one of important regulators of OXT gene expression

Function of the cytoplasmic tail of human calcitonin receptor-like receptor in complex with receptor activity-modifying protein 2

Energy Technology Data Exchange (ETDEWEB)

Kuwasako, Kenji, E-mail: kuwasako@fc.miyazaki-u.ac.jp [Frontier Science Research Center, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki 889-1692 (Japan); Kitamura, Kazuo; Nagata, Sayaka; Hikosaka, Tomomi [Division of Circulation and Body Fluid Regulation, Faculty of Medicine, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki 889-1692 (Japan); Kato, Johji [Frontier Science Research Center, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki 889-1692 (Japan)

2010-02-12

Receptor activity-modifying protein 2 (RAMP2) enables calcitonin receptor-like receptor (CRLR) to form an adrenomedullin (AM)-specific receptor. Here we investigated the function of the cytoplasmic C-terminal tail (C-tail) of human (h)CRLR by co-transfecting its C-terminal mutants into HEK-293 cells stably expressing hRAMP2. Deleting the C-tail from CRLR disrupted AM-evoked cAMP production or receptor internalization, but did not affect [{sup 125}I]AM binding. We found that CRLR residues 428-439 are required for AM-evoked cAMP production, though deleting this region had little effect on receptor internalization. Moreover, pretreatment with pertussis toxin (100 ng/mL) led to significant increases in AM-induced cAMP production via wild-type CRLR/RAMP2 complexes. This effect was canceled by deleting CRLR residues 454-457, suggesting Gi couples to this region. Flow cytometric analysis revealed that CRLR truncation mutants lacking residues in the Ser/Thr-rich region extending from Ser{sup 449} to Ser{sup 467} were unable to undergo AM-induced receptor internalization and, in contrast to the effect on wild-type CRLR, overexpression of GPCR kinases-2, -3 and -4 failed to promote internalization of CRLR mutants lacking residues 449-467. Thus, the hCRLR C-tail is crucial for AM-evoked cAMP production and internalization of the CRLR/RAMP2, while the receptor internalization is dependent on the aforementioned GPCR kinases, but not Gs coupling.

Synergistic regulation of the mouse orphan nuclear receptor SHP gene promoter by CLOCK-BMAL1 and LRH-1

International Nuclear Information System (INIS)

Oiwa, Ako; Kakizawa, Tomoko; Miyamoto, Takahide; Yamashita, Koh; Jiang, Wei; Takeda, Teiji; Suzuki, Satoru; Hashizume, Kiyoshi

2007-01-01

Small heterodimer partner (SHP; NR0B2) is an orphan nuclear receptor and acts as a repressor for wide variety of nuclear hormone receptors. We demonstrated here that mouse SHP mRNA showed a circadian expression pattern in the liver. Transient transfection of the mSHP promoter demonstrated that CLOCK-BMAL1, core circadian clock components, bound to E-box (CACGTG), and stimulated the promoter activity by 4-fold. Liver receptor homologue-1 (LRH-1; NR5A2) stimulated the mSHP promoter, and CLOCK-BMAL1 synergistically enhanced the LRH-1-mediated transactivation. Interestingly, SHP did not affect the CLOCK-BMAL1-mediated promoter activity, but strongly repressed the synergistic activation of CLOCK-BMAL1 and LRH-1. Furthermore, in vitro pull-down assays revealed the existence of direct protein-protein interaction between LRH-1 and CLOCK. In summary, this study shows that CLOCK-BMAL1, LRH-1 and SHP coordinately regulate the mSHP gene to generate the circadian oscillation. The cyclic expression of mSHP may affect daily activity of other nuclear receptors and contribute to circadian liver functions

Impulsive traits and 5-HT2A receptor promoter polymorphism in alcohol dependents: Possible association but no influence of personality disorders

OpenAIRE

Preuss, Ulrich W.; Koller, G.; Bondy, Brigitta; Bahlmann, Miriam; Soyka, Michael

2001-01-01

Objective: Impulsive behavior in alcoholics puts them at serious risk of severer course of disease and has been related to the serotonergic neurotransmission dysfunction. The aim of this study is to investigate the association between impulsive aggression in alcohol dependents with regard to the G-1438A polymorphism in the promoter region of the 5-HT2A receptor gene. Furthermore, we investigated the statistical interaction between 5-HT2A alleles, antisocial personality disorder (APD) and impu...

Variants in the dopamine-4-receptor gene promoter are not associated with sensation seeking in skiers.

Directory of Open Access Journals (Sweden)

Cynthia J Thomson

Full Text Available Sensation seeking is a personality trait that has been associated with disinhibited behaviours including substance use and gambling, but also with high-risk sport practices including skydiving, paragliding, and downhill skiing. Twin studies have shown that sensation seeking is moderately heritable, and candidate genes encoding components involved in dopaminergic transmission have been investigated as contributing to this type of behaviour. To determine whether variants in the regulatory regions of the dopamine-4-receptor gene (DRD4 influenced sport-specific sensation seeking, we analyzed five polymorphisms (-1106T/C, -906T/C, -809G/A, -291C/T, 120-bp duplication in the promoter region of the gene in a cohort of skiers and snowboarders (n = 599 that represented a broad range of sensation seeking behaviours. We grouped subjects by genotype at each of the five loci and compared impulsive sensation seeking and domain-specific (skiing sensation seeking between groups. There were no significant associations between genotype(s and general or domain-specific sensation seeking in the skiers and snowboarders, suggesting that while DRD4 has previously been implicated in sensation seeking, the promoter variants investigated in this study do not contribute to sensation seeking in this athlete population.

Supersensitive Kappa Opioid Receptors Promotes Ethanol Withdrawal-Related Behaviors and Reduce Dopamine Signaling in the Nucleus Accumbens.

Science.gov (United States)

Rose, Jamie H; Karkhanis, Anushree N; Chen, Rong; Gioia, Dominic; Lopez, Marcelo F; Becker, Howard C; McCool, Brian A; Jones, Sara R

2016-05-01

Chronic ethanol exposure reduces dopamine transmission in the nucleus accumbens, which may contribute to the negative affective symptoms associated with ethanol withdrawal. Kappa opioid receptors have been implicated in withdrawal-induced excessive drinking and anxiety-like behaviors and are known to inhibit dopamine release in the nucleus accumbens. The effects of chronic ethanol exposure on kappa opioid receptor-mediated changes in dopamine transmission at the level of the dopamine terminal and withdrawal-related behaviors were examined. Five weeks of chronic intermittent ethanol exposure in male C57BL/6 mice were used to examine the role of kappa opioid receptors in chronic ethanol-induced increases in ethanol intake and marble burying, a measure of anxiety/compulsive-like behavior. Drinking and marble burying were evaluated before and after chronic intermittent ethanol exposure, with and without kappa opioid receptor blockade by nor-binaltorphimine (10mg/kg i.p.). Functional alterations in kappa opioid receptors were assessed using fast scan cyclic voltammetry in brain slices containing the nucleus accumbens. Chronic intermittent ethanol-exposed mice showed increased ethanol drinking and marble burying compared with controls, which was attenuated with kappa opioid receptor blockade. Chronic intermittent ethanol-induced increases in behavior were replicated with kappa opioid receptor activation in naïve mice. Fast scan cyclic voltammetry revealed that chronic intermittent ethanol reduced accumbal dopamine release and increased uptake rates, promoting a hypodopaminergic state of this region. Kappa opioid receptor activation with U50,488H concentration-dependently decreased dopamine release in both groups; however, this effect was greater in chronic intermittent ethanol-treated mice, indicating kappa opioid receptor supersensitivity in this group. These data suggest that the chronic intermittent ethanol-induced increase in ethanol intake and anxiety

Cross-talk between an activator of nuclear receptors-mediated transcription and the D1 dopamine receptor signaling pathway.

Science.gov (United States)

Schmidt, Azriel; Vogel, Robert; Rutledge, Su Jane; Opas, Evan E; Rodan, Gideon A; Friedman, Eitan

2005-03-01

Nuclear receptors are transcription factors that usually interact, in a ligand-dependent manner, with specific DNA sequences located within promoters of target genes. The nuclear receptors can also be controlled in a ligand-independent manner via the action of membrane receptors and cellular signaling pathways. 5-Tetradecyloxy-2-furancarboxylic acid (TOFA) was shown to stimulate transcription from the MMTV promoter via chimeric receptors that consist of the DNA binding domain of GR and the ligand binding regions of the PPARbeta or LXRbeta nuclear receptors (GR/PPARbeta and GR/LXRbeta). TOFA and hydroxycholesterols also modulate transcription from NF-kappaB- and AP-1-controlled reporter genes and induce neurite differentiation in PC12 cells. In CV-1 cells that express D(1) dopamine receptors, D(1) dopamine receptor stimulation was found to inhibit TOFA-stimulated transcription from the MMTV promoter that is under the control of chimeric GR/PPARbeta and GR/LXRbeta receptors. Treatment with the D(1) dopamine receptor antagonist, SCH23390, prevented dopamine-mediated suppression of transcription, and by itself increased transcription controlled by GR/LXRbeta. Furthermore, combined treatment of CV-1 cells with TOFA and SCH23390 increased transcription controlled by the GR/LXRbeta chimeric receptor synergistically. The significance of this in vitro synergy was demonstrated in vivo, by the observation that SCH23390 (but not haloperidol)-mediated catalepsy in rats was potentiated by TOFA, thus showing that an agent that mimics the in vitro activities of compounds that activate members of the LXR and PPAR receptor families can influence D1 dopamine receptor elicited responses.

Identification and annotation of promoter regions in microbial

Indian Academy of Sciences (India)

2007-06-15

Jun 15, 2007 ... Analysis of various predicted structural properties of promoter regions in prokaryotic as well as eukaryotic genomes had earlier indicated that they have several common features, such as lower stability, higher curvature and less bendability, when compared with their neighboring regions. Based on the ...

Lactate Receptor Sites Link Neurotransmission, Neurovascular Coupling, and Brain Energy Metabolism

DEFF Research Database (Denmark)

Lauritzen, Knut H; Morland, Cecilie; Puchades, Maja

2013-01-01

The G-protein-coupled lactate receptor, GPR81 (HCA1), is known to promote lipid storage in adipocytes by downregulating cAMP levels. Here, we show that GPR81 is also present in the mammalian brain, including regions of the cerebral neocortex and hippocampus, where it can be activated by physiolog......The G-protein-coupled lactate receptor, GPR81 (HCA1), is known to promote lipid storage in adipocytes by downregulating cAMP levels. Here, we show that GPR81 is also present in the mammalian brain, including regions of the cerebral neocortex and hippocampus, where it can be activated...

Characterization of the promoter of human CRTh2, a prostaglandin D{sub 2} receptor

Energy Technology Data Exchange (ETDEWEB)

Quapp, Russell; Madsen, Norman [Department of Medicine, Division of Pulmonary Medicine, Pulmonary Research Group, 574B Heritage Medical Research Centre, University of Alberta, Edmonton, AB, T6G 2S2 (Canada); Cameron, Lisa [Department of Medicine, Division of Pulmonary Medicine, Pulmonary Research Group, 574B Heritage Medical Research Centre, University of Alberta, Edmonton, AB, T6G 2S2 (Canada)

2007-11-30

Chemoattractant-receptor homologous molecule expressed on Th2 cells (CRTh2) is a receptor for prostaglandin (PG)D{sub 2}, a lipid mediator involved in allergic inflammation. CRTh2 is expressed by Th2 cells, eosinophils and basophils and PDG{sub 2}-CRTh2 signaling induces calcium mobilization, cell migration and expression of the Th2 cytokines IL-4, IL-5, and IL-13. Despite the role of CRTh2 in allergic inflammation, transcriptional regulation of this gene has not been studied. Here, we demonstrated that a reporter construct of the CRTh2 promoter was induced following T cell stimulation. This activity could be further enhanced by over-expression of GATA-3, but not NFAT2 or STAT6. Electromobility shift assay demonstrated GATA-3 binding to a probe from the CRTh2 promoter. This study provides the first detailed analysis of transcriptional regulation of the human CRTh2 promoter. These findings may help identify strategies to attenuate expression of this gene and influence the maintenance and proliferation of Th2 cells in allergic inflammation.

Region-specific expression and hormonal regulation of the first exon variants of rat prolactin receptor mRNA in rat brain and anterior pituitary gland.

Science.gov (United States)

Nogami, H; Hoshino, R; Ogasawara, K; Miyamoto, S; Hisano, S

2007-08-01

Recent studies have revealed the occurrence of five first exon variants of the rat prolactin receptor mRNA, suggesting that multiple promoters direct prolactin receptor transcription in response to different regulatory factors. In the present study, regional expression of these first exon variants, as well as two prolactin receptor subtypes generated by alternative splicing, was examined in the brains and anterior pituitary glands of female rats. Expression of the long-form was detected in the choroid plexus, hypothalamus, hippocampus, cerebral cortex and anterior pituitary gland, whereas the short form was detected only in the choroid plexus. E1-3 mRNA, a first exon variant, was detected in the choroid plexus, hypothalamus, and anterior pituitary gland, whereas E1-4 was detected only in the choroid plexus. Other variants were not detectable by the polymerase chain reaction protocol employed in this study. Ovariectomy increased the short form in the choroid plexus and the E1-3 expression in the choroid plexus and pituitary gland, but changes in the long-form and E1-4 expression were minimal. Replacement of oestrogens and prolactin suggest that oestrogens down-regulate E1-3 expression in the choroid plexus and pituitary gland, and that the negative effect of oestrogen is mediated by prolactin in the pituitary gland. The present results revealed the region-specific promoter usage in prolactin receptor mRNA transcription, as well as the involvement of oestrogens in the regulation of E1-3 mRNA expression in the brain and pituitary gland.

Promotion of cancer cell invasiveness and metastasis emergence caused by olfactory receptor stimulation.

Directory of Open Access Journals (Sweden)

Guenhaël Sanz

Full Text Available Olfactory receptors (ORs are expressed in the olfactory epithelium, where they detect odorants, but also in other tissues with additional functions. Some ORs are even overexpressed in tumor cells. In this study, we identified ORs expressed in enterochromaffin tumor cells by RT-PCR, showing that single cells can co-express several ORs. Some of the receptors identified were already reported in other tumors, but they are orphan (without known ligand, as it is the case for most of the hundreds of human ORs. Thus, genes coding for human ORs with known ligands were transfected into these cells, expressing functional heterologous ORs. The in vitro stimulation of these cells by the corresponding OR odorant agonists promoted cell invasion of collagen gels. Using LNCaP prostate cancer cells, the stimulation of the PSGR (Prostate Specific G protein-coupled Receptor, an endogenously overexpressed OR, by β-ionone, its odorant agonist, resulted in the same phenotypic change. We also showed the involvement of a PI3 kinase γ dependent signaling pathway in this promotion of tumor cell invasiveness triggered by OR stimulation. Finally, after subcutaneous inoculation of LNCaP cells into NSG immunodeficient mice, the in vivo stimulation of these cells by the PSGR agonist β-ionone significantly enhanced metastasis emergence and spreading.

Characterization of dFOXO binding sites upstream of the Insulin Receptor P2 promoter across the Drosophila phylogeny.

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Dorcas J Orengo

Full Text Available The insulin/TOR signal transduction pathway plays a critical role in determining such important traits as body and organ size, metabolic homeostasis and life span. Although this pathway is highly conserved across the animal kingdom, the affected traits can exhibit important differences even between closely related species. Evolutionary studies of regulatory regions require the reliable identification of transcription factor binding sites. Here we have focused on the Insulin Receptor (InR expression from its P2 promoter in the Drosophila genus, which in D. melanogaster is up-regulated by hypophosphorylated Drosophila FOXO (dFOXO. We have finely characterized this transcription factor binding sites in vitro along the 1.3 kb region upstream of the InR P2 promoter in five Drosophila species. Moreover, we have tested the effect of mutations in the characterized dFOXO sites of D. melanogaster in transgenic flies. The number of experimentally established binding sites varies across the 1.3 kb region of any particular species, and their distribution also differs among species. In D. melanogaster, InR expression from P2 is differentially affected by dFOXO binding sites at the proximal and distal halves of the species 1.3 kb fragment. The observed uneven distribution of binding sites across this fragment might underlie their differential contribution to regulate InR transcription.

Differences in signal activation by LH and hCG are mediated by the LH/CG receptor`s extracellular hinge region

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Paul eGrzesik

2015-09-01

Full Text Available The human lutropin/choriogonadotropin receptor (LHCGR can be activated by binding two slightly different gonadotropic glycoprotein hormones, choriogonadotropin (CG - secreted by the placenta, and lutropin (LH - produced by the pituitary. They induce different signaling profiles at the LHCGR. This cannot be explained by binding to the receptor's leucine-rich repeat domain (LRRD, as this binding is similar for the two hormones. We therefore speculate that there are previously unknown differences in the hormone/receptor interaction at the extracellular hinge region, which might help to understand functional differences between the two hormones. We have therefore performed a detailed study of the binding and action of LH and CG at the LHCGR hinge region. We focused on a primate-specific additional exon in the hinge region, which is located between LRRD and the serpentine domain. The segment of the hinge region encoded by exon10 was previously reported to be only relevant to hLH signaling, as the exon10-deletion receptor exhibits decreased hLH signaling, but unchanged hCG signaling. We designed an advanced homology model of the hormone/LHCGR complex, followed by experimental characterization of relevant fragments in the hinge region. In addition, we examined predictions of a helical exon10-encoded conformation by block-wise polyalanine (helix supporting mutations. These helix preserving modifications showed no effect on hormone induced signaling. However, introduction of a structure-disturbing double-proline mutant LHCGR-Q303P/E305P within the exon10-helix has, in contrast to exon10 deletion, no impact on hLH, but only on hCG signaling. This opposite effect on signaling by hLH and hCG can be explained by distinct sites of hormone interaction in the hinge region s. In conclusion, our analysis provides details of the differences between hLH- and hCG-induced signaling that are mainly determined in the L2-beta loop of the hormones and in the hinge region

Epigenetic transgenerational actions of vinclozolin on promoter regions of the sperm epigenome.

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Carlos Guerrero-Bosagna

2010-09-01

Full Text Available Previous observations have demonstrated that embryonic exposure to the endocrine disruptor vinclozolin during gonadal sex determination promotes transgenerational adult onset disease such as male infertility, kidney disease, prostate disease, immune abnormalities and tumor development. The current study investigates genome-wide promoter DNA methylation alterations in the sperm of F3 generation rats whose F0 generation mother was exposed to vinclozolin. A methylated DNA immunoprecipitation with methyl-cytosine antibody followed by a promoter tilling microarray (MeDIP-Chip procedure was used to identify 52 different regions with statistically significant altered methylation in the sperm promoter epigenome. Mass spectrometry bisulfite analysis was used to map the CpG DNA methylation and 16 differential DNA methylation regions were confirmed, while the remainder could not be analyzed due to bisulfite technical limitations. Analysis of these validated regions identified a consensus DNA sequence (motif that associated with 75% of the promoters. Interestingly, only 16.8% of a random set of 125 promoters contained this motif. One candidate promoter (Fam111a was found to be due to a copy number variation (CNV and not a methylation change, suggesting initial alterations in the germline epigenome may promote genetic abnormalities such as induced CNV in later generations. This study identifies differential DNA methylation sites in promoter regions three generations after the initial exposure and identifies common genome features present in these regions. In addition to primary epimutations, a potential indirect genetic abnormality was identified, and both are postulated to be involved in the epigenetic transgenerational inheritance observed. This study confirms that an environmental agent has the ability to induce epigenetic transgenerational changes in the sperm epigenome.

Epigenetic transgenerational actions of vinclozolin on promoter regions of the sperm epigenome.

Science.gov (United States)

Guerrero-Bosagna, Carlos; Settles, Matthew; Lucker, Ben; Skinner, Michael K

2010-09-30

Previous observations have demonstrated that embryonic exposure to the endocrine disruptor vinclozolin during gonadal sex determination promotes transgenerational adult onset disease such as male infertility, kidney disease, prostate disease, immune abnormalities and tumor development. The current study investigates genome-wide promoter DNA methylation alterations in the sperm of F3 generation rats whose F0 generation mother was exposed to vinclozolin. A methylated DNA immunoprecipitation with methyl-cytosine antibody followed by a promoter tilling microarray (MeDIP-Chip) procedure was used to identify 52 different regions with statistically significant altered methylation in the sperm promoter epigenome. Mass spectrometry bisulfite analysis was used to map the CpG DNA methylation and 16 differential DNA methylation regions were confirmed, while the remainder could not be analyzed due to bisulfite technical limitations. Analysis of these validated regions identified a consensus DNA sequence (motif) that associated with 75% of the promoters. Interestingly, only 16.8% of a random set of 125 promoters contained this motif. One candidate promoter (Fam111a) was found to be due to a copy number variation (CNV) and not a methylation change, suggesting initial alterations in the germline epigenome may promote genetic abnormalities such as induced CNV in later generations. This study identifies differential DNA methylation sites in promoter regions three generations after the initial exposure and identifies common genome features present in these regions. In addition to primary epimutations, a potential indirect genetic abnormality was identified, and both are postulated to be involved in the epigenetic transgenerational inheritance observed. This study confirms that an environmental agent has the ability to induce epigenetic transgenerational changes in the sperm epigenome.

Role of ELA region in auto-activation of mutant KIT receptor: a molecular dynamics simulation insight.

Science.gov (United States)

Purohit, Rituraj

2014-01-01

KIT receptor is the prime target in gastrointestinal stromal tumor (GISTs) therapy. Second generation inhibitor, Sunitinib, binds to an inactivated conformation of KIT receptor and stabilizes it in order to prevent tumor formation. Here, we investigated the dynamic behavior of wild type and mutant D816H KIT receptor, and emphasized the extended A-loop (EAL) region (805-850) by conducting molecular dynamics simulation (∼100 ns). We analyzed different properties such as root mean square cutoff or deviation, root mean square fluctuation, radius of gyration, solvent-accessible surface area, hydrogen bonding network analysis, and essential dynamics. Apart from this, clustering and cross-correlation matrix approach was used to explore the conformational space of the wild type and mutant EAL region of KIT receptor. Molecular dynamics analysis indicated that mutation (D816H) was able to alter intramolecular hydrogen bonding pattern and affected the structural flexibility of EAL region. Moreover, flexible secondary elements, specially, coil and turns were dominated in EAL region of mutant KIT receptor during simulation. This phenomenon increased the movement of EAL region which in turn helped in shifting the equilibrium towards the active kinase conformation. Our atomic investigation of mutant KIT receptor which emphasized on EAL region provided a better insight into the understanding of Sunitinib resistance mechanism of KIT receptor and would help to discover new therapeutics for KIT-based resistant tumor cells in GIST therapy.

Unveiling DNA structural properties of promoter regions of ...

Indian Academy of Sciences (India)

Aditya Kumar

Unveiling DNA structural properties of promoter regions of prokaryotic transcriptome and their role in gene expression. Aditya Kumar. Assistant Professor. Molecular Biology & Biotechnology. Tezpur University. Tezpur – 784028, Assam ...

A Bacterial Receptor PcrK Senses the Plant Hormone Cytokinin to Promote Adaptation to Oxidative Stress

Directory of Open Access Journals (Sweden)

Fang-Fang Wang

2017-12-01

Full Text Available Summary: Recognition of the host plant is a prerequisite for infection by pathogenic bacteria. However, how bacterial cells sense plant-derived stimuli, especially chemicals that function in regulating plant development, remains completely unknown. Here, we have identified a membrane-bound histidine kinase of the phytopathogenic bacterium Xanthomonas campestris, PcrK, as a bacterial receptor that specifically detects the plant cytokinin 2-isopentenyladenine (2iP. 2iP binds to the extracytoplasmic region of PcrK to decrease its autokinase activity. Through a four-step phosphorelay, 2iP stimulation decreased the phosphorylation level of PcrR, the cognate response regulator of PcrK, to activate the phosphodiesterase activity of PcrR in degrading the second messenger 3′,5′-cyclic diguanylic acid. 2iP perception by the PcrK-PcrR remarkably improves bacterial tolerance to oxidative stress by regulating the transcription of 56 genes, including the virulence-associated TonB-dependent receptor gene ctrA. Our results reveal an evolutionarily conserved, inter-kingdom signaling by which phytopathogenic bacteria intercept a plant hormone signal to promote adaptation to oxidative stress. : How pathogenic bacteria use receptors to recognize the signals of the host plant is unknown. Wang et al. have identified a bacterial receptor histidine kinase that specifically senses the plant hormone cytokinin. Through a four-step phosphorelay, cytokinin perception triggers degradation of a second messenger, c-di-GMP, to activate the bacterial response to oxidative stress. Keywords: histidine kinase, ligand, cytokinin, autokinase activity, phosphorelay, response regulator, two-component signal transduction system, Xanthomonas campestris pv. campestris, virulence, oxidative stress

Novel variants in the putative peroxisome proliferator-activated receptor {gamma} promoter and relationships with obesity in men

DEFF Research Database (Denmark)

Larsen, Thomas M; Larsen, Lesli H; Torekov, Signe K

2005-01-01

Yet unidentified variants within the peroxisome proliferator-activated receptor gamma (PPARgamma) 2 promoter may explain the inconsistent reports on associations between variants in the coding region and obesity or diabetes. Thus, we examined the putative PPARgamma2 promoter (-3371 to +43 bp......) for variants in 83 subjects with obesity or type 2 diabetes. We identified eight variants, seven of which were novel, including -792A>G, -816C>T, -882T>C, -1505G>A, -1881C>T, -1884T>A, -2604T>C, and -2953A>G. The variants -816C>T, -1505G>A, -1881C>T, and -2604T>C were in total linkage disequilibrium......, and there was a high degree of linkage disequilibrium between several of the novel variants and Pro12Ala. The novel variants were, together with Pro12Ala and 1431C>T, examined for relationships with obesity among 234 men with early-onset obesity with a BMI at age approximately 20 years of 33.2+/-2.5 kg/m2 and 323...

The main immunogenic region of acetylcholine receptors does not provoke the formation of antibodies of a predominant idiotype.

Science.gov (United States)

Killen, J A; Hochschwender, S M; Lindstrom, J M

1985-08-01

Anti-idiotype antibodies were induced in rats by immunization with rat monoclonal antibodies to the main immunogenic region of acetylcholine receptors. These anti-idiotype antibodies showed very little crossreaction with other rat monoclonal antibodies which bind to the same region of the receptor. When the rats producing these anti-idiotype antibodies were immunized with receptor, they showed no net decrease in anti-receptor antibody production. These data indicate that, although more than half of the antibodies produced by rats immunized with receptor are directed at a small region, many anti-receptor idiotypes are involved in this response and anti-idiotype therapy is not beneficial.

Cancer cell-selective promoter recognition accompanies antitumor effect by glucocorticoid receptor-targeted gold nanoparticle

Science.gov (United States)

Sau, Samaresh; Agarwalla, Pritha; Mukherjee, Sudip; Bag, Indira; Sreedhar, Bojja; Pal-Bhadra, Manika; Patra, Chitta Ranjan; Banerjee, Rajkumar

2014-05-01

Nanoparticles, such as gold nanoparticles (GNP), upon convenient modifications perform multi tasks catering to many biomedical applications. However, GNP or any other type of nanoparticles is yet to achieve the feat of intracellular regulation of endogenous genes of choice such as through manipulation of a gene-promoter in a chromosome. As for gene modulation and delivery, GNP (or other nanoparticles) showed only limited gene therapy potential, which relied on the delivery of `exogenous' genes invoking gene knockdown or replacement. Practically, there are no instances for the nanoparticle-mediated promoter regulation of `endogenous' genes, more so, as a cancer selective phenomenon. In this regard, we report the development of a simple, easily modifiable GNP-formulation, which promoted/up-regulated the expression of a specific category of `endogenous' genes, the glucocorticoid responsive genes. This genetic up-regulation was induced in only cancer cells by modified GNP-mediated transcriptional activation of its cytoplasmic receptor, glucocorticoid receptor (GR). Normal cells and their GR remained primarily unperturbed by this GNP-formulation. The most potent gene up-regulating GNP-formulation down-regulated a cancer-specific proliferative signal, phospho-Akt in cancer cells, which accompanied retardation of tumor growth in the murine melanoma model. We show that GR-targeted GNPs may find potential use in the targeting and modulation of genetic information in cancer towards developing novel anticancer therapeutics.Nanoparticles, such as gold nanoparticles (GNP), upon convenient modifications perform multi tasks catering to many biomedical applications. However, GNP or any other type of nanoparticles is yet to achieve the feat of intracellular regulation of endogenous genes of choice such as through manipulation of a gene-promoter in a chromosome. As for gene modulation and delivery, GNP (or other nanoparticles) showed only limited gene therapy potential, which relied

Social Media Marketing as a tool for promoting the regional investment portals

Directory of Open Access Journals (Sweden)

Alisa Yu. Fadeyeva

2016-01-01

Full Text Available Objective to investigate the potential of Social Media Marketing as a tool for promoting regional investment portals in the information environment to identify the most effective ways of its implementation and to determine the level of mastering of this tool by the Russian regions. Methods general scientific methods observation comparison analysis induction deduction analogy classification. Results the analysis showed that today Social Media Marketing is an essential tool for interaction with the investment community and one of the most effective ways to promote the regional portal which allows to increase the knowledge of and loyalty to the brand to increase the targeted website traffic to increase the awareness of investors about the specific features of the portal and the regional development agenciesrsquo functioning to promptly receive information about the investment environment and to establish contacts with investors. At the same time the study of SMMactivity in the Russian regions revealed a very low level of quality of communication with investors through social networks. Scientific novelty for the first time the article investigates the significance and makes the comparative analysis of the Social Media Marketing channels with regard to investment promotion agencies as well as the results of the regional structures functioning for effective communication through social networks. Practical significance the main results of the research can be used by the regional investment agencies in order to promote their websites increase the quality of communication with investors and promote the investment attractiveness of the region as a whole. nbsp

Thyroid hormone receptor binds to a site in the rat growth hormone promoter required for induction by thyroid hormone

International Nuclear Information System (INIS)

Koenig, R.J.; Brent, G.A.; Warne, R.L.; Larsen, P.R.; Moore, D.D.

1987-01-01

Transcription of the rat growth hormone (rGH) gene in pituitary cells is increased by addition of thyroid hormone (T3). This induction is dependent on the presence of specific sequences just upstream of the rGH promoter. The authors have partially purified T3 receptor from rat liver and examined its interaction with these rGH sequences. They show here that T3 receptor binds specifically to a site just upstream of the basal rGH promoter. This binding site includes two copies of a 7-base-pair direct repeat, the centers of which are separated by 10 base pairs. Deletions that specifically remove the T3 receptor binding site drastically reduce response to T3 in transient transfection experiments. These results demonstrate that T3 receptor can recognize specific DNA sequences and suggest that it can act directly as a positive transcriptional regulatory factor

Neuron-derived orphan receptor 1 promoted human pulmonary artery smooth muscle cells proliferation.

Science.gov (United States)

Wang, Chang-Guo; Lei, Wei; Li, Chang; Zeng, Da-Xiong; Huang, Jian-An

2015-05-01

As a transcription factor of the nuclear receptor superfamily, neuron-derived orphan receptor 1 (NOR1) is induced rapidly in response to various extracellular stimuli. But, it is still unclear its role in pulmonary artery smooth muscle cells proliferation. Human PASMCs were cultured in vitro and stimulated by serum. The special antisense oligodeoxynucleotides (AS-ODNs) were used to knockdown human NOR1 gene expression. Real-time PCR and Western-blot were used to evaluate the gene expression and protein levels. Fetal bovine serum (FBS) induced human PASMCs proliferation in a dose dependent manner. Furthermore, FBS promoted NOR1 gene expression in a dose dependent manner and a time dependent manner. 10% FBS induced a maximal NOR1 mRNA levels at 2 h. FBS also induced a significant higher NOR1 protein levels as compared with control. The NOR1 over-expressed plasmid significantly promoted DNA synthesis and cells proliferation. Moreover, the special AS-ODNs against human NOR1 not only prevented NOR1 expression but also inhibited DNA synthesis and cells proliferation significantly. The NOR1 over-expression plasmid could up-regulate cyclin D1 expression markedly, but the AS-ODNs inhibited cyclin D1 expression significantly. So, we concluded that NOR1 could promote human PASMCs proliferation. Cyclin D1 might be involved in this process.

Receptor protein tyrosine phosphatase alpha enhances rheumatoid synovial fibroblast signaling and promotes arthritis in mice

NARCIS (Netherlands)

Stanford, Stephanie M; Svensson, Mattias N D; Sacchetti, Cristiano; Pilo, Caila A; Wu, Dennis J; Kiosses, William B; Hellvard, Annelie; Bergum, Brith; Aleman Muench, German R; Elly, Christian; Liu, Yun-Cai; den Hertog, Jeroen; Elson, Ari; Sap, Jan; Mydel, Piotr; Boyle, David L; Corr, Maripat; Firestein, Gary S; Bottini, Nunzio

2016-01-01

OBJECTIVE: During rheumatoid arthritis (RA), fibroblast-like synoviocytes (FLS) critically promote disease pathogenesis by aggressively invading the joint extracellular matrix. The focal adhesion kinase (FAK) signaling pathway is emerging as a contributor to RA FLS anomalous behavior. The receptor

Transcriptional activation of melanocortin 2 receptor accessory protein by PPARγ in adipocytes

Energy Technology Data Exchange (ETDEWEB)

Kim, Nam Soo; Kim, Yoon-Jin [Department of Biology, Research Institute for Basic Science, Kyung Hee University, Seoul 130-701 (Korea, Republic of); Cho, Si Young [R and D Center, Amore Pacific Corporation, Yongin-si, Gyeonggi-do 446-729 (Korea, Republic of); Lee, Tae Ryong, E-mail: trlee@amorepacific.com [R and D Center, Amore Pacific Corporation, Yongin-si, Gyeonggi-do 446-729 (Korea, Republic of); Kim, Sang Hoon, E-mail: shkim@khu.ac.kr [Department of Biology, Research Institute for Basic Science, Kyung Hee University, Seoul 130-701 (Korea, Republic of)

2013-09-27

Highlights: •MRAP enhanced HSL expression. •ACTH-mediated MRAP reduced glycerol release. •PPARγ induced MRAP expression. •PPARγ bound to the MRAP promoter. -- Abstract: Adrenocorticotropic hormone (ACTH) in rodents decreases lipid accumulation and body weight. Melanocortin receptor 2 (MC2R) and MC2R accessory protein (MRAP) are specific receptors for ACTH in adipocytes. Peroxisome proliferator-activated receptor γ (PPARγ) plays a role in the transcriptional regulation of metabolic pathways such as adipogenesis and β-oxidation of fatty acids. In this study we investigated the transcriptional regulation of MRAP expression during differentiation of 3T3-L1 cells. Stimulation with ACTH affected lipolysis in murine mature adipocytes via MRAP. Putative peroxisome proliferator response element (PPRE) was identified in the MRAP promoter region. In chromatin immunoprecipitation and reporter assays, we observed binding of PPARγ to the MRAP promoter. The mutagenesis experiments showed that the −1209/−1198 region of the MRAP promoter could function as a PPRE site. These results suggest that PPARγ is required for transcriptional activation of the MRAP gene during adipogenesis, which contributes to understanding of the molecular mechanism of lipolysis in adipocytes.

Transcriptional activation of melanocortin 2 receptor accessory protein by PPARγ in adipocytes

International Nuclear Information System (INIS)

Kim, Nam Soo; Kim, Yoon-Jin; Cho, Si Young; Lee, Tae Ryong; Kim, Sang Hoon

2013-01-01

Highlights: •MRAP enhanced HSL expression. •ACTH-mediated MRAP reduced glycerol release. •PPARγ induced MRAP expression. •PPARγ bound to the MRAP promoter. -- Abstract: Adrenocorticotropic hormone (ACTH) in rodents decreases lipid accumulation and body weight. Melanocortin receptor 2 (MC2R) and MC2R accessory protein (MRAP) are specific receptors for ACTH in adipocytes. Peroxisome proliferator-activated receptor γ (PPARγ) plays a role in the transcriptional regulation of metabolic pathways such as adipogenesis and β-oxidation of fatty acids. In this study we investigated the transcriptional regulation of MRAP expression during differentiation of 3T3-L1 cells. Stimulation with ACTH affected lipolysis in murine mature adipocytes via MRAP. Putative peroxisome proliferator response element (PPRE) was identified in the MRAP promoter region. In chromatin immunoprecipitation and reporter assays, we observed binding of PPARγ to the MRAP promoter. The mutagenesis experiments showed that the −1209/−1198 region of the MRAP promoter could function as a PPRE site. These results suggest that PPARγ is required for transcriptional activation of the MRAP gene during adipogenesis, which contributes to understanding of the molecular mechanism of lipolysis in adipocytes

Transrepression of the estrogen receptor promoter by calcitriol in human breast cancer cells via two negative vitamin D response elements.

Science.gov (United States)

Swami, Srilatha; Krishnan, Aruna V; Peng, Lihong; Lundqvist, Johan; Feldman, David

2013-08-01

Calcitriol (1,25-dihydroxyvitamin D3), the hormonally active metabolite of vitamin D, exerts its anti-proliferative activity in breast cancer (BCa) cells by multiple mechanisms including the downregulation of the expression of estrogen receptor α (ER). We analyzed an ∼3.5 kb ER promoter sequence and demonstrated the presence of two potential negative vitamin D response elements (nVDREs), a newly identified putative nVDRE upstream at -2488 to -2473 bp (distal nVDRE) and a previously published sequence (proximal nVDRE) at -94 to -70 bp proximal to the P1 start site. Transactivation analysis using ER promoter deletion constructs and heterologous promoter-reporter constructs revealed that both nVDREs functioned to mediate calcitriol transrepression. In the electrophoretic mobility shift assay, the vitamin D receptor (VDR) showed strong binding to both nVDREs in the presence of calcitriol, and the chromatin immunoprecipitation assay demonstrated the recruitment of the VDR to the distal nVDRE site. Mutations in the 5' hexameric DNA sequence of the distal nVDRE resulted in the loss of calcitriol-mediated transrepression and the inhibition of protein-DNA complex formation, demonstrating the importance of these nucleotides in VDR DNA binding and transrepression. A putative nuclear factor-Y (NFY) binding site, identified within the distal nVDRE, led to the findings that NFY bound to the distal nVDRE site interfered with the binding of the VDR at the site and reduced calcitriol-mediated transrepression. In conclusion, the ER promoter region contains two negative VDREs that act in concert to bind to the VDR and both nVDREs are required for the maximal inhibition of ER expression by calcitriol. The suppression of ER expression and estrogen-mediated signaling by calcitriol in BCa cells suggests that vitamin D may be useful in the treatment of ER+ BCa.

Dioxin increases the interaction between aryl hydrocarbon receptor and estrogen receptor alpha at human promoters

DEFF Research Database (Denmark)

Ahmed, Shaaima; Valen, Eivind; Sandelin, Albin Gustav

2009-01-01

genes with little knowledge of what was occurring at other genomic regions. In this study, we showed using chromatin immunoprecipitation followed by hybridization to promoter focused microarrays (ChIP-chip) that 2,3,7,8-tetrachlorodibenzo-p-dioxin treatment significantly increased the overlap of genomic...... , suggesting that AHR was the important factor determining the recruitment of ER to these regions. RNA interference-mediated knockdown of AHR confirmed its requirement for the recruitment of ER to some, but not all, of the shared regions. Our findings demonstrate not only that dioxin induces the recruitment...

Selective androgen receptor modulators as function promoting therapies.

Science.gov (United States)

Bhasin, Shalender; Jasuja, Ravi

2009-05-01

The past decade has witnessed an unprecedented discovery effort to develop selective androgen receptor modulators (SARMs) that improve physical function and bone health without adversely affecting the prostate and cardiovascular outcomes. This review describes the historical evolution, the rationale for SARM development, and the mechanisms of testosterone action and SARM selectivity. Although steroidal SARMs have been around since the 1940s, a number of nonsteroidal SARMs that do not serve as substrates for CYP19 aromatase or 5alpha-reductase, act as full agonists in muscle and bone and as partial agonists in prostate are in development. The differing interactions of steroidal and nonsteroidal compounds with androgen receptor (AR) contribute to their unique pharmacologic actions. Ligand binding induces specific conformational changes in the ligand-binding domain, which could modulate surface topology and protein-protein interactions between AR and coregulators, resulting in tissue-specific gene regulation. Preclinical studies have demonstrated the ability of SARMs to increase muscle and bone mass in preclinical rodent models with varying degree of prostate sparing. Phase I trials of SARMs in humans have reported modest increments in fat-free mass. SARMs hold promise as a new class of function promoting anabolic therapies for a number of clinical indications, including functional limitations associated with aging and chronic disease, frailty, cancer cachexia, and osteoporosis.

Identification and annotation of promoter regions in microbial ...

Indian Academy of Sciences (India)

PRAKASH KUMAR

2007-06-15

Jun 15, 2007 ... It remains important, not only to detect rarely expressed genes but also for ... well as in identifying genes associated with rRNA, tRNA and ... DNA stability; free energy calculation; promoter; upstream and downstream region.

Excessive D1 Dopamine Receptor Activation in the Dorsal Striatum Promotes Autistic-Like Behaviors.

Science.gov (United States)

Lee, Yunjin; Kim, Hannah; Kim, Ji-Eun; Park, Jin-Young; Choi, Juli; Lee, Jung-Eun; Lee, Eun-Hwa; Han, Pyung-Lim

2018-07-01

The dopamine system has been characterized in motor function, goal-directed behaviors, and rewards. Recent studies recognize various dopamine system genes as being associated with autism spectrum disorder (ASD). However, how dopamine system dysfunction induces ASD pathophysiology remains unknown. In the present study, we demonstrated that mice with increased dopamine functions in the dorsal striatum via the suppression of dopamine transporter expression in substantia nigra neurons or the optogenetic stimulation of the nigro-striatal circuitry exhibited sociability deficits and repetitive behaviors relevant to ASD pathology in animal models, while these behavioral changes were blocked by a D1 receptor antagonist. Pharmacological activation of D1 dopamine receptors in normal mice or the genetic knockout (KO) of D2 dopamine receptors also produced typical autistic-like behaviors. Moreover, the siRNA-mediated inhibition of D2 dopamine receptors in the dorsal striatum was sufficient to replicate autistic-like phenotypes in D2 KO mice. Intervention of D1 dopamine receptor functions or the signaling pathways-related D1 receptors in D2 KO mice produced anti-autistic effects. Together, our results indicate that increased dopamine function in the dorsal striatum promotes autistic-like behaviors and that the dorsal striatum is the neural correlate of ASD core symptoms.

Characterization of the human Activin-A receptor type II-like kinase 1 (ACVRL1 promoter and its regulation by Sp1

Directory of Open Access Journals (Sweden)

Botella Luisa M

2010-06-01

Full Text Available Abstract Background Activin receptor-like kinase 1 (ALK1 is a Transforming Growth Factor-β (TGF-β receptor type I, mainly expressed in endothelial cells that plays a pivotal role in vascular remodelling and angiogenesis. Mutations in the ALK1 gene (ACVRL1 give rise to Hereditary Haemorrhagic Telangiectasia, a dominant autosomal vascular dysplasia caused by a haploinsufficiency mechanism. In spite of its patho-physiological relevance, little is known about the transcriptional regulation of ACVRL1. Here, we have studied the different origins of ACVRL1 transcription, we have analyzed in silico its 5'-proximal promoter sequence and we have characterized the role of Sp1 in the transcriptional regulation of ACVRL1. Results We have performed a 5'Rapid Amplification of cDNA Ends (5'RACE of ACVRL1 transcripts, finding two new transcriptional origins, upstream of the one previously described, that give rise to a new exon undiscovered to date. The 5'-proximal promoter region of ACVRL1 (-1,035/+210 was analyzed in silico, finding that it lacks TATA/CAAT boxes, but contains a remarkably high number of GC-rich Sp1 consensus sites. In cells lacking Sp1, ACVRL1 promoter reporters did not present any significant transcriptional activity, whereas increasing concentrations of Sp1 triggered a dose-dependent stimulation of its transcription. Moreover, silencing Sp1 in HEK293T cells resulted in a marked decrease of ACVRL1 transcriptional activity. Chromatin immunoprecipitation assays demonstrated multiple Sp1 binding sites along the proximal promoter region of ACVRL1 in endothelial cells. Furthermore, demethylation of CpG islands, led to an increase in ACVRL1 transcription, whereas in vitro hypermethylation resulted in the abolishment of Sp1-dependent transcriptional activation of ACVRL1. Conclusions Our results describe two new transcriptional start sites in ACVRL1 gene, and indicate that Sp1 is a key regulator of ACVRL1 transcription, providing new insights into

Adrenergic Agonists Bind to Adrenergic-Receptor-Like Regions of the Mu Opioid Receptor, Enhancing Morphine and Methionine-Enkephalin Binding: A New Approach to "Biased Opioids"?

Science.gov (United States)

Root-Bernstein, Robert; Turke, Miah; Subhramanyam, Udaya K Tiruttani; Churchill, Beth; Labahn, Joerg

2018-01-17

Extensive evidence demonstrates functional interactions between the adrenergic and opioid systems in a diversity of tissues and organs. While some effects are due to receptor and second messenger cross-talk, recent research has revealed an extracellular, allosteric opioid binding site on adrenergic receptors that enhances adrenergic activity and its duration. The present research addresses whether opioid receptors may have an equivalent extracellular, allosteric adrenergic binding site that has similar enhancing effects on opioid binding. Comparison of adrenergic and opioid receptor sequences revealed that these receptors share very significant regions of similarity, particularly in some of the extracellular and transmembrane regions associated with adrenergic binding in the adrenergic receptors. Five of these shared regions from the mu opioid receptor (muOPR) were synthesized as peptides and tested for binding to adrenergic, opioid and control compounds using ultraviolet spectroscopy. Adrenergic compounds bound to several of these muOPR peptides with low micromolar affinity while acetylcholine, histamine and various adrenergic antagonists did not. Similar studies were then conducted with purified, intact muOPR with similar results. Combinations of epinephrine with methionine enkephalin or morphine increased the binding of both by about half a log unit. These results suggest that muOPR may be allosterically enhanced by adrenergic agonists.

In silico comparative genomic analysis of GABAA receptor transcriptional regulation

Directory of Open Access Journals (Sweden)

Joyce Christopher J

2007-06-01

Full Text Available Abstract Background Subtypes of the GABAA receptor subunit exhibit diverse temporal and spatial expression patterns. In silico comparative analysis was used to predict transcriptional regulatory features in individual mammalian GABAA receptor subunit genes, and to identify potential transcriptional regulatory components involved in the coordinate regulation of the GABAA receptor gene clusters. Results Previously unreported putative promoters were identified for the β2, γ1, γ3, ε, θ and π subunit genes. Putative core elements and proximal transcriptional factors were identified within these predicted promoters, and within the experimentally determined promoters of other subunit genes. Conserved intergenic regions of sequence in the mammalian GABAA receptor gene cluster comprising the α1, β2, γ2 and α6 subunits were identified as potential long range transcriptional regulatory components involved in the coordinate regulation of these genes. A region of predicted DNase I hypersensitive sites within the cluster may contain transcriptional regulatory features coordinating gene expression. A novel model is proposed for the coordinate control of the gene cluster and parallel expression of the α1 and β2 subunits, based upon the selective action of putative Scaffold/Matrix Attachment Regions (S/MARs. Conclusion The putative regulatory features identified by genomic analysis of GABAA receptor genes were substantiated by cross-species comparative analysis and now require experimental verification. The proposed model for the coordinate regulation of genes in the cluster accounts for the head-to-head orientation and parallel expression of the α1 and β2 subunit genes, and for the disruption of transcription caused by insertion of a neomycin gene in the close vicinity of the α6 gene, which is proximal to a putative critical S/MAR.

Protein arginine methyltransferase 5 functions as an epigenetic activator of the androgen receptor to promote prostate cancer cell growth.

Science.gov (United States)

Deng, X; Shao, G; Zhang, H-T; Li, C; Zhang, D; Cheng, L; Elzey, B D; Pili, R; Ratliff, T L; Huang, J; Hu, C-D

2017-03-02

Protein arginine methyltransferase 5 (PRMT5) is an emerging epigenetic enzyme that mainly represses transcription of target genes via symmetric dimethylation of arginine residues on histones H4R3, H3R8 and H2AR3. Accumulating evidence suggests that PRMT5 may function as an oncogene to drive cancer cell growth by epigenetic inactivation of several tumor suppressors. Here, we provide evidence that PRMT5 promotes prostate cancer cell growth by epigenetically activating transcription of the androgen receptor (AR) in prostate cancer cells. Knockdown of PRMT5 or inhibition of PRMT5 by a specific inhibitor reduces the expression of AR and suppresses the growth of multiple AR-positive, but not AR-negative, prostate cancer cells. Significantly, knockdown of PRMT5 in AR-positive LNCaP cells completely suppresses the growth of xenograft tumors in mice. Molecular analysis reveals that PRMT5 binds to the proximal promoter region of the AR gene and contributes mainly to the enriched symmetric dimethylation of H4R3 in the same region. Mechanistically, PRMT5 is recruited to the AR promoter by its interaction with Sp1, the major transcription factor responsible for AR transcription, and forms a complex with Brg1, an ATP-dependent chromatin remodeler, on the proximal promoter region of the AR gene. Furthermore, PRMT5 expression in prostate cancer tissues is significantly higher than that in benign prostatic hyperplasia tissues, and PRMT5 expression correlates positively with AR expression at both the protein and mRNA levels. Taken together, our results identify PRMT5 as a novel epigenetic activator of AR in prostate cancer. Given that inhibiting AR transcriptional activity or androgen synthesis remains the major mechanism of action for most existing anti-androgen agents, our findings also raise an interesting possibility that targeting PRMT5 may represent a novel approach for prostate cancer treatment by eliminating AR expression.

Ku proteins function as corepressors to regulate farnesoid X receptor-mediated gene expression

International Nuclear Information System (INIS)

Ohno, Masae; Kunimoto, Masaaki; Nishizuka, Makoto; Osada, Shigehiro; Imagawa, Masayoshi

2009-01-01

The farnesoid X receptor (FXR; NR1H4) is a member of the nuclear receptor superfamily and regulates the expression of genes involved in enterohepatic circulation and the metabolism of bile acids. Based on functional analyses, nuclear receptors are divided into regions A-F. To explore the cofactors interacting with FXR, we performed a pull-down assay using GST-fused to the N-terminal A/B region and the C region, which are required for the ligand-independent transactivation and DNA-binding, respectively, of FXR, and nuclear extracts from HeLa cells. We identified DNA-dependent protein kinase catalytic subunit (DNA-PKcs), Ku80, and Ku70 as FXR associated factors. These proteins are known to have an important role in DNA repair, recombination, and transcription. DNA-PKcs mainly interacted with the A/B region of FXR, whereas the Ku proteins interacted with the C region and with the D region (hinge region). Chromatin immunoprecipitation assays revealed that the Ku proteins associated with FXR on the bile salt export pump (BSEP) promoter. Furthermore, we demonstrated that ectopic expression of the Ku proteins decreased the promoter activity and expression of BSEP gene mediated by FXR. These results suggest that the Ku proteins function as corepressors for FXR.

E1A FUNCTIONS AS A COACTIVATOR OF RETINOIC ACID-DEPENDENT RETINOIC ACID RECEPTOR-BETA-2 PROMOTER ACTIVATION

NARCIS (Netherlands)

KRUYT, FAE; FOLKERS, GE; WALHOUT, AJM; VANDERLEEDE, BM; VANDERSAAG, PT; Kruyt, Frank

The retinoic acid (RA) receptor (RAR) beta2 promoter is strongly activated by RA in embryonal carcinoma (EC) cells. We examined this activation in the P19 EC-derived END-2 cell line and in E1A-expressing counterparts and found strong RA-dependent RARbeta2 promoter activation in the E1A-expressing

Gelidium amansii promotes dendritic spine morphology and synaptogenesis, and modulates NMDA receptor-mediated postsynaptic current.

Science.gov (United States)

Hannan, Md Abdul; Mohibbullah, Md; Hong, Yong-Ki; Nam, Joo Hyun; Moon, Il Soo

2014-01-01

Neurotrophic factors are essential for the differentiation and maturation of developing neurons as well as providing survival support to the mature neurons. Moreover, therapeutically neurotrophic factors are promising to reconstruct partially damaged neuronal networks in neurodegenerative diseases. In the previous study, we reported that the ethanol extract of an edible marine alga, Gelidium amansii (GAE) had shown promising effects in the development and maturation of both axon and dendrites of hippocampal neurons. Here, we demonstrate that in primary culture of hippocampal neurons (1) GAE promotes a significant increase in the number of filopodia and dendritic spines; (2) promotes synaptogenesis; (3) enhances N-methyl-D-aspartic acid (NMDA) receptor recruitment; and (4) modulates NMDA-receptor-mediated postsynaptic current. Taken together these findings that GAE might be involved in both morphological and functional maturation of neurons suggest the possibility that GAE may constitute a promising candidate for novel compounds for the prevention and treatment of neurodegenerative diseases.

The genomic structure of the human UFO receptor.

Science.gov (United States)

Schulz, A S; Schleithoff, L; Faust, M; Bartram, C R; Janssen, J W

1993-02-01

Using a DNA transfection-tumorigenicity assay we have recently identified the UFO oncogene. It encodes a tyrosine kinase receptor characterized by the juxtaposition of two immunoglobulin-like and two fibronectin type III repeats in its extracellular domain. Here we describe the genomic organization of the human UFO locus. The UFO receptor is encoded by 20 exons that are distributed over a region of 44 kb. Different isoforms of UFO mRNA are generated by alternative splicing of exon 10 and differential usage of two imperfect polyadenylation sites resulting in the presence or absence of 1.5-kb 3' untranslated sequences. Primer extension and S1 nuclease analyses revealed multiple transcriptional initiation sites including a major site 169 bp upstream of the translation start site. The promoter region is GC rich, lacks TATA and CAAT boxes, but contains potential recognition sites for a variety of trans-acting factors, including Sp1, AP-2 and the cyclic AMP response element-binding protein. Proto-UFO and its oncogenic counterpart exhibit identical cDNA and promoter regions sequences. Possible modes of UFO activation are discussed.

Biological activity of cloned mammary tumor virus DNA fragments that bind purified glucocorticoid receptor protein in vitro

International Nuclear Information System (INIS)

Yamamoto, K.R.; Payvar, F.; Firestone, G.L.; Maler, B.A.; Wrange, O.; Carlstedt-Duke, J.; Gustafsson, J.A.; Chandler, V.L.; Karolinska Institutet, Stockholm, Sweden)

1983-01-01

To test whether high-affinity receptor:DNA interactions can be correlated with receptor effects on promoter function in vivo, we have mapped in greater detail the receptor-binding regions on murine mammary tumor virus DNA, using both nitrocellulose-filter binding and electron microscopy. Recombinant plasmids bearing these receptor-binding domains have been transfected into cultured cells, and the expression of the plasmid sequences has been monitored for hormonal regulation. The results are considered in terms of a speculative proposal that the glucocorticoid receptor may effect changes in promoter activity via specific alteration of chromatin and/or DNA structure. 37 references, 6 figures, 2 tables

Business networking for SMEs as a means to promote regional competitiveness: A Theoretical Framework

OpenAIRE

Vitor Braga

2004-01-01

The competitiveness of regions, as a means of promoting the competitiveness of a country as a whole, has been one of the main topics on the agenda of policy makers over the last decades. Several attempts at promoting competitiveness have been made with different degrees of success. In most cases, public investment in the regions was perceived as the solution to promote regional competitiveness and top-down policies were implemented. However, competitiveness also has an important dimension tha...

Identification of Insulin-Like Growth Factor-I Receptor (IGF-IR) Gene Promoter-Binding Proteins in Estrogen Receptor (ER)-Positive and ER-Depleted Breast Cancer Cells

International Nuclear Information System (INIS)

Sarfstein, Rive; Belfiore, Antonino; Werner, Haim

2010-01-01

The insulin-like growth factor I receptor (IGF-IR) has been implicated in the etiology of breast cancer. Overexpression of the IGF-IR gene is a typical feature of most primary breast cancers, whereas low IGF-IR levels are seen at advanced stages. Hence, evaluation of IGF-IR levels might be important for assessing prognosis. In the present study, we employed a proteomic approach based on DNA affinity chromatography followed either by mass spectroscopy (MS) or Western blot analysis to identify transcription factors that may associate with the IGF-IR promoter in estrogen receptor (ER)-positive and ER-depleted breast cancer cells. A biotinylated IGF-IR promoter fragment was bound to streptavidin magnetic beads and incubated with nuclear extracts of breast cancer cells. IGF-IR promoter-binding proteins were eluted with high salt and analyzed by MS and Western blots. Among the proteins that were found to bind to the IGF-IR promoter we identified zinc finger transcription factors Sp1 and KLF6, ER-α, p53, c-jun, and poly (ADP-ribosylation) polymerase. Furthermore, chromatin immune-precipitation (ChIP) analysis confirmed the direct in vivo binding of some of these transcription factors to IGF-IR promoter DNA. The functional relevance of binding data was assessed by cotransfection experiments with specific expression vectors along with an IGF-IR promoter reporter. In summary, we identified nuclear proteins that are potentially responsible for the differential expression of the IGF-IR gene in ER-positive and ER-depleted breast cancer cells

Identification of Insulin-Like Growth Factor-I Receptor (IGF-IR) Gene Promoter-Binding Proteins in Estrogen Receptor (ER)-Positive and ER-Depleted Breast Cancer Cells

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Sarfstein, Rive [Department of Human Molecular Genetics and Biochemistry, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978 (Israel); Belfiore, Antonino [Department of Clinical and Experimental Medicine, University Magna Graecia of Catanzaro, Catanzaro 88100 (Italy); Werner, Haim, E-mail: hwerner@post.tau.ac.il [Department of Human Molecular Genetics and Biochemistry, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978 (Israel)

2010-03-25

The insulin-like growth factor I receptor (IGF-IR) has been implicated in the etiology of breast cancer. Overexpression of the IGF-IR gene is a typical feature of most primary breast cancers, whereas low IGF-IR levels are seen at advanced stages. Hence, evaluation of IGF-IR levels might be important for assessing prognosis. In the present study, we employed a proteomic approach based on DNA affinity chromatography followed either by mass spectroscopy (MS) or Western blot analysis to identify transcription factors that may associate with the IGF-IR promoter in estrogen receptor (ER)-positive and ER-depleted breast cancer cells. A biotinylated IGF-IR promoter fragment was bound to streptavidin magnetic beads and incubated with nuclear extracts of breast cancer cells. IGF-IR promoter-binding proteins were eluted with high salt and analyzed by MS and Western blots. Among the proteins that were found to bind to the IGF-IR promoter we identified zinc finger transcription factors Sp1 and KLF6, ER-α, p53, c-jun, and poly (ADP-ribosylation) polymerase. Furthermore, chromatin immune-precipitation (ChIP) analysis confirmed the direct in vivo binding of some of these transcription factors to IGF-IR promoter DNA. The functional relevance of binding data was assessed by cotransfection experiments with specific expression vectors along with an IGF-IR promoter reporter. In summary, we identified nuclear proteins that are potentially responsible for the differential expression of the IGF-IR gene in ER-positive and ER-depleted breast cancer cells.

Multiple promoters drive tissue-specific expression of the human M2 muscarinic acetylcholine receptor gene

Czech Academy of Sciences Publication Activity Database

Krejčí, Alena; Bruce, A. W.; Doležal, Vladimír; Tuček, Stanislav; Buckley, N. J.

2004-01-01

Roč. 91, č. 1 (2004), s. 88-98 ISSN 0022-3042 R&D Projects: GA AV ČR IAA5011306 Institutional research plan: CEZ:AV0Z5011922 Keywords : M2 muscarinic receptor * neuron-restrictive silence factor * promoter Subject RIV: ED - Physiology Impact factor: 4.824, year: 2004

mTORC2 promotes type I insulin-like growth factor receptor and insulin receptor activation through the tyrosine kinase activity of mTOR.

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Yin, Yancun; Hua, Hui; Li, Minjing; Liu, Shu; Kong, Qingbin; Shao, Ting; Wang, Jiao; Luo, Yuanming; Wang, Qian; Luo, Ting; Jiang, Yangfu

2016-01-01

Mammalian target of rapamycin (mTOR) is a core component of raptor-mTOR (mTORC1) and rictor-mTOR (mTORC2) complexes that control diverse cellular processes. Both mTORC1 and mTORC2 regulate several elements downstream of type I insulin-like growth factor receptor (IGF-IR) and insulin receptor (InsR). However, it is unknown whether and how mTOR regulates IGF-IR and InsR themselves. Here we show that mTOR possesses unexpected tyrosine kinase activity and activates IGF-IR/InsR. Rapamycin induces the tyrosine phosphorylation and activation of IGF-IR/InsR, which is largely dependent on rictor and mTOR. Moreover, mTORC2 promotes ligand-induced activation of IGF-IR/InsR. IGF- and insulin-induced IGF-IR/InsR phosphorylation is significantly compromised in rictor-null cells. Insulin receptor substrate (IRS) directly interacts with SIN1 thereby recruiting mTORC2 to IGF-IR/InsR and promoting rapamycin- or ligand-induced phosphorylation of IGF-IR/InsR. mTOR exhibits tyrosine kinase activity towards the general tyrosine kinase substrate poly(Glu-Tyr) and IGF-IR/InsR. Both recombinant mTOR and immunoprecipitated mTORC2 phosphorylate IGF-IR and InsR on Tyr1131/1136 and Tyr1146/1151, respectively. These effects are independent of the intrinsic kinase activity of IGF-IR/InsR, as determined by assays on kinase-dead IGF-IR/InsR mutants. While both rictor and mTOR immunoprecitates from rictor(+/+) MCF-10A cells exhibit tyrosine kinase activity towards IGF-IR and InsR, mTOR immunoprecipitates from rictor(-/-) MCF-10A cells do not induce IGF-IR and InsR phosphorylation. Phosphorylation-deficient mutation of residue Tyr1131 in IGF-IR or Tyr1146 in InsR abrogates the activation of IGF-IR/InsR by mTOR. Finally, overexpression of rictor promotes IGF-induced cell proliferation. Our work identifies mTOR as a dual-specificity kinase and clarifies how mTORC2 promotes IGF-IR/InsR activation.

O-GlcNAcylation of Orphan Nuclear Receptor Estrogen-Related Receptor γ Promotes Hepatic Gluconeogenesis.

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Misra, Jagannath; Kim, Don-Kyu; Jung, Yoon Seok; Kim, Han Byeol; Kim, Yong-Hoon; Yoo, Eun-Kyung; Kim, Byung Gyu; Kim, Sunghoon; Lee, In-Kyu; Harris, Robert A; Kim, Jeong-Sun; Lee, Chul-Ho; Cho, Jin Won; Choi, Hueng-Sik

2016-10-01

Estrogen-related receptor γ (ERRγ) is a major positive regulator of hepatic gluconeogenesis. Its transcriptional activity is suppressed by phosphorylation signaled by insulin in the fed state, but whether posttranslational modification alters its gluconeogenic activity in the fasted state is not known. Metabolically active hepatocytes direct a small amount of glucose into the hexosamine biosynthetic pathway, leading to protein O-GlcNAcylation. In this study, we demonstrate that ERRγ is O-GlcNAcylated by O-GlcNAc transferase in the fasted state. This stabilizes the protein by inhibiting proteasome-mediated protein degradation, increasing ERRγ recruitment to gluconeogenic gene promoters. Mass spectrometry identifies two serine residues (S317, S319) present in the ERRγ ligand-binding domain that are O-GlcNAcylated. Mutation of these residues destabilizes ERRγ protein and blocks the ability of ERRγ to induce gluconeogenesis in vivo. The impact of this pathway on gluconeogenesis in vivo was confirmed by the observation that decreasing the amount of O-GlcNAcylated ERRγ by overexpressing the deglycosylating enzyme O-GlcNAcase decreases ERRγ-dependent glucose production in fasted mice. We conclude that O-GlcNAcylation of ERRγ serves as a major signal to promote hepatic gluconeogenesis. © 2016 by the American Diabetes Association.

Corticosteroid receptors adopt distinct cyclical transcriptional signatures.

Science.gov (United States)

Le Billan, Florian; Amazit, Larbi; Bleakley, Kevin; Xue, Qiong-Yao; Pussard, Eric; Lhadj, Christophe; Kolkhof, Peter; Viengchareun, Say; Fagart, Jérôme; Lombès, Marc

2018-05-07

Mineralocorticoid receptors (MRs) and glucocorticoid receptors (GRs) are two closely related hormone-activated transcription factors that regulate major pathophysiologic functions. High homology between these receptors accounts for the crossbinding of their corresponding ligands, MR being activated by both aldosterone and cortisol and GR essentially activated by cortisol. Their coexpression and ability to bind similar DNA motifs highlight the need to investigate their respective contributions to overall corticosteroid signaling. Here, we decipher the transcriptional regulatory mechanisms that underlie selective effects of MRs and GRs on shared genomic targets in a human renal cellular model. Kinetic, serial, and sequential chromatin immunoprecipitation approaches were performed on the period circadian protein 1 ( PER1) target gene, providing evidence that both receptors dynamically and cyclically interact at the same target promoter in a specific and distinct transcriptional signature. During this process, both receptors regulate PER1 gene by binding as homo- or heterodimers to the same promoter region. Our results suggest a novel level of MR-GR target gene regulation, which should be considered for a better and integrated understanding of corticosteroid-related pathophysiology.-Le Billan, F., Amazit, L., Bleakley, K., Xue, Q.-Y., Pussard, E., Lhadj, C., Kolkhof, P., Viengchareun, S., Fagart, J., Lombès, M. Corticosteroid receptors adopt distinct cyclical transcriptional signatures.

ING3 promotes prostate cancer growth by activating the androgen receptor.

Science.gov (United States)

Nabbi, Arash; McClurg, Urszula L; Thalappilly, Subhash; Almami, Amal; Mobahat, Mahsa; Bismar, Tarek A; Binda, Olivier; Riabowol, Karl T

2017-05-16

The androgen receptor (AR) is a major driver of prostate cancer, and increased AR levels and co-activators of the receptor promote the development of prostate cancer. INhibitor of Growth (ING) proteins target lysine acetyltransferase or lysine deacetylase complexes to the histone H3K4Me3 mark of active transcription, to affect chromatin structure and gene expression. ING3 is a stoichiometric member of the TIP60 lysine acetyltransferase complex implicated in prostate cancer development. Biopsies of 265 patients with prostate cancer were stained for ING3, pan-cytokeratin, and DNA. LNCaP and C4-2 androgen-responsive cells were used for in vitro assays including immunoprecipitation, western blotting, Luciferase reporter assay and quantitative polymerase chain reaction. Cell viability and migration assays were performed in prostate cancer cell lines using scrambled siRNA or siRNA targeting ING3. We find that ING3 levels and AR activity positively correlate in prostate cancer. ING3 potentiates androgen effects, increasing expression of androgen-regulated genes and androgen response element-driven reporters to promote growth and anchorage-independent growth. Conversely, ING3 knockdown inhibits prostate cancer cell growth and invasion. ING3 activates the AR by serving as a scaffold to increase interaction between TIP60 and the AR in the cytoplasm, enhancing receptor acetylation and translocation to the nucleus. Activation is independent of ING3's ability to target the TIP60 complex to H3K4Me3, identifying a previously unknown chromatin-independent cytoplasmic activity for ING3. In agreement with in vitro observations, analysis of The Cancer Genome Atlas (TCGA) data (n = 498) and a prostate cancer tissue microarray (n = 256) show that ING3 levels are higher in aggressive prostate cancers, with high levels of ING3 predicting shorter patient survival in a low AR subgroup. Including ING3 levels with currently used indicators such as the Gleason score provides more

Adrenergic Agonists Bind to Adrenergic-Receptor-Like Regions of the Mu Opioid Receptor, Enhancing Morphine and Methionine-Enkephalin Binding: A New Approach to “Biased Opioids”?

Science.gov (United States)

Turke, Miah; Subhramanyam, Udaya K. Tiruttani; Churchill, Beth; Labahn, Joerg

2018-01-01

Extensive evidence demonstrates functional interactions between the adrenergic and opioid systems in a diversity of tissues and organs. While some effects are due to receptor and second messenger cross-talk, recent research has revealed an extracellular, allosteric opioid binding site on adrenergic receptors that enhances adrenergic activity and its duration. The present research addresses whether opioid receptors may have an equivalent extracellular, allosteric adrenergic binding site that has similar enhancing effects on opioid binding. Comparison of adrenergic and opioid receptor sequences revealed that these receptors share very significant regions of similarity, particularly in some of the extracellular and transmembrane regions associated with adrenergic binding in the adrenergic receptors. Five of these shared regions from the mu opioid receptor (muOPR) were synthesized as peptides and tested for binding to adrenergic, opioid and control compounds using ultraviolet spectroscopy. Adrenergic compounds bound to several of these muOPR peptides with low micromolar affinity while acetylcholine, histamine and various adrenergic antagonists did not. Similar studies were then conducted with purified, intact muOPR with similar results. Combinations of epinephrine with methionine enkephalin or morphine increased the binding of both by about half a log unit. These results suggest that muOPR may be allosterically enhanced by adrenergic agonists. PMID:29342106

Adrenergic Agonists Bind to Adrenergic-Receptor-Like Regions of the Mu Opioid Receptor, Enhancing Morphine and Methionine-Enkephalin Binding: A New Approach to “Biased Opioids”?

Directory of Open Access Journals (Sweden)

Robert Root-Bernstein

2018-01-01

Full Text Available Extensive evidence demonstrates functional interactions between the adrenergic and opioid systems in a diversity of tissues and organs. While some effects are due to receptor and second messenger cross-talk, recent research has revealed an extracellular, allosteric opioid binding site on adrenergic receptors that enhances adrenergic activity and its duration. The present research addresses whether opioid receptors may have an equivalent extracellular, allosteric adrenergic binding site that has similar enhancing effects on opioid binding. Comparison of adrenergic and opioid receptor sequences revealed that these receptors share very significant regions of similarity, particularly in some of the extracellular and transmembrane regions associated with adrenergic binding in the adrenergic receptors. Five of these shared regions from the mu opioid receptor (muOPR were synthesized as peptides and tested for binding to adrenergic, opioid and control compounds using ultraviolet spectroscopy. Adrenergic compounds bound to several of these muOPR peptides with low micromolar affinity while acetylcholine, histamine and various adrenergic antagonists did not. Similar studies were then conducted with purified, intact muOPR with similar results. Combinations of epinephrine with methionine enkephalin or morphine increased the binding of both by about half a log unit. These results suggest that muOPR may be allosterically enhanced by adrenergic agonists.

Control of transcriptional repression of the vitellogenin receptor gene in largemouth bass (Micropterus salmoides) by select estrogen receptors isotypes.

Science.gov (United States)

Dominguez, Gustavo A; Bisesi, Joseph H; Kroll, Kevin J; Denslow, Nancy D; Sabo-Attwood, Tara

2014-10-01

The vitellogenin receptor (Vtgr) plays an important role in fish reproduction. This receptor functions to incorporate vitellogenin (Vtg), a macromolecule synthesized and released from the liver in the bloodstream, into oocytes where it is processed into yolk. Although studies have focused on the functional role of Vtgr in fish, the mechanistic control of this gene is still unexplored. Here we report the identification and analysis of the first piscine 5' regulatory region of the vtgr gene which was cloned from largemouth bass (Micropterus salmoides). Using this putative promoter sequence, we investigated a role for hormones, including insulin and 17β-estradiol (E2), in transcriptional regulation through cell-based reporter assays. No effect of insulin was observed, however, E2 was able to repress transcriptional activity of the vtgr promoter through select estrogen receptor subtypes, Esr1 and Esr2a but not Esr2b. Electrophoretic mobility shift assay demonstrated that Esr1 likely interacts with the vtgr promoter region through half ERE and/or SP1 sites, in part. Finally we also show that ethinylestradiol (EE2), but not bisphenol-A (BPA), represses promoter activity similarly to E2. These results reveal for the first time that the Esr1 isoform may play an inhibitory role in the expression of LMB vtgr mRNA under the influence of E2, and potent estrogens such as EE2. In addition, this new evidence suggests that vtgr may be a target of select endocrine disrupting compounds through environmental exposures. © The Author 2014. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Cell-cell adhesion mediated by binding of membrane-anchored transforming growth factor α to epidermal growth factor receptors promotes cell proliferation

International Nuclear Information System (INIS)

Anklesaria, P.; Greenberger, J.S.; Teixido, J.; Laiho, M.; Massague, J.; Pierce, J.H.

1990-01-01

The precursor for transforming growth factor α, pro-TGF-α, is a cell surface glycoprotein that can establish contact with epidermal growth factor (EGF) receptors on adjacent cells. To examine whether the pro-TGF-α/EGF receptor pair can simultaneously mediate cell adhesion and promote cell proliferation, the authors have expressed pro-TGF-α in a bone marrow stromal cell line labeled with [ 35 S] cysteine. Expression of pro-TGF-α allows these cells to support long-term attachment of an EGF/interleukin-3-dependent hematopoietic progenitor cell line that expresses EGF receptors but is unable to adhere to normal stroma. This interaction is inhibited by soluble EGF receptor ligands. Further, the hematopoietic progenitor cells replicate their DNA while they are attached to the stromal cell layer and become foci of sustained cell proliferation. Thus, pro-TGF-α and the EGF receptor can function as mediators of intercellular adhesion and this interaction may promote a mitogenic response. They propose the term juxtacrine to designate this form of stimulation between adjacent cells

Analysis of tissue-specific region in sericin 1 gene promoter of Bombyx mori

Energy Technology Data Exchange (ETDEWEB)

Yan, Liu [College of Biomedical Engineering and Instrument Science, Zhejiang University, Hangzhou 310027 (China); Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Lian, Yu [College of Biomedical Engineering and Instrument Science, Zhejiang University, Hangzhou 310027 (China); Zhejiang Province Key Laboratory of Preventive Veterinary Medicine, Institute of Preventive Veterinary Medicine, Zhejiang University, Hangzhou 310029 (China); Xiuyang, Guo [Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Tingqing, Guo [Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Shengpeng, Wang [Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Changde, Lu [Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China)

2006-03-31

The gene encoding sericin 1 (Ser1) of silkworm (Bombyx mori) is specifically expressed in the middle silk gland cells. To identify element involved in this transcription-dependent spatial restriction, truncation of the 5' terminal from the sericin 1 (Ser1) promoter is studied in vivo. A 209 bp DNA sequence upstream of the transcriptional start site (-586 to -378) is found to be responsible for promoting tissue-specific transcription. Analysis of this 209 bp region by overlapping deletion studies showed that a 25 bp region (-500 to -476) suppresses the ectopic expression of the Ser1 promoter. An unknown factor abundant in fat body nuclear extracts is shown to bind to this 25 bp fragment. These results suggest that this 25 bp region and the unknown factor are necessary for determining the tissue-specificity of the Ser1 promoter.

P2Y2 Receptor and EGFR Cooperate to Promote Prostate Cancer Cell Invasion via ERK1/2 Pathway.

Science.gov (United States)

Li, Wei-Hua; Qiu, Ying; Zhang, Hong-Quan; Tian, Xin-Xia; Fang, Wei-Gang

2015-01-01

As one member of G protein-coupled P2Y receptors, P2Y2 receptor can be equally activated by extracellular ATP and UTP. Our previous studies have proved that activation of P2Y2 receptor by extracellular ATP could promote prostate cancer cell invasion and metastasis in vitro and in vivo via regulating the expressions of some epithelial-mesenchymal transition/invasion-related genes (including IL-8, E-cadherin, Snail and Claudin-1), and the most significant change in expression of IL-8 was observed after P2Y2 receptor activation. However, the signaling pathway downstream of P2Y2 receptor and the role of IL-8 in P2Y2-mediated prostate cancer cell invasion remain unclear. Here, we found that extracellular ATP/UTP induced activation of EGFR and ERK1/2. After knockdown of P2Y2 receptor, the ATP -stimulated phosphorylation of EGFR and ERK1/2 was significantly suppressed. Further experiments showed that inactivation of EGFR and ERK1/2 attenuated ATP-induced invasion and migration, and suppressed ATP-mediated IL-8 production. In addition, knockdown of IL-8 inhibited ATP-mediated invasion and migration of prostate cancer cells. These findings suggest that P2Y2 receptor and EGFR cooperate to upregulate IL-8 production via ERK1/2 pathway, thereby promoting prostate cancer cell invasion and migration. Thus blocking of the P2Y2-EGFR-ERK1/2 pathway may provide effective therapeutic interventions for prostate cancer.

Lack of death receptor 4 (DR4) expression through gene promoter methylation in gastric carcinoma.

Science.gov (United States)

Lee, Kyung Hwa; Lim, Sang Woo; Kim, Ho Gun; Kim, Dong Yi; Ryu, Seong Yeob; Joo, Jae Kyun; Kim, Jung Chul; Lee, Jae Hyuk

2009-07-01

To determine the underlying mechanism for the differential expression, the extent of promoter methylation in tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-related genes acting downstream of TRAIL was examined in early and advanced gastric carcinomas. The extent of promoter methylation in the DR4, DR5, DcR1, DcR2, and CASP8 genes was quantified using bisulfite modification and methylation-specific polymerase chain reaction. The promoters for DcR1, DcR2, and CASP8 were largely unmethylated in early gastric carcinoma, advanced gastric carcinoma, and controls, with no significant difference among them. Protein levels of DR4, DcR1, and DcR2 as revealed by immunohistochemistry correlated with the extent of the respective promoter methylation (P < 0.05 in all cases). Hypomethylation, rather than hypermethylation, of the DR4 promoter was noted in invasive gastric malignancies, with statistical significance (P = 0.003). The promoter methylation status of TRAIL receptors in gastric carcinoma may have clinical implications for improving therapeutic strategies in patients with gastric carcinoma.

Region-selective effects of neuroinflammation and antioxidant treatment on peripheral benzodiazepine receptors and NMDA receptors in the rat brain

Energy Technology Data Exchange (ETDEWEB)

Biegon, A.; Alvarado, M.; Budinger, T.F.; Grossman, R.; Hensley, K.; West, M.S.; Kotake, Y.; Ono, M.; Floyd, R.A.

2001-12-10

Following induction of acute neuroinflammation by intracisternal injection of endotoxin (lipopolysaccharide) in rats, quantitative autoradiography was used to assess the regional level of microglial activation and glutamate (NMDA) receptor binding. The possible protective action of the antioxidant phenyl-tert-butyl nitrone in this model was tested by administering the drug in the drinking water for 6 days starting 24 hours after endotoxin injection. Animals were killed 7 days post-injection and consecutive cryostat brain sections labeled with [3H]PK11195 as a marker of activated microglia and [125I]iodoMK801 as a marker of the open-channel, activated state of NMDA receptors. Lipopolysaccharide increased [3H]PK11195 binding in the brain, with the largest increases (2-3 fold) in temporal and entorhinal cortex, hippocampus, and substantia innominata. A significant (>50 percent) decrease in [125I]iodoMK801 binding was found in the same brain regions. Phenyl-tert-butyl nitrone treatment resulted in a partial inhibition ({approx}25 percent decrease) of the lipopolysaccharide-induced increase in [3H]PK11195 binding but completely reversed the lipopolysaccharide-induced decrease in [125I]iodoMK80 binding in the entorhinal cortex, hippocampus, and substantia innominata. Loss of NMDA receptor function in cortical and hippocampal regions may contribute to the cognitive deficits observed in diseases with a neuroinflammatory component, such as meningitis or Alzheimer's disease.

Perspectives on Promoting Regional Renewable Energy Research and Development

International Nuclear Information System (INIS)

Dresselhaus, M.

2008-01-01

Recent discussions at the Washington International Renewable Energy Conference (WIREC), hosted in March 2008 by the United States Government, with nearly 9000 participants including 103 ministers from 126 countries, concluded that a major acceleration in the adoption of renewable energy technologies was needed by mid-century. Because of different climatic conditions and societal preferences, regional cooperation is expected to play a major role in the efficient adoption of appropriate renewable energy technologies, and countries with special expertise in specific technologies seem eager to collaborate internationally to promote global goals in renewable energy. A review will be given of what we learned from this conference about renewable energy research and development strategies with a special focus given to using this basic knowledge base to promote the development of renewable energy technologies appropriate to specific regions of the world.(author)

Notch receptor expression in neurogenic regions of the adult zebrafish brain.

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Vanessa de Oliveira-Carlos

Full Text Available The adult zebrash brain has a remarkable constitutive neurogenic capacity. The regulation and maintenance of its adult neurogenic niches are poorly understood. In mammals, Notch signaling is involved in stem cell maintenance both in embryonic and adult CNS. To better understand how Notch signaling is involved in stem cell maintenance during adult neurogenesis in zebrafish we analysed Notch receptor expression in five neurogenic zones of the adult zebrafish brain. Combining proliferation and glial markers we identified several subsets of Notch receptor expressing cells. We found that 90 [Formula: see text] of proliferating radial glia express notch1a, notch1b and notch3. In contrast, the proliferating non-glial populations of the dorsal telencephalon and hypothalamus rarely express notch3 and about half express notch1a/1b. In the non-proliferating radial glia notch3 is the predominant receptor throughout the brain. In the ventral telencephalon and in the mitotic area of the optic tectum, where cells have neuroepithelial properties, notch1a/1b/3 are expressed in most proliferating cells. However, in the cerebellar niche, although progenitors also have neuroepithelial properties, only notch1a/1b are expressed in a high number of PCNA [Formula: see text] cells. In this region notch3 expression is mostly in Bergmann glia and at low levels in few PCNA [Formula: see text] cells. Additionally, we found that in the proliferation zone of the ventral telencephalon, Notch receptors display an apical high to basal low gradient of expression. Notch receptors are also expressed in subpopulations of oligodendrocytes, neurons and endothelial cells. We suggest that the partial regional heterogeneity observed for Notch expression in progenitor cells might be related to the cellular diversity present in each of these neurogenic niches.

Tamoxifen induces the expression of maspin through estrogen receptor-alpha.

Science.gov (United States)

Liu, Zesheng; Shi, Heidi Y; Nawaz, Zafar; Zhang, Ming

2004-06-08

Maspin (mammary serine protease inhibitor) is a tumor suppressor gene that plays an important role in inhibiting tumor growth, invasion and metastasis. Maspin expression is down regulated at transcription level in primary and metastatic breast tumor cells. Previous studies on hormonal regulation of maspin prompt us to test whether an estrogen antagonist tamoxifen (TAM) can exert its anti-tumor function by up regulating maspin gene expression. For this purpose, we first tested whether maspin promoter could be activated in normal and several breast tumor cells. We then carried out a series of promoter analysis in which estrogen receptors and TAM were reconstituted in an in vitro cell culture system. Here we report our new finding that tumor suppresser gene maspin is one of the TAM target genes. TAM induces a maspin/luciferase reporter in cell culture and this induction requires the presence of (estrogen receptor alpha) ERalpha but not estrogen receptor-beta (ERbeta). Maspin promoter deletion and mutation analysis showed that the cis element(s) within a region between -90and+87 bp but not the HRE site (-272 bp) was involved in TAM induction of maspin expression. TAM bound ERalpha may directly control maspin gene expression through the interaction with cofactor (s). Analysis using several ERalpha mutants showed that the N-terminal A/B motif (AF-1) was critical for maspin basal level transcription activation. An ERalpha mutant with point mutations at DNA binding domain abolished estrogen induction of an ERE-luciferase reporter but was still active in activating maspin promoter by TAM. LBD-AF2 domain was required for ERalpha-dependent TAM induction. Deletion of LBD-AF2 or a point mutation in the ERalpha LBD-AF2 region (LBDmtL539A) completely abolished the activation of maspin promoter, suggesting that TAM induction of maspin involves the recruitment of cofactor(s) by ERalpha to the maspin promoter region. This finding indicates that one of the pathways for cancer

Dynamic mobility of functional GABAA receptors at inhibitory synapses.

Science.gov (United States)

Thomas, Philip; Mortensen, Martin; Hosie, Alastair M; Smart, Trevor G

2005-07-01

Importing functional GABAA receptors into synapses is fundamental for establishing and maintaining inhibitory transmission and for controlling neuronal excitability. By introducing a binding site for an irreversible inhibitor into the GABAA receptor alpha1 subunit channel lining region that can be accessed only when the receptor is activated, we have determined the dynamics of receptor mobility between synaptic and extrasynaptic locations in hippocampal pyramidal neurons. We demonstrate that the cell surface GABAA receptor population shows no fast recovery after irreversible inhibition. In contrast, after selective inhibition, the synaptic receptor population rapidly recovers by the import of new functional entities within minutes. The trafficking pathways that promote rapid importation of synaptic receptors do not involve insertion from intracellular pools, but reflect receptor diffusion within the plane of the membrane. This process offers the synapse a rapid mechanism to replenish functional GABAA receptors at inhibitory synapses and a means to control synaptic efficacy.

Regional differences in gene expression and promoter usage in aged human brains

KAUST Repository

Pardo, Luba M.

2013-02-19

To characterize the promoterome of caudate and putamen regions (striatum), frontal and temporal cortices, and hippocampi from aged human brains, we used high-throughput cap analysis of gene expression to profile the transcription start sites and to quantify the differences in gene expression across the 5 brain regions. We also analyzed the extent to which methylation influenced the observed expression profiles. We sequenced more than 71 million cap analysis of gene expression tags corresponding to 70,202 promoter regions and 16,888 genes. More than 7000 transcripts were differentially expressed, mainly because of differential alternative promoter usage. Unexpectedly, 7% of differentially expressed genes were neurodevelopmental transcription factors. Functional pathway analysis on the differentially expressed genes revealed an overrepresentation of several signaling pathways (e.g., fibroblast growth factor and wnt signaling) in hippocampus and striatum. We also found that although 73% of methylation signals mapped within genes, the influence of methylation on the expression profile was small. Our study underscores alternative promoter usage as an important mechanism for determining the regional differences in gene expression at old age.

Growing Significance of EU Institutions in Promotion of Inter-regional policies

Directory of Open Access Journals (Sweden)

Ella V. Ermakova

2014-01-01

Full Text Available The article explores the variety of tools and vehicles applied within the EU to expand the prerogative of the regions of the EU member states. The author uses as an example the inter-regional policies in Belgium in respect of the Flemish Region and the Walloon Region. The author analyzes the mechanisms of promotion of external regional relations in Belgium as a means of addressing different problems both on national and all-European level, supporting the arguments and conclusions by examples of relevant EU initiatives. The article details the activities of the EU Regional Committee (RC, the EU advisory body with the powers of political initiative, upholding the principle ofsubsidarity in the implementation of the EU member states' regional policies. The involvement of the Flemish Region and the Walloon Region in the activities of EU RC is described and summarized. As a case study, the article deals with Belgium's rotating six months presidency in the EUin 2010 when the country, which was going through a severe political crisis with no federal government in place, was represented by the two regions. The special focus of the article is on the strategic EU program "Europe2020" and its implementation by the regions of Belgium. There is an account of the initiatives undertaken by the Flemish Region and the Walloon Region within the framework of this program outlining the interaction of the two regions. The author provides a comprehensive analysis of the involvement of the Flemish Region and the Walloon Region with various EU institutions describing how each party achieves the promotion of its regional interests. Within this context, it is a noteworthy development that the Flemish Region is participating in the international program "Pact 2020" on energy all by its own. The article features quotations by Flemish and Walloon political figures which serve as an illustration of the prevailing attitudes in the Belgian society to the process of

Zebrafish GDNF and its co-receptor GFRα1 activate the human RET receptor and promote the survival of dopaminergic neurons in vitro.

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Tuulia Saarenpää

Full Text Available Glial cell line-derived neurotrophic factor (GDNF is a ligand that activates, through co-receptor GDNF family receptor alpha-1 (GFRα1 and receptor tyrosine kinase "RET", several signaling pathways crucial in the development and sustainment of multiple neuronal populations. We decided to study whether non-mammalian orthologs of these three proteins have conserved their function: can they activate the human counterparts? Using the baculovirus expression system, we expressed and purified Danio rerio RET, and its binding partners GFRα1 and GDNF, and Drosophila melanogaster RET and two isoforms of co-receptor GDNF receptor-like. Our results report high-level insect cell expression of post-translationally modified and dimerized zebrafish RET and its binding partners. We also found that zebrafish GFRα1 and GDNF are comparably active as mammalian cell-produced ones. We also report the first measurements of the affinity of the complex to RET in solution: at least for zebrafish, the Kd for GFRα1-GDNF binding RET is 5.9 μM. Surprisingly, we also found that zebrafish GDNF as well as zebrafish GFRα1 robustly activated human RET signaling and promoted the survival of cultured mouse dopaminergic neurons with comparable efficiency to mammalian GDNF, unlike E. coli-produced human proteins. These results contradict previous studies suggesting that mammalian GFRα1 and GDNF cannot bind and activate non-mammalian RET and vice versa.

The Toll-like receptor 5 ligand flagellin promotes asthma by priming allergic responses to indoor allergens.

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Wilson, Rhonda H; Maruoka, Shuichiro; Whitehead, Gregory S; Foley, Julie F; Flake, Gordon P; Sever, Michelle L; Zeldin, Darryl C; Kraft, Monica; Garantziotis, Stavros; Nakano, Hideki; Cook, Donald N

2012-11-01

Allergic asthma is a complex disease characterized by eosinophilic pulmonary inflammation, mucus production and reversible airway obstruction. Exposure to indoor allergens is a risk factor for asthma, but this disease is also associated with high household levels of total and particularly Gram-negative bacteria. The ability of bacterial products to act as adjuvants suggests they might promote asthma by priming allergic sensitization to inhaled allergens. In support of this idea, house dust extracts (HDEs) can activate antigen-presenting dendritic cells (DCs) in vitro and promote allergic sensitization to inhaled innocuous proteins in vivo. It is unknown which microbial products provide most of the adjuvant activity in HDEs. A screen for adjuvant activity of microbial products revealed that the bacterial protein flagellin (FLA) stimulated strong allergic airway responses to an innocuous inhaled protein, ovalbumin (OVA). Moreover, Toll-like receptor 5 (TLR5), the mammalian receptor for FLA, was required for priming strong allergic responses to natural indoor allergens present in HDEs. In addition, individuals with asthma have higher serum levels of FLA-specific antibodies as compared to nonasthmatic individuals. Together, these findings suggest that household FLA promotes the development of allergic asthma by TLR5-dependent priming of allergic responses to indoor allergens.

Role of the hinge region of glucocorticoid receptor for HEXIM1-mediated transcriptional repression

International Nuclear Information System (INIS)

Yoshikawa, Noritada; Shimizu, Noriaki; Sano, Motoaki; Ohnuma, Kei; Iwata, Satoshi; Hosono, Osamu; Fukuda, Keiichi; Morimoto, Chikao

2008-01-01

We previously reported that HEXIM1 (hexamethylene bisacetamide-inducible protein 1), which suppresses transcription elongation via sequestration of positive transcription elongation factor b (P-TEFb) using 7SK RNA as a scaffold, directly associates with glucocorticoid receptor (GR) to suppress glucocorticoid-inducible gene activation. Here, we revealed that the hinge region of GR is essential for its interaction with HEXIM1, and that oxosteroid receptors including GR show sequence homology in their hinge region and interact with HEXIM1, whereas the other members of nuclear receptors do not. We also showed that HEXIM1 suppresses GR-mediated transcription in two ways: sequestration of P-TEFb by HEXIM1 and direct interaction between GR and HEXIM1. In contrast, peroxisome proliferator-activated receptor γ-dependent gene expression is negatively modulated by HEXIM1 solely via sequestration of P-TEFb. We, therefore, conclude that HEXIM1 may act as a gene-selective transcriptional regulator via direct interaction with certain transcriptional regulators including GR and contribute to fine-tuning of, for example, glucocorticoid-mediated biological responses

Systematic screening for mutations in the promoter and the coding region of the 5-HT{sub 1A} gene

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Erdmann, J.; Shimron-Abarbanell, D.; Cichon, S. [Univ. of Bonn (Germany)] [and others

1995-10-09

In the present study we sought to identify genetic variation in the 5-HT{sub 1A} receptor gene which through alteration of protein function or level of expression might contribute to the genetic predisposition to neuropsychiatric diseases. Genomic DNA samples from 159 unrelated subjects (including 45 schizophrenic, 46 bipolar affective, and 43 patients with Tourette`s syndrome, as well as 25 healthy controls) were investigated by single-strand conformation analysis. Overlapping PCR (polymerase chain reaction) fragments covered the whole coding sequence as well as the 5{prime} untranslated region of the 5-HT{sub 1A} gene. The region upstream to the coding sequence we investigated contains a functional promoter. We found two rare nucleotide sequence variants. Both mutations are located in the coding region of the gene: a coding mutation (A{yields}G) in nucleotide position 82 which leads to an amino acid exchange (Ile{yields}Val) in position 28 of the receptor protein and a silent mutation (C{yields}T) in nucleotide position 549. The occurrence of the Ile-28-Val substitution was studied in an extended sample of patients (n = 352) and controls (n = 210) but was found in similar frequencies in all groups. Thus, this mutation is unlikely to play a significant role in the genetic predisposition to the diseases investigated. In conclusion, our study does not provide evidence that the 5-HT{sub 1A} gene plays either a major or a minor role in the genetic predisposition to schizophrenia, bipolar affective disorder, or Tourette`s syndrome. 29 refs., 4 figs., 1 tab.

Regional Delivery of Chimeric Antigen Receptor (CAR) T-Cells for Cancer Therapy.

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Sridhar, Praveen; Petrocca, Fabio

2017-07-18

Chimeric Antigen Receptor (CAR) T-cells are T-cells with recombinant receptors targeted to tumor antigens. CAR-T cell therapy has emerged as a mode of immunotherapy and is now being extensively explored in hematologic cancer. In contrast, CAR-T cell use in solid tumors has been hampered by multiple obstacles. Several approaches have been taken to circumvent these obstacles, including the regional delivery of CAR-T cells. Regional CAR-T cell delivery can theoretically compensate for poor T-cell trafficking and tumor antigen specificity while avoiding systemic toxicity associated with intravenous delivery. We reviewed completed clinical trials for the treatment of glioblastoma and metastatic colorectal cancer and examined the data in these studies for safety, efficacy, and potential advantages that regional delivery may confer over systemic delivery. Our appraisal of the available literature revealed that regional delivery of CAR-T cells in both glioblastoma and hepatic colorectal metastases was generally well tolerated and efficacious in select instances. We propose that the regional delivery of CAR-T cells is an area of potential growth in the solid tumor immunotherapy, and look towards future clinical trials in head and neck cancer, mesothelioma, and peritoneal carcinomatosis as the use of this technique expands.

Regional Delivery of Chimeric Antigen Receptor (CAR T-Cells for Cancer Therapy

Directory of Open Access Journals (Sweden)

Praveen Sridhar

2017-07-01

Full Text Available Chimeric Antigen Receptor (CAR T-cells are T-cells with recombinant receptors targeted to tumor antigens. CAR-T cell therapy has emerged as a mode of immunotherapy and is now being extensively explored in hematologic cancer. In contrast, CAR-T cell use in solid tumors has been hampered by multiple obstacles. Several approaches have been taken to circumvent these obstacles, including the regional delivery of CAR-T cells. Regional CAR-T cell delivery can theoretically compensate for poor T-cell trafficking and tumor antigen specificity while avoiding systemic toxicity associated with intravenous delivery. We reviewed completed clinical trials for the treatment of glioblastoma and metastatic colorectal cancer and examined the data in these studies for safety, efficacy, and potential advantages that regional delivery may confer over systemic delivery. Our appraisal of the available literature revealed that regional delivery of CAR-T cells in both glioblastoma and hepatic colorectal metastases was generally well tolerated and efficacious in select instances. We propose that the regional delivery of CAR-T cells is an area of potential growth in the solid tumor immunotherapy, and look towards future clinical trials in head and neck cancer, mesothelioma, and peritoneal carcinomatosis as the use of this technique expands.

Refinement of glucagon-like peptide 1 docking to its intact receptor using mid-region photolabile probes and molecular modeling.

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Miller, Laurence J; Chen, Quan; Lam, Polo C-H; Pinon, Delia I; Sexton, Patrick M; Abagyan, Ruben; Dong, Maoqing

2011-05-06

The glucagon-like peptide 1 (GLP1) receptor is an important drug target within the B family of G protein-coupled receptors. Its natural agonist ligand, GLP1, has incretin-like actions and the receptor is a recognized target for management of type 2 diabetes mellitus. Despite recent solution of the structure of the amino terminus of the GLP1 receptor and several close family members, the molecular basis for GLP1 binding to and activation of the intact receptor remains unclear. We previously demonstrated molecular approximations between amino- and carboxyl-terminal residues of GLP1 and its receptor. In this work, we study spatial approximations with the mid-region of this peptide to gain insights into the orientation of the intact receptor and the ligand-receptor complex. We have prepared two new photolabile probes incorporating a p-benzoyl-l-phenylalanine into positions 16 and 20 of GLP1(7-36). Both probes bound to the GLP1 receptor specifically and with high affinity. These were each fully efficacious agonists, stimulating cAMP accumulation in receptor-bearing CHO cells in a concentration-dependent manner. Each probe specifically labeled a single receptor site. Protease cleavage and radiochemical sequencing identified receptor residue Leu(141) above transmembrane segment one as its site of labeling for the position 16 probe, whereas the position 20 probe labeled receptor residue Trp(297) within the second extracellular loop. Establishing ligand residue approximation with this loop region is unique among family members and may help to orient the receptor amino-terminal domain relative to its helical bundle region.

Epigenetic regulation of the glucocorticoid receptor promoter 1(7) in adult rats.

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Witzmann, Simone R; Turner, Jonathan D; Mériaux, Sophie B; Meijer, Onno C; Muller, Claude P

2012-11-01

Regulation of glucocorticoid receptor (GR) levels is an important stress adaptation mechanism. Transcription factor Nfgi-a and environmentally induced Gr promoter 1 7 methylation have been implicated in fine-tuning the expression of Gr 1 7 transcripts. Here, we investigated Gr promoter 1 7 methylation and Gr 1 7 expression in adult rats exposed to either acute or chronic stress paradigms. A strong negative correlation was observed between the sum of promoter-wide methylation levels and Gr 1 7 transcript levels, independent of the stressor. Methylation of individual sites did not, however, correlate with transcript levels. This suggested that promoter 1 7 was directly regulated by promoter-wide DNA methylation. Although acute stress increased Ngfi-a expression in the hypothalamic paraventricular nucleus (PVN), Gr 1 7 transcript levels remained unaffected despite low methylation levels. Acute stress had little effect on these low methylation levels, except at four hippocampal CpGs. Chronic stress altered the corticosterone response to an acute stressor. In the adrenal and pituitary glands, but not in the brain, this was accompanied by an increase in methylation levels in orchestrated clusters rather than individual CpGs. PVN methylation levels, unaffected by acute or chronic stress, were significantly more variable within- than between-groups, suggesting that they were instated probably during the perinatal period and represent a pre-established trait. Thus, in addition to the known perinatal programming, the Gr 1 7 promoter is epigenetically regulated by chronic stress in adulthood, and retains promoter-wide tissue-specific plasticity. Differences in methylation susceptibility between the PVN in the perinatal period and the peripheral HPA axis tissues in adulthood may represent an important "trait" vs. "state" regulation of the Gr gene.

Structural Diversity in Conserved Regions Like the DRY-Motif among Viral 7TM Receptors-A Consequence of Evolutionary Pressure?

DEFF Research Database (Denmark)

Mølleskov-Jensen, Ann-Sofie; Sparre-Ulrich, Alexander Hovard; Davis-Poynter, Nicholas

2012-01-01

Several herpes- and poxviruses have captured chemokine receptors from their hosts and modified these to their own benefit. The human and viral chemokine receptors belong to class A 7 transmembrane (TM) receptors which are characterized by several structural motifs like the DRY-motif in TM3...... and the C-terminal tail. In the DRY-motif, the arginine residue serves important purposes by being directly involved in G protein coupling. Interestingly, among the viral receptors there is a greater diversity in the DRY-motif compared to their endogenous receptor homologous. The C-terminal receptor tail...... constitutes another regulatory region that through a number of phosphorylation sites is involved in signaling, desensitization, and internalization. Also this region is more variable among virus-encoded 7TM receptors compared to human class A receptors. In this review we will focus on these two structural...

A composite method based on formal grammar and DNA structural features in detecting human polymerase II promoter region.

Directory of Open Access Journals (Sweden)

Sutapa Datta

Full Text Available An important step in understanding gene regulation is to identify the promoter regions where the transcription factor binding takes place. Predicting a promoter region de novo has been a theoretical goal for many researchers for a long time. There exists a number of in silico methods to predict the promoter region de novo but most of these methods are still suffering from various shortcomings, a major one being the selection of appropriate features of promoter region distinguishing them from non-promoters. In this communication, we have proposed a new composite method that predicts promoter sequences based on the interrelationship between structural profiles of DNA and primary sequence elements of the promoter regions. We have shown that a Context Free Grammar (CFG can formalize the relationships between different primary sequence features and by utilizing the CFG, we demonstrate that an efficient parser can be constructed for extracting these relationships from DNA sequences to distinguish the true promoter sequences from non-promoter sequences. Along with CFG, we have extracted the structural features of the promoter region to improve upon the efficiency of our prediction system. Extensive experiments performed on different datasets reveals that our method is effective in predicting promoter sequences on a genome-wide scale and performs satisfactorily as compared to other promoter prediction techniques.

A Composite Method Based on Formal Grammar and DNA Structural Features in Detecting Human Polymerase II Promoter Region

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Datta, Sutapa; Mukhopadhyay, Subhasis

2013-01-01

An important step in understanding gene regulation is to identify the promoter regions where the transcription factor binding takes place. Predicting a promoter region de novo has been a theoretical goal for many researchers for a long time. There exists a number of in silico methods to predict the promoter region de novo but most of these methods are still suffering from various shortcomings, a major one being the selection of appropriate features of promoter region distinguishing them from non-promoters. In this communication, we have proposed a new composite method that predicts promoter sequences based on the interrelationship between structural profiles of DNA and primary sequence elements of the promoter regions. We have shown that a Context Free Grammar (CFG) can formalize the relationships between different primary sequence features and by utilizing the CFG, we demonstrate that an efficient parser can be constructed for extracting these relationships from DNA sequences to distinguish the true promoter sequences from non-promoter sequences. Along with CFG, we have extracted the structural features of the promoter region to improve upon the efficiency of our prediction system. Extensive experiments performed on different datasets reveals that our method is effective in predicting promoter sequences on a genome-wide scale and performs satisfactorily as compared to other promoter prediction techniques. PMID:23437045

Aryl hydrocarbon receptor downregulates MYCN expression and promotes cell differentiation of neuroblastoma.

Directory of Open Access Journals (Sweden)

Pei-Yi Wu

Full Text Available Neuroblastoma (NB is the most common malignant disease of infancy. MYCN amplification is a prognostic factor for NB and is a sign of highly malignant disease and poor patient prognosis. In this study, we aimed to investigate novel MYCN-related genes and assess how they affect NB cell behavior. The different gene expression found in 10 MYCN amplification NB tumors and 10 tumors with normal MYCN copy number were analyzed using tissue oligonucleotide microarrays. Ingenuity Pathway Analysis was subsequently performed to identify the potential genes involved in MYCN regulation pathways. Aryl hydrocarbon receptor (AHR, a receptor for dioxin-like compounds, was found to be inversely correlated with MYCN expression in NB tissues. This correlation was confirmed in a further 14 human NB samples. Moreover, AHR expression in NB tumors was found to correlate highly with histological grade of differentiation. In vitro studies revealed that AHR overexpression in NB cells induced spontaneous cell differentiation. In addition, it was found that ectopic expression of AHR suppressed MYCN promoter activity resulting in downregulation of MYCN expression. The suppression effect of AHR on the transcription of MYCN was compensated for by E2F1 overexpression, indicating that E2F1 is involved in the AHR-regulating MYCN pathway. Furthermore, AHR shRNA promotes the expression of E2F1 and MYCN in NB cells. These findings suggest that AHR is one of the upstream regulators of MYCN. Through the modulation of E2F1, AHR regulates MYCN gene expression, which may in turn affect NB differentiation.

COLOCALIZATION OF MUSCARINIC AND NICOTINIC RECEPTORS IN CHOLINOCEPTIVE NEURONS OF THE SUPRACHIASMATIC REGION IN YOUNG AND AGED RATS

NARCIS (Netherlands)

VANDERZEE, EA; STREEFLAND, C; STROSBERG, AD; SCHRODER, H; LUITEN, PGM; Schröder, H.

1991-01-01

In the present study muscarinic and nicotinic cholinergic receptors in the SCN region were demonstrated and analyzed, employing monoclonal antibodies to purified muscarinic and nicotinic cholinergic receptor proteins. A near-total colocalization of the two acetylcholine receptor subclasses in

Class A scavenger receptor promotes osteoclast differentiation via the enhanced expression of receptor activator of NF-{kappa}B (RANK)

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Takemura, Kenichi [Department of Cell Pathology, Graduate School of Medical Sciences, Kumamoto University, 1-1-1 Honjo, Kumamoto 860-8556 (Japan); Department of Orthopaedic and Neuro-Musculoskeletal Surgery, Graduate School of Medical Sciences, Kumamoto University, Kumamoto (Japan); Sakashita, Naomi; Fujiwara, Yukio; Komohara, Yoshihiro; Lei, XiaoFeng; Ohnishi, Koji [Department of Cell Pathology, Graduate School of Medical Sciences, Kumamoto University, 1-1-1 Honjo, Kumamoto 860-8556 (Japan); Suzuki, Hiroshi [National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Hokkaido (Japan); Kodama, Tatsuhiko [Department of Molecular Biology and Medicine, Research Center for Advanced Science and Technology, The University of Tokyo, Tokyo (Japan); Mizuta, Hiroshi [Department of Orthopaedic and Neuro-Musculoskeletal Surgery, Graduate School of Medical Sciences, Kumamoto University, Kumamoto (Japan); Takeya, Motohiro, E-mail: takeya@kumamoto-u.ac.jp [Department of Cell Pathology, Graduate School of Medical Sciences, Kumamoto University, 1-1-1 Honjo, Kumamoto 860-8556 (Japan)

2010-01-22

Osteoclasts originate from bone marrow monocyte/macrophage lineage cells, and their differentiation depends on macrophage colony-stimulating factor (M-CSF) and receptor activator nuclear factor kappa B (RANK) ligand. Class A scavenger receptor (SR-A) is one of the principal functional molecules of macrophages, and its level of expression declines during osteoclast differentiation. To investigate the role of SR-A in osteoclastogenesis, we examined pathological changes in femoral bone and the expression levels of osteoclastogenesis-related molecules in SR-A{sup -/-} mice. The femoral osseous density of SR-A{sup -/-} mice was higher than that of SR-A{sup +/+} mice, and the number of multinucleated osteoclasts was significantly decreased. An in vitro differentiation assay revealed that the differentiation of multinucleated osteoclasts from bone marrow-derived progenitor cells is impaired in SR-A{sup -/-} mice. Elimination of SR-A did not alter the expression level of the M-CSF receptor, c-fms; however, the expression levels of RANK and RANK-related osteoclast-differentiation molecules such as nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 1 (NFATc1) and microphthalmia-associated transcription factor (MITF) significantly decreased. Furthermore, acetylated low-density lipoprotein (AcLDL), an SR-A ligand, significantly increased the expression level of RANK and MITF during osteoclast differentiation. These data indicate that SR-A promotes osteoclastogenesis via augmentation of the expression level of RANK and its related molecules.

Promotion of Wound Healing by an Agonist of Adenosine A2A Receptor Is Dependent on Tissue Plasminogen Activator.

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Montesinos, M Carmen; Desai-Merchant, Avani; Cronstein, Bruce N

2015-12-01

Impaired wound healing, as it occurs in diabetes mellitus or long-term corticoid treatment, is commonly associated with disability, diminished quality of life, and high economic costs. Selective agonists of the A2A receptor subtype of adenosine, an endogenous regulator of inflammation, promote tissue repair in animal models, both healthy and with impaired healing. Plasmin-mediated proteolysis of fibrin and other matrix proteins is essential for cell migration at sites of injury. Since adenosine A2A receptor activation increases plasminogen activator release from macrophages and mast cells, we studied the effect of a selective agonist, CGS-21680, on full-thickness excisional wound closure in wild-type, urokinase plasminogen activator (uPA)-deficient, and tissue plasminogen activator (tPA)-deficient mice. Wound closure was impaired in tPA- and uPA-deficient mice as compared with wild-type mice, and topical application of CGS-21680 significantly increased the rate at which wounds closed in wild-type mice and uPA-deficient mice, but not in tPA-deficient mice. Immunostaining of tissue sections showed that tPA was present in endothelial cells and histiocytes by day 3 post-wound and also by day 6. In contrast, uPA was more prominent in these cell types only by day 6 post-wound. Our results confirm that plasminogen activation contributes to wound repair and are consistent with the hypothesis that adenosine A2A receptor activation promotes wound closure by a mechanism that depends upon tPA, but not uPA. Moreover, our results suggest that topical adenosine A2A receptor agonists may be useful in promotion of wound closure in patients with impaired wound healing.

Disruption of histone modification and CARM1 recruitment by arsenic represses transcription at glucocorticoid receptor-regulated promoters.

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Barr, Fiona D; Krohmer, Lori J; Hamilton, Joshua W; Sheldon, Lynn A

2009-08-26

Chronic exposure to inorganic arsenic (iAs) found in the environment is one of the most significant and widespread environmental health risks in the U.S. and throughout the world. It is associated with a broad range of health effects from cancer to diabetes as well as reproductive and developmental anomalies. This diversity of diseases can also result from disruption of metabolic and other cellular processes regulated by steroid hormone receptors via aberrant transcriptional regulation. Significantly, exposure to iAs inhibits steroid hormone-mediated gene activation. iAs exposure is associated with disease, but is also used therapeutically to treat specific cancers complicating an understanding of iAs action. Transcriptional activation by steroid hormone receptors is accompanied by changes in histone and non-histone protein post-translational modification (PTM) that result from the enzymatic activity of coactivator and corepressor proteins such as GRIP1 and CARM1. This study addresses how iAs represses steroid receptor-regulated gene transcription. PTMs on histones H3 and H4 at the glucocorticoid receptor (GR)-activated mouse mammary tumor virus (MMTV) promoter were identified by chromatin immunoprecipitation analysis following exposure to steroid hormone+/-iAs. Histone H3K18 and H3R17 amino acid residues had significantly different patterns of PTMs after treatment with iAs. Promoter interaction of the coactivator CARM1 was disrupted, but the interaction of GRIP1, a p160 coactivator through which CARM1 interacts with a promoter, was intact. Over-expression of CARM1 was able to fully restore and GRIP1 partially restored iAs-repressed transcription indicating that these coactivators are functionally associated with iAs-mediated transcriptional repression. Both are essential for robust transcription at steroid hormone regulated genes and both are associated with disease when inappropriately expressed. We postulate that iAs effects on CARM1 and GRIP1 may underlie some

Reversible and regionally selective downregulation of brain cannabinoid CB1 receptors in chronic daily cannabis smokers

OpenAIRE

Hirvonen, J; Goodwin, RS; Li, C-T; Terry, GE; Zoghbi, SS; Morse, C; Pike, VW; Volkow, ND; Huestis, MA; Innis, RB

2011-01-01

Chronic cannabis (marijuana, hashish) smoking can result in dependence. Rodent studies show reversible downregulation of brain cannabinoid CB1 (cannabinoid receptor type 1) receptors after chronic exposure to cannabis. However, whether downregulation occurs in humans who chronically smoke cannabis is unknown. Here we show, using positron emission tomography imaging, reversible and regionally selective downregulation of brain cannabinoid CB1 receptors in human subjects who chronically smoke ca...

The N-terminal region of the dopamine D2 receptor, a rhodopsin-like GPCR, regulates correct integration into the plasma membrane and endocytic routes

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Cho, DI; Min, C; Jung, KS; Cheong, SY; Zheng, M; Cheong, SJ; Oak, MH; Cheong, JH; Lee, BK; Kim, KM

2012-01-01

BACKGROUND AND PURPOSE Functional roles of the N-terminal region of rhodopsin-like GPCR family remain unclear. Using dopamine D2 and D3 receptors as a model system, we probed the roles of the N-terminal region in the signalling, intracellular trafficking of receptor proteins, and explored the critical factors that determine the functionality of the N-terminal region. EXPERIMENTAL APPROACH The N-terminal region of the D2 receptor was gradually shortened or switched with that of the D3 receptor or a non-specific sequence (FLAG), or potential N-terminal glycosylation sites were mutated. Effects of these manipulations on surface expression, internalization, post-endocytic behaviours and signalling were determined. KEY RESULTS Shortening the N-terminal region of the D2 receptor enhanced receptor internalization and impaired surface expression and signalling; ligand binding, desensitization and down-regulation were not affected but their association with a particular microdomain, caveolae, was disrupted. Replacement of critical residues within the N-terminal region with the FLAG epitope failed to restore surface expression but partially restored the altered internalization and signalling. When the N-terminal regions were switched between D2 and D3 receptors, cell surface expression pattern of each receptor was switched. Mutations of potential N-terminal glycosylation sites inhibited surface expression but enhanced internalization of D2 receptors. CONCLUSIONS AND IMPLICATIONS Shortening of N-terminus or mutation of glycosylation sites located within the N-terminus enhanced receptor internalization but impaired the surface expression of D2 receptors. The N-terminal region of the D2 receptor, in a sequence-specific manner, controls the receptor's conformation and integration into the plasma membrane, which determine its subcellular localization, intracellular trafficking and signalling properties. PMID:22117524

Selection for Unequal Densities of Sigma70 Promoter-like Signalsin Different Regions of Large Bacterial Genomes

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Huerta, Araceli M.; Francino, M. Pilar; Morett, Enrique; Collado-Vides, Julio

2006-03-01

The evolutionary processes operating in the DNA regions that participate in the regulation of gene expression are poorly understood. In Escherichia coli, we have established a sequence pattern that distinguishes regulatory from nonregulatory regions. The density of promoter-like sequences, that are recognizable by RNA polymerase and may function as potential promoters, is high within regulatory regions, in contrast to coding regions and regions located between convergently-transcribed genes. Moreover, functional promoter sites identified experimentally are often found in the subregions of highest density of promoter-like signals, even when individual sites with higher binding affinity for RNA polymerase exist elsewhere within the regulatory region. In order to investigate the generality of this pattern, we have used position weight matrices describing the -35 and -10 promoter boxes of E. coli to search for these motifs in 43 additional genomes belonging to most established bacterial phyla, after specific calibration of the matrices according to the base composition of the noncoding regions of each genome. We have found that all bacterial species analyzed contain similar promoter-like motifs, and that, in most cases, these motifs follow the same genomic distribution observed in E. coli. Differential densities between regulatory and nonregulatory regions are detectable in most bacterial genomes, with the exception of those that have experienced evolutionary extreme genome reduction. Thus, the phylogenetic distribution of this pattern mirrors that of genes and other genomic features that require weak selection to be effective in order to persist. On this basis, we suggest that the loss of differential densities in the reduced genomes of host-restricted pathogens and symbionts is the outcome of a process of genome degradation resulting from the decreased efficiency of purifying selection in highly structured small populations. This implies that the differential

Wake-promoting effects of ONO-4127Na, a prostaglandin DP1 receptor antagonist, in hypocretin/orexin deficient narcoleptic mice.

Science.gov (United States)

Sagawa, Yohei; Sato, Masatoshi; Sakai, Noriaki; Chikahisa, Sachiko; Chiba, Shintaro; Maruyama, Takashi; Yamamoto, Junki; Nishino, Seiji

2016-11-01

Prostaglandin (PG)D2 is an endogenous sleep substance, and a series of animal studies reported that PGD2 or PGD2 receptor (DP1) agonists promote sleep, while DP1 antagonists promote wakefulness. This suggests the possibility of use of PG DP1 antagonists as wake-promoting compounds. We therefore evaluated the wake-promoting effects of ONO-4127Na, a DP1 antagonist, in a mouse model of narcolepsy (i.e., orexin/ataxin-3 transgenic mice) and compared those to effects of modafinil. ONO-4127Na perfused in the basal forebrain (BF) area potently promoted wakefulness in both wild type and narcoleptic mice, and the wake-promoting effects of ONO-4127Na at 2.93 × 10(-4) M roughly corresponded to those of modafinil at 100 mg/kg (p.o.). The wake promoting effects of ONO-4127Na was observed both during light and dark periods, and much larger effects were seen during the light period when mice slept most of the time. ONO-4127Na, when perfused in the hypothalamic area, had no effects on sleep. We further demonstrated that wake-promoting effects of ONO-4127Na were abolished in DP1 KO mice, confirming that the wake-promoting effect of ONO-4127Na is mediated by blockade of the PG DP1 receptors located in the BF area. ONO-4127Na reduced DREM, an EEG/EMG assessment of behavioral cataplexy in narcoleptic mice, suggesting that ONO-4127Na is likely to have anticataplectic effects. DP1 antagonists may be a new class of compounds for the treatment of narcolepsy-cataplexy, and further studies are warranted. Copyright © 2016 Elsevier Ltd. All rights reserved.

Mechanosensitive promoter region in the human HB-GAM gene

DEFF Research Database (Denmark)

Liedert, Astrid; Kassem, Moustapha; Claes, Lutz

2009-01-01

Mechanical loading is essential for maintaining bone mass in the adult skeleton. However, the underlying process of the transfer of the physical stimulus into a biochemical response, which is termed mechanotransduction is poorly understood. Mechanotransduction results in the modulation of gene...... cells. Analysis of the human HB-GAM gene upstream regulatory region with luciferase reporter gene assays revealed that the upregulation of HB-GAM expression occurred at the transcriptional level and was mainly dependent on the HB-GAM promoter region most upstream containing three potential AP-1 binding...

A basic peptide within the juxtamembrane region is required for EGF receptor dimerization.

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Aifa, Sami; Aydin, Jan; Nordvall, Gunnar; Lundström, Ingemar; Svensson, Samuel P S; Hermanson, Ola

2005-01-01

The epidermal growth factor receptor (EGFR) is fundamental for normal cell growth and organ development, but has also been implicated in various pathologies, notably tumors of epithelial origin. We have previously shown that the initial 13 amino acids (P13) within the intracellular juxtamembrane region (R645-R657) are involved in the interaction with calmodulin, thus indicating an important role for this region in EGFR function. Here we show that P13 is required for proper dimerization of the receptor. We expressed either the intracellular domain of EGFR (TKJM) or the intracellular domain lacking P13 (DeltaTKJM) in COS-7 cells that express endogenous EGFR. Only TKJM was immunoprecipitated with an antibody directed against the extracellular part of EGFR, and only TKJM was tyrosine phosphorylated by endogenous EGFR. Using SK-N-MC cells, which do not express endogenous EGFR, that were stably transfected with either wild-type EGFR or recombinant full-length EGFR lacking P13 demonstrated that P13 is required for appropriate receptor dimerization. Furthermore, mutant EGFR lacking P13 failed to be autophosphorylated. P13 is rich in basic amino acids and in silico modeling of the EGFR in conjunction with our results suggests a novel role for the juxtamembrane domain (JM) of EGFR in mediating intracellular dimerization and thus receptor kinase activation and function.

Molecular characterization of the haptoglobin.hemoglobin receptor CD163. Ligand binding properties of the scavenger receptor cysteine-rich domain region

DEFF Research Database (Denmark)

Madsen, Mette; Møller, Holger J; Nielsen, Marianne Jensby

2004-01-01

CD163 is the macrophage receptor for endocytosis of haptoglobin.hemoglobin complexes. The extracellular region consisting of nine scavenger receptor cysteine rich (SRCR) domains also circulates in plasma as a soluble protein. By ligand binding analysis of a broad spectrum of soluble CD163...... truncation variants, the amino-terminal third of the SRCR region was shown to be crucial for the binding of haptoglobin.hemoglobin complexes. By Western blotting of the CD163 variants, a panel of ten monoclonal antibodies was mapped to SRCR domains 1, 3, 4, 6, 7, and 9, respectively. Only the two antibodies...... to CD163 demonstrated that optimal ligand binding requires physiological plasma calcium concentrations, and an immediate ligand release occurs at the low calcium concentrations measured in acidifying endosomes. In conclusion, SRCR domain 3 of CD163 is an exposed domain and a critical determinant...

Disruption of retinoic acid receptor alpha reveals the growth promoter face of retinoic acid.

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Giulia Somenzi

2007-09-01

Full Text Available Retinoic acid (RA, the bioactive derivative of Vitamin A, by epigenetically controlling transcription through the RA-receptors (RARs, exerts a potent antiproliferative effect on human cells. However, a number of studies show that RA can also promote cell survival and growth. In the course of one of our studies we observed that disruption of RA-receptor alpha, RARalpha, abrogates the RA-mediated growth-inhibitory effects and unmasks the growth-promoting face of RA (Ren et al., Mol. Cell. Biol., 2005, 25:10591. The objective of this study was to investigate whether RA can differentially govern cell growth, in the presence and absence of RARalpha, through differential regulation of the "rheostat" comprising ceramide (CER, the sphingolipid with growth-inhibitory activity, and sphingosine-1-phosphate (S1P, the sphingolipid with prosurvival activity.We found that functional inhibition of endogenous RARalpha in breast cancer cells by using either RARalpha specific antagonists or a dominant negative RARalpha mutant hampers on one hand the RA-induced upregulation of neutral sphingomyelinase (nSMase-mediated CER synthesis, and on the other hand the RA-induced downregulation of sphingosine kinase 1, SK1, pivotal for S1P synthesis. In association with RA inability to regulate the sphingolipid rheostat, cells not only survive, but also grow more in response to RA both in vitro and in vivo. By combining genetic, pharmacological and biochemical approaches, we mechanistically demonstrated that RA-induced growth is, at least in part, due to non-RAR-mediated activation of the SK1-S1P signaling.In the presence of functional RARalpha, RA inhibits cell growth by concertedly, and inversely, modulating the CER and S1P synthetic pathways. In the absence of a functional RARalpha, RA-in a non-RAR-mediated fashion-promotes cell growth by activating the prosurvival S1P signaling. These two distinct, yet integrated processes apparently concur to the growth-promoter effects

Signaling cross-talk between peroxisome proliferator-activated receptor/retinoid X receptor and estrogen receptor through estrogen response elements.

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Keller, H; Givel, F; Perroud, M; Wahli, W

1995-07-01

Peroxisome proliferator-activated receptors (PPARs) and retinoid X receptors (RXRs) are nuclear hormone receptors that are activated by fatty acids and 9-cis-retinoic acid, respectively. PPARs and RXRs form heterodimers that activate transcription by binding to PPAR response elements (PPREs) in the promoter of target genes. The PPREs described thus far consist of a direct tandem repeat of the AGGTCA core element with one intervening nucleotide. We show here that the vitellogenin A2 estrogen response element (ERE) can also function as a PPRE and is bound by a PPAR/RXR heterodimer. Although this heterodimer can bind to several other ERE-related palindromic response elements containing AGGTCA half-sites, only the ERE is able to confer transactivation of test reporter plasmids, when the ERE is placed either close to or at a distance from the transcription initiation site. Examination of natural ERE-containing promoters, including the pS2, very-low-density apolipoprotein II and vitellogenin A2 genes, revealed considerable differences in the binding of PPAR/RXR heterodimers to these EREs. In their natural promoter context, these EREs did not allow transcriptional activation by PPARs/RXRs. Analysis of this lack of stimulation of the vitellogenin A2 promoter demonstrated that PPARs/RXRs bind to the ERE but cannot transactivate due to a nonpermissive promoter structure. As a consequence, PPARs/RXRs inhibit transactivation by the estrogen receptor through competition for ERE binding. This is the first example of signaling cross-talk between PPAR/RXR and estrogen receptor.

Identification of a single-nucleotide insertion in the promoter region affecting the sodC promoter activity in Brucella neotomae.

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Dina A Moustafa

Full Text Available Brucella neotomae is not known to be associated with clinical disease in any host species. Previous research suggested that B. neotomae might not express detectable levels of Cu/Zn superoxide dismutase (SOD, a periplasmic enzyme known to be involved in protecting Brucella from oxidative bactericidal effects of host phagocytes. This study was undertaken to investigate the genetic basis for the disparity in SOD expression in B. neotomae. Our Western blot and SOD enzyme assay analyses indicated that B. neotomae does express SOD, but at a substantially reduced level. Nucleotide sequence analysis of region upstream to the sodC gene identified a single-nucleotide insertion in the potential promoter region. The same single-nucleotide insertion was also detected in the sodC promoter of B. suis strain Thomsen, belonging to biovar 2 in which SOD expression was undetectable previously. Examination of the sodC promoter activities using translational fusion constructs with E. coli β-galactosidase demonstrated that the B. neotomae and B. suis biovar 2 promoters were very weak in driving gene expression. Site-directed mutation studies indicated that the insertion of A in the B. neotomae sodC promoter reduced the promoter activity. Increasing the level of SOD expression in B. neotomae through complementation with B. abortus sodC gene did not alter the bacterial survival in J774A.1 macrophage-like cells and in tissues of BALB/c and C57BL/6 mice. These results for the first time demonstrate the occurrence of a single-nucleotide polymorphism affecting promoter function and gene expression in Brucella.

Clinical significance of promoter region hypermethylation of microRNA-148a in gastrointestinal cancers

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Sun JX

2014-05-01

Full Text Available Jingxu Sun,1,* Yongxi Song,1,* Zhenning Wang,1 Guoli Wang,2 Peng Gao,1 Xiaowan Chen,1 Zhaohua Gao,1 Huimian Xu1 1Department of Surgical Oncology and General Surgery, First Hospital of China Medical University, Shenyang, People’s Republic of China; 2Department of Biochemistry and Molecular Biology, China Medical University, Shenyang, People’s Republic of China *These authors contributed equally to this work Background: MicroRNAs are associated with tumor genesis and progression in various carcinomas. MicroRNA-148a (miR-148a was reported to have low expression in gastrointestinal cancers, and might be regulated by promoter region DNA methylation. Methods: Bisulfite-modified sequencing was used to determine the promoter region DNA methylation status of human gastrointestinal cancer cell lines. Expression levels of miR-148a in cell lines treated with 5-aza-2′-deoxycytidine were determined by quantitative real-time polymerase chain reaction. Total DNA was extracted from the tissues of 64 patients with gastric cancer and 51 patients with colorectal cancer. Methylation status was determined by methylation-specific polymerase chain reaction. All statistical analyses were performed with SPSS 17.0 software. Results: The promoter regions of genes in human gastrointestinal cancer cell lines were all hypermethylated, except for HT-29, and the expression of miR-148a tended to be higher than in controls after treatment with 5-aza-2′-deoxycytidine. The methylation-specific polymerase chain reaction results showed that 56.25% of gastric cancer tissues and 19.61% of colorectal cancer tissues were hypermethylated. A strong correlation was found between the expression of miR-148a and the methylation status of promoter regions (P<0.001, chi-square test and Pearson’s correlation. Furthermore, promoter region CpG site hypermethylation of miR-148a was correlated with increased tumor size (P=0.01 in gastric cancer after analyzing the correlation between

G-protein-coupled receptor 81 promotes a malignant phenotype in breast cancer through angiogenic factor secretion.

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Lee, Yu Jin; Shin, Kyeong Jin; Park, Soo-Ah; Park, Kyeong Su; Park, Seorim; Heo, Kyun; Seo, Young-Kyo; Noh, Dong-Young; Ryu, Sung Ho; Suh, Pann-Ghill

2016-10-25

G-protein-coupled receptor 81 (GPR81) functions as a receptor for lactate and plays an important role in the regulation of anti-lipolytic effects in adipocytes. However, to data, a role for GPR81 in the tumor microenvironment has not been clearly defined. Here, GPR81 expression in breast cancer patients and several breast cancer cell lines was significantly increased compared with normal mammary tissues and cells. GPR81 knockdown resulted in impaired breast cancer growth and led to apoptosis both in vitro and in vivo. Furthermore, the inhibition of GPR81 signaling suppressed angiogenesis through a phosphoinositide 3-OH kinase (PI3K)/Akt-cAMP response element binding protein (CREB) pathway, which led to decreased production of the pro-angiogenic mediator amphiregulin (AREG). Overall, these findings identify GPR81 as a tumor-promoting receptor in breast cancer progression and suggest a novel mechanism that regulates GPR81-dependent activation of the PI3K/Akt signaling axis in tumor microenvironment.

Brand Products of Regional Cuisine in the Promotion of Tourism in Roztocze

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Bekier-Jaworska Ewa

2015-03-01

Full Text Available Introduction. There has been a trend over the last few years of using specialties of regional cuisine as an independent tourist attraction. The creation of local brands is an important element in the promotion of a given region and it also influences the development of culinary tourism. The aim of the studies conducted was to identify regional dishes - a choice of dishes that could be described as 'brand dishes' and the use of those dishes as tourist attractions in Roztocze. Material and methods. Studies were conducted on a group of students studying tourism and recreation at State Higher School of Vocational Education (PWSZ in Zamość using a questionnaire. Results. The questionnaire provided an assessment of the levels of knowledge of regional cuisine among Polish and Ukrainian students, identified the most characteristic dishes and selected brand products, and helped to arrive at a suitable method of promotion. Conclusions. Nationality, family customs and selection of local restaurants highly influence knowledge of regional cuisine. Interviewees decided that the most outstanding products from Roztocze were Zwierzyniec beer, and Biłgoraj pie. Regional products should be used as a tourist attraction in Roztocze.

The Pentapeptide RM-131 Promotes Food Intake and Adiposity in Wildtype Mice but Not in Mice Lacking the Ghrelin Receptor

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Fischer, Katrin; Finan, Brian; Clemmensen, Christoffer

2014-01-01

The gastrointestinal peptide hormone ghrelin is the endogenous ligand of the growth hormone secretagogue receptor (a.k.a. ghrelin receptor, GHR). Currently, ghrelin is the only circulating peripheral hormone with the ability to promote a positive energy balance by stimulating food intake while...... decreasing energy expenditure and body fat utilization, as defined in rodents. Based on these and additional, beneficial effects on metabolism, the endogenous ghrelin system is considered an attractive target to treat diverse pathological conditions including those associated with eating/wasting disorders...... and cachexia. As the pharmacological potential of ghrelin is hampered by its relatively short half-life, ghrelin analogs with enhanced pharmacokinetics offer the potential to sustainably improve metabolism. One of these ghrelin analogs is the pentapeptide RM-131, which promotes food intake and adiposity...

Leptin Receptor Deficiency is Associated With Upregulation of Cannabinoid 1 Receptors in Limbic Brain Regions

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THANOS, PANAYOTIS K.; RAMALHETE, ROBERTO C.; MICHAELIDES, MICHAEL; PIYIS, YIANNI K.; WANG, GENE-JACK; VOLKOW, NORA D.

2009-01-01

Leptin receptor dysfunction results in overeating and obesity. Leptin regulates hypothalamic signaling that underlies the motivation to hyperphagia, but the interaction between leptin and cannabinoid signaling is poorly understood. We evaluated the role of cannabinoid 1 receptors (CB1R) in overeating and the effects of food deprivation on CB1R in the brain. One-month-old Zucker rats were divided into unrestricted and restricted (fed 70% of unrestricted rats) diet groups and maintained until adulthood (4 months). Levels of relative binding sites of CB1R (CB1R binding levels) were assessed using [3H] SR141716A in vitro autoradiography. These levels were higher (except cerebellum and hypothalamus) at 4 months than at 1 month of age. One month CB1R binding levels for most brain regions did not differ between Ob and Lean (Le) rats (except in frontal and cingulate cortices in Le and in the hypothalamus in Ob). Four month Ob rats had higher CB1R binding levels than Le in most brain regions and food restriction was associated with higher CB1R levels in all brain regions in Ob, but not in Le rats. CB1R binding levels increased between adolescence and young adulthood which we believe was influenced by leptin and food availability. The high levels of CB1R in Ob rats suggest that leptin's inhibition of food-intake is in part mediated by downregulation of CB1R and that leptin interferes with CB1R upregulation under food-deprivation conditions. These results are consistent with prior findings showing increased levels of endogenous cannabinoids in the Ob rats corroborating the regulation of cannabinoid signaling by leptin. PMID:18563836

Promoter-region hypermethylation and expression downregulation of Yy1 (Yin yang 1) in preneoplastic liver lesions in a thioacetamide rat hepatocarcinogenesis model

International Nuclear Information System (INIS)

Abe, Hajime; Ogawa, Takashi; Wang, Liyun; Kimura, Masayuki; Tanaka, Takeshi; Morita, Reiko; Yoshida, Toshinori; Shibutani, Makoto

2014-01-01

Thioacetamide (TAA) has been used to develop a rodent model for hepatocarcinogenesis. To determine the genes with epigenetic modifications in early hepatocarcinogenesis, we did a genome-wide scan for hypermethylated promoter regions using CpG island microarrays in TAA-promoted rat liver tissue. Eight genes were selected based on the microarray profile; of these, Yy1 and Wdr45b were confirmed to be hypermethylated by methylation-specific polymerase chain reaction (PCR) and pyrosequencing and downregulated by real-time reverse transcription PCR. Non-neoplastic liver cells had nuclear Yy1 immunoreactivity, while preneoplastic foci with glutathione S-transferase placental form (GST-P) immunoreactivity had decreased Yy1 immunoreactivity. The incidence of these foci was proportional to the dose of TAA administered. Co-expression analysis of gene products downstream of Yy1 revealed increased nuclear phospho-c-Myc + foci as well as nuclear and cytoplasmic p21 Cip1+ foci in Yy1 − or GST-P + foci in response to TAA-promotion dose. Although the absolute number of cells was low, the incidence of death receptor 5 − foci was increased in Yy1 − foci in proportion to the TAA dose. Yy1 − /GST-P + foci revealed a higher number of proliferating cell nuclear antigen (PCNA)-immunoreactive cells than Yy1 + /GST-P + foci, while cleaved caspase-3 + cells were unchanged between Yy1 – /GST-P + and Yy1 + /GST-P + foci. In the case of Wdr45b, most GST-P + foci were Wdr45b – and were not increased by TAA promotion. These results suggest involvement of Yy1 in the epigenetic gene regulation at the early stages of TAA promoted cell proliferation and concomitant cell cycle arrest in preneoplastic lesions. - Highlights: • Epigenetically downregulated genes were searched in TAA-promnoted rat livers. • Yy1 and Wdr45b showed promoter-region hypermethylation and mRNA downregulation. • TAA promoted increase of preneoplastic Yy1 – /GST-P + foci showing high proliferation. • TAA

The glycocalyx promotes cooperative binding and clustering of adhesion receptors.

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Xu, Guang-Kui; Qian, Jin; Hu, Jinglei

2016-05-18

Cell adhesion plays a pivotal role in various biological processes, e.g., immune responses, cancer metastasis, and stem cell differentiation. The adhesion behaviors depend subtly on the binding kinetics of receptors and ligands restricted at the cell-substrate interfaces. Although much effort has been directed toward investigating the kinetics of adhesion molecules, the role of the glycocalyx, anchored on cell surfaces as an exterior layer, is still unclear. In this paper, we propose a theoretical approach to study the collective binding kinetics of a few and a large number of binders in the presence of the glycocalyx, representing the cases of initial and mature adhesions of cells, respectively. The analytical results are validated by finding good agreement with our Monte Carlo simulations. In the force loading case, the on-rate and affinity increase as more bonds form, whereas this cooperative effect is not observed in the displacement loading case. The increased thickness and stiffness of the glycocalyx tend to decrease the affinity for a few bonds, while they have less influence on the affinity for a large number of bonds. Moreover, for a flexible membrane with thermally-excited shape fluctuations, the glycocalyx is exhibited to promote the formation of bond clusters, mainly due to the cooperative binding of binders. This study helps to understand the cooperative kinetics of adhesion receptors under physiologically relevant loading conditions and sheds light on the novel role of the glycocalyx in cell adhesion.

Combgap Promotes Ovarian Niche Development and Chromatin Association of EcR-Binding Regions in BR-C.

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Hitrik, Anna; Popliker, Malka; Gancz, Dana; Mukamel, Zohar; Lifshitz, Aviezer; Schwartzman, Omer; Tanay, Amos; Gilboa, Lilach

2016-11-01

The development of niches for tissue-specific stem cells is an important aspect of stem cell biology. Determination of niche size and niche numbers during organogenesis involves precise control of gene expression. How this is achieved in the context of a complex chromatin landscape is largely unknown. Here we show that the nuclear protein Combgap (Cg) supports correct ovarian niche formation in Drosophila by controlling ecdysone-Receptor (EcR)- mediated transcription and long-range chromatin contacts in the broad locus (BR-C). Both cg and BR-C promote ovarian growth and the development of niches for germ line stem cells. BR-C levels were lower when Combgap was either reduced or over-expressed, indicating an intricate regulation of the BR-C locus by Combgap. Polytene chromosome stains showed that Cg co-localizes with EcR, the major regulator of BR-C, at the BR-C locus and that EcR binding to chromatin was sensitive to changes in Cg levels. Proximity ligation assay indicated that the two proteins could reside in the same complex. Finally, chromatin conformation analysis revealed that EcR-bound regions within BR-C, which span ~30 KBs, contacted each other. Significantly, these contacts were stabilized in an ecdysone- and Combgap-dependent manner. Together, these results highlight Combgap as a novel regulator of chromatin structure that promotes transcription of ecdysone target genes and ovarian niche formation.

Identification of functional DNA variants in the constitutive promoter region of MDM2

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Lalonde Marie-Eve

2012-09-01

Full Text Available Abstract Although mutations in the oncoprotein murine double minute 2 (MDM2 are rare, MDM2 gene overexpression has been observed in several human tumors. Given that even modest changes in MDM2 levels might influence the p53 tumor suppressor signaling pathway, we postulated that sequence variation in the promoter region of MDM2 could lead to disregulated expression and variation in gene dosage. Two promoters have been reported for MDM2; an internal promoter (P2, which is located near the end of intron 1 and is p53-responsive, and an upstream constitutive promoter (P1, which is p53-independent. Both promoter regions contain DNA variants that could influence the expression levels of MDM2, including the well-studied single nucleotide polymorphism (SNP SNP309, which is located in the promoter P2; i.e., upstream of exon 2. In this report, we screened the promoter P1 for DNA variants and assessed the functional impact of the corresponding SNPs. Using the dbSNP database and genotyping validation in individuals of European descent, we identified three common SNPs (−1494 G > A; indel 40 bp; and −182 C > G. Three major promoter haplotypes were inferred by using these three promoter SNPs together with rs2279744 (SNP309. Following subcloning into a gene reporter system, we found that two of the haplotypes significantly influenced MDM2 promoter activity in a haplotype-specific manner. Site-directed mutagenesis experiments indicated that the 40 bp insertion/deletion variation is causing the observed allelic promoter activity. This study suggests that part of the variability in the MDM2 expression levels could be explained by allelic p53-independent P1 promoter activity.

Histone H4 Lys 20 methyltransferase SET8 promotes androgen receptor-mediated transcription activation in prostate cancer

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Yao, Lushuai [Laboratory of Genome Variations and Precision Bio-Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Li, Yanyan; Du, Fengxia [Laboratory of Genome Variations and Precision Bio-Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101 (China); Han, Xiao [Laboratory of Genome Variations and Precision Bio-Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Li, Xiaohua [Laboratory of Genome Variations and Precision Bio-Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101 (China); Niu, Yuanjie [Chawnshang Chang Sex Hormone Research Center, Tianjin Institute of Urology, Tianjin Medical University, Tianjin 300070 (China); Ren, Shancheng, E-mail: renshancheng@gmail.com [Department of Urology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai 200433 (China); Sun, Yingli, E-mail: sunyl@big.ac.cn [Laboratory of Genome Variations and Precision Bio-Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101 (China)

2014-07-18

Highlights: • Dihydrotestosterone stimulates H4K20me1 enrichment at the PSA promoter. • SET8 promotes AR-mediated transcription activation. • SET8 interacts with AR and promotes cell proliferation. - Abstract: Histone methylation status in different lysine residues has an important role in transcription regulation. The effect of H4K20 monomethylation (H4K20me1) on androgen receptor (AR)-mediated gene transcription remains unclear. Here we show that AR agonist stimulates the enrichment of H4K20me1 and SET8 at the promoter of AR target gene PSA in an AR dependent manner. Furthermore, SET8 is crucial for the transcription activation of PSA. Co-immunoprecipitation analyses demonstrate that SET8 interacts with AR. Therefore, we conclude that SET8 is involved in AR-mediated transcription activation, possibly through its interaction with AR and H4K20me1 modification.

Wake-promoting actions of median nerve stimulation in TBI-induced coma: An investigation of orexin-A and orexin receptor 1 in the hypothalamic region.

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Zhong, Ying-Jun; Feng, Zhen; Wang, Liang; Wei, Tian-Qi

2015-09-01

A coma is a serious complication, which can occur following traumatic brain injury (TBI), for which no effective treatment has been established. Previous studies have suggested that neural electrical stimulation, including median nerve stimulation (MNS), may be an effective method for treating patients in a coma, and orexin‑A, an excitatory hypothalamic neuropeptide, may be involved in wakefulness. However, the exact mechanisms underlying this involvement remain to be elucidated. The present study aimed to examine the arousal‑promoting role of MNS in rats in a TBI‑induced coma and to investigate the potential mechanisms involved. A total of 90 rats were divided into three groups, comprising a control group, sham‑stimulated (TBI) group and a stimulated (TBI + MNS) group. MNS was performed on the animals, which were in a TBI‑induced comatose state. Changes in the behavior of the rats were observed following MNS. Subsequently, hypothalamic tissues were extracted from the rats 6, 12 and 24 h following TBI or MNS, respectively. The expression levels of orexin‑A and orexin receptor‑1 (OX1R) in the hypothalamus were examined using immunohistochemistry, western blotting and an enzyme‑linked immunosorbent assay. The results demonstrated that 21 rats subjected to TBI‑induced coma exhibited a restored righting reflex and response to pain stimuli following MNS. In addition, ignificant differences in the expression levels of orexin‑A and OXIR were observed among the three groups and among the time‑points. Orexin‑A and OX1R were upregulated following MNS. The rats in the stimulated group reacted to the MNS and exhibited a re‑awakening response. The results of the present study indicated that MNS may be a therapeutic option for TBI‑induced coma. The mechanism may be associated with increasing expression levels of the excitatory hypothalamic neuropeptide, orexin-A, and its receptor, OX1R, in the hypothalamic region.

CD81 Receptor Regions outside the Large Extracellular Loop Determine Hepatitis C Virus Entry into Hepatoma Cells

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Pia Banse

2018-04-01

Full Text Available Hepatitis C virus (HCV enters human hepatocytes using four essential entry factors, one of which is human CD81 (hCD81. The tetraspanin hCD81 contains a large extracellular loop (LEL, which interacts with the E2 glycoprotein of HCV. The role of the non-LEL regions of hCD81 (intracellular tails, four transmembrane domains, small extracellular loop and intracellular loop is poorly understood. Here, we studied the contribution of these domains to HCV susceptibility of hepatoma cells by generating chimeras of related tetraspanins with the hCD81 LEL. Our results show that non-LEL regions in addition to the LEL determine susceptibility of cells to HCV. While closely related tetraspanins (X. tropicalis CD81 and D. rerio CD81 functionally complement hCD81 non-LEL regions, distantly related tetraspanins (C. elegans TSP9 amd D. melanogaster TSP96F do not and tetraspanins with intermediate homology (hCD9 show an intermediate phenotype. Tetraspanin homology and susceptibility to HCV correlate positively. For some chimeras, infectivity correlates with surface expression. In contrast, the hCD9 chimera is fully surface expressed, binds HCV E2 glycoprotein but is impaired in HCV receptor function. We demonstrate that a cholesterol-coordinating glutamate residue in CD81, which hCD9 lacks, promotes HCV infection. This work highlights the hCD81 non-LEL regions as additional HCV susceptibility-determining factors.

Sox2 Is an Androgen Receptor-Repressed Gene That Promotes Castration-Resistant Prostate Cancer

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Kregel, Steven; Kiriluk, Kyle J.; Rosen, Alex M.; Cai, Yi; Reyes, Edwin E.; Otto, Kristen B.; Tom, Westin; Paner, Gladell P.; Szmulewitz, Russell Z.; Vander Griend, Donald J.

2013-01-01

Despite advances in detection and therapy, castration-resistant prostate cancer continues to be a major clinical problem. The aberrant activity of stem cell pathways, and their regulation by the Androgen Receptor (AR), has the potential to provide insight into novel mechanisms and pathways to prevent and treat advanced, castrate-resistant prostate cancers. To this end, we investigated the role of the embryonic stem cell regulator Sox2 [SRY (sex determining region Y)-box 2] in normal and malignant prostate epithelial cells. In the normal prostate, Sox2 is expressed in a portion of basal epithelial cells. Prostate tumors were either Sox2-positive or Sox2-negative, with the percentage of Sox2-positive tumors increasing with Gleason Score and metastases. In the castration-resistant prostate cancer cell line CWR-R1, endogenous expression of Sox2 was repressed by AR signaling, and AR chromatin-IP shows that AR binds the enhancer element within the Sox2 promoter. Likewise, in normal prostate epithelial cells and human embryonic stem cells, increased AR signaling also decreases Sox2 expression. Resistance to the anti-androgen MDV3100 results in a marked increase in Sox2 expression within three prostate cancer cell lines, and in the castration-sensitive LAPC-4 prostate cancer cell line ectopic expression of Sox2 was sufficient to promote castration-resistant tumor formation. Loss of Sox2 expression in the castration-resistant CWR-R1 prostate cancer cell line inhibited cell growth. Up-regulation of Sox2 was not associated with increased CD133 expression but was associated with increased FGF5 (Fibroblast Growth Factor 5) expression. These data propose a model of elevated Sox2 expression due to loss of AR-mediated repression during castration, and consequent castration-resistance via mechanisms not involving induction of canonical embryonic stem cell pathways. PMID:23326489

Natural selection in a population of Drosophila melanogaster explained by changes in gene expression caused by sequence variation in core promoter regions.

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Sato, Mitsuhiko P; Makino, Takashi; Kawata, Masakado

2016-02-09

Understanding the evolutionary forces that influence variation in gene regulatory regions in natural populations is an important challenge for evolutionary biology because natural selection for such variations could promote adaptive phenotypic evolution. Recently, whole-genome sequence analyses have identified regulatory regions subject to natural selection. However, these studies could not identify the relationship between sequence variation in the detected regions and change in gene expression levels. We analyzed sequence variations in core promoter regions, which are critical regions for gene regulation in higher eukaryotes, in a natural population of Drosophila melanogaster, and identified core promoter sequence variations associated with differences in gene expression levels subjected to natural selection. Among the core promoter regions whose sequence variation could change transcription factor binding sites and explain differences in expression levels, three core promoter regions were detected as candidates associated with purifying selection or selective sweep and seven as candidates associated with balancing selection, excluding the possibility of linkage between these regions and core promoter regions. CHKov1, which confers resistance to the sigma virus and related insecticides, was identified as core promoter regions that has been subject to selective sweep, although it could not be denied that selection for variation in core promoter regions was due to linked single nucleotide polymorphisms in the regulatory region outside core promoter regions. Nucleotide changes in core promoter regions of CHKov1 caused the loss of two basal transcription factor binding sites and acquisition of one transcription factor binding site, resulting in decreased gene expression levels. Of nine core promoter regions regions associated with balancing selection, brat, and CG9044 are associated with neuromuscular junction development, and Nmda1 are associated with learning

Association between a promoter dopamine D2 receptor gene variant and the personality trait detachment.

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Jönsson, Erik G; Cichon, Sven; Gustavsson, J Petter; Grünhage, Frank; Forslund, Kaj; Mattila-Evenden, Marja; Rylander, Gunnar; Asberg, Marie; Farde, Lars; Propping, Peter; Nöthen, Markus M

2003-04-01

Personality traits have shown considerable heritable components. Striatal dopamine D(2) receptor density, as determined by positron-emission tomography, has been associated with detached personality, as assessed by the Karolinska Scales of Personality. A putative functional promoter polymorphism in the dopamine D(2) receptor gene (DRD2), -141C ins/del, has been associated with dopamine D(2) receptor density. In this study healthy subjects (n = 235) who filled in at least one of several personality questionnaires (Karolinska Scales of Personality, Swedish Universities Scales of Personality, Health-relevant Five-factor Personality Inventory, and Temperament and Character Inventory) were analyzed with regard to the DRD2 -141C ins/del variant. There was an association (p =.001) between the DRD2 -141C ins/del variant and Karolinska Scales of Personality Detachment scale, indicating higher scores in subjects with the -141C del variant. There were also associations between the DRD2 -141C ins/del variant and a number of Karolinska Scales of Personality and Swedish Universities Scales of Personality Neuroticism-related scales, but of these only Swedish Universities Scales of Personality Lack of Assertiveness scale (p =.001) survived correction for multiple testing. These results add further support for the involvement of dopamine D(2) receptor in certain personality traits. The results should be treated with caution until replicated.

Modulation of thyroid hormone receptor transactivation by the early region 1A (E1A-like inhibitor of differentiation 1 (EID1

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Diana Vargas

2008-01-01

Full Text Available Transcriptional activation (TA mediated by the effect of thyroid hormones on target genes requires co-activator proteins such as the early region 1A (E1A associated 300 kDa binding protein (p300 and the cAMP response element binding protein (CREB binding protein (CBP, known as the p300/CBP complex, which acetylate histones 3 and 4 to allow transcriptional machinery access to the target gene promoter. Little is known on the role of p300 in thyroid hormone receptor (TR mediated TA but the E1A-like inhibitor of differentiation 1 (EID1, an inhibitor of p300 histone acetyltransferase (HAT, is a functional homolog of E1A and may inhibit myogenic differentiation factor D (MyoD transcriptional activity and reduces muscle cell differentiation. We evaluated the influence of EID1 on TR-mediated transcriptional activity using transfection and mammalian two-hybrid studies to show that EID1 may partially reduces TA activity of the TR receptor, probably due to p300 blockage since EID1 mutants cannot reduce TR-mediated TA. The EID1 does not affect the function of p160 co-activator proteins (160 kDa proteins of steroid receptor co-activators and is functionally independent of co-repressor proteins or TR binding. Summarizing, EID1 reduces TR-mediated transcriptional activity by blocking p300 and may play an important role in thyroid receptor activity in muscle and other tissues.

The human oxytocin gene promoter is regulated by estrogens.

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Richard, S; Zingg, H H

1990-04-15

Gonadal steroids affect brain function primarily by altering the expression of specific genes, yet the specific mechanisms by which neuronal target genes undergo such regulation are unknown. Recent evidence suggests that the expression of the neuropeptide gene for oxytocin (OT) is modulated by estrogens. We therefore examined the possibility that this regulation occurred via a direct interaction of the estrogen-receptor complex with cis-acting elements flanking the OT gene. DNA-mediated gene transfer experiments were performed using Neuro-2a neuroblastoma cells and chimeric plasmids containing portions of the human OT gene 5'-glanking region linked to the chloramphenicol acetyltransferase gene. We identified a 19-base pair region located at -164 to -146 upstream of the transcription start site which is capable of conferring estrogen responsiveness to the homologous as well as to a heterologous promoter. The hormonal response is strictly dependent on the presence of intracellular estrogen receptors, since estrogen induced stimulation occurred only in Neuro-2a cells co-transfected with an expression vector for the human estrogen receptor. The identified region contains a novel imperfect palindrome (GGTGACCTTGACC) with sequence similarity to other estrogen response elements (EREs). To define cis-acting elements that function in synergism with the ERE, sequences 3' to the ERE were deleted, including the CCAAT box, two additional motifs corresponding to the right half of the ERE palindrome (TGACC), as well as a CTGCTAA heptamer similar to the "elegans box" found in Caenorhabditis elegans. Interestingly, optimal function of the identified ERE was fully independent of these elements and only required a short promoter region (-49 to +36). Our studies define a molecular mechanism by which estrogens can directly modulate OT gene expression. However, only a subset of OT neurons are capable of binding estrogens, therefore, direct action of estrogens on the OT gene may be

Role of sigma-1 receptors in neurodegenerative diseases

Directory of Open Access Journals (Sweden)

Linda Nguyen

2015-01-01

Full Text Available Neurodegenerative diseases with distinct genetic etiologies and pathological phenotypes appear to share common mechanisms of neuronal cellular dysfunction, including excitotoxicity, calcium dysregulation, oxidative damage, ER stress and mitochondrial dysfunction. Glial cells, including microglia and astrocytes, play an increasingly recognized role in both the promotion and prevention of neurodegeneration. Sigma receptors, particularly the sigma-1 receptor subtype, which are expressed in both neurons and glia of multiple regions within the central nervous system, are a unique class of intracellular proteins that can modulate many biological mechanisms associated with neurodegeneration. These receptors therefore represent compelling putative targets for pharmacologically treating neurodegenerative disorders. In this review, we provide an overview of the biological mechanisms frequently associated with neurodegeneration, and discuss how sigma-1 receptors may alter these mechanisms to preserve or restore neuronal function. In addition, we speculate on their therapeutic potential in the treatment of various neurodegenerative disorders.

Two negative cis-regulatory regions involved in fruit-specific promoter activity from watermelon (Citrullus vulgaris S.).

Science.gov (United States)

Yin, Tao; Wu, Hanying; Zhang, Shanglong; Lu, Hongyu; Zhang, Lingxiao; Xu, Yong; Chen, Daming; Liu, Jingmei

2009-01-01

A 1.8 kb 5'-flanking region of the large subunit of ADP-glucose pyrophosphorylase, isolated from watermelon (Citrullus vulgaris S.), has fruit-specific promoter activity in transgenic tomato plants. Two negative regulatory regions, from -986 to -959 and from -472 to -424, were identified in this promoter region by fine deletion analyses. Removal of both regions led to constitutive expression in epidermal cells. Gain-of-function experiments showed that these two regions were sufficient to inhibit RFP (red fluorescent protein) expression in transformed epidermal cells when fused to the cauliflower mosaic virus (CaMV) 35S minimal promoter. Gel mobility shift experiments demonstrated the presence of leaf nuclear factors that interact with these two elements. A TCCAAAA motif was identified in these two regions, as well as one in the reverse orientation, which was confirmed to be a novel specific cis-element. A quantitative beta-glucuronidase (GUS) activity assay of stable transgenic tomato plants showed that the activities of chimeric promoters harbouring only one of the two cis-elements, or both, were approximately 10-fold higher in fruits than in leaves. These data confirm that the TCCAAAA motif functions as a fruit-specific element by inhibiting gene expression in leaves.

CDK11p58 represses vitamin D receptor-mediated transcriptional activation through promoting its ubiquitin-proteasome degradation

International Nuclear Information System (INIS)

Chi, Yayun; Hong, Yi; Zong, Hongliang; Wang, Yanlin; Zou, Weiying; Yang, Junwu; Kong, Xiangfei; Yun, Xiaojing; Gu, Jianxin

2009-01-01

Vitamin D receptor (VDR) is a member of the nuclear receptor superfamily and regulates transcription of target genes. In this study, we identified CDK11 p58 as a novel protein involved in the regulation of VDR. CDK11 p58 , a member of the large family of p34cdc2-related kinases, is associated with cell cycle progression, tumorigenesis, and apoptotic signaling. Our study demonstrated that CDK11 p58 interacted with VDR and repressed VDR-dependent transcriptional activation. Furthermore, overexpression of CDK11 p58 decreased the stability of VDR through promoting its ubiquitin-proteasome-mediated degradation. Taken together, these results suggest that CDK11 p58 is involved in the negative regulation of VDR.

Methylation of the leukocyte glucocorticoid receptor gene promoter in adults: associations with early adversity and depressive, anxiety and substance-use disorders.

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Tyrka, A R; Parade, S H; Welch, E S; Ridout, K K; Price, L H; Marsit, C; Philip, N S; Carpenter, L L

2016-07-05

Early adversity increases risk for developing psychopathology. Epigenetic modification of stress reactivity genes is a likely mechanism contributing to this risk. The glucocorticoid receptor (GR) gene is of particular interest because of the regulatory role of the GR in hypothalamic-pituitary-adrenal (HPA) axis function. Mounting evidence suggests that early adversity is associated with GR promoter methylation and gene expression. Few studies have examined links between GR promoter methylation and psychopathology, and findings to date have been mixed. Healthy adult participants (N=340) who were free of psychotropic medications reported on their childhood experiences of maltreatment and parental death and desertion. Lifetime depressive and anxiety disorders and past substance-use disorders were assessed using the Structured Clinical Interview for the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition. Methylation of exon 1F of the GR gene (NR3C1) was examined in leukocyte DNA via pyrosequencing. On a separate day, a subset of the participants (n=231) completed the dexamethasone/corticotropin-releasing hormone (Dex/CRH) test. Childhood adversity and a history of past substance-use disorder and current or past depressive or anxiety disorders were associated with lower levels of NR3C1 promoter methylation across the region as a whole and at individual CpG sites (Pdisorder. GR promoter methylation was linked to altered cortisol responses to the Dex/CRH test (Pdepressive, anxiety and substance-use disorders in adults. This finding stands in contrast to our prior work, but is consistent with emerging findings, suggesting complexity in the regulation of this gene.

Osteoblast-specific transcription factor Osterix increases vitamin D receptor gene expression in osteoblasts.

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Chi Zhang

Full Text Available Osterix (Osx is an osteoblast-specific transcription factor required for osteoblast differentiation from mesenchymal stem cells. In Osx knock-out mice, no bone formation occurs. The vitamin D receptor (VDR is a member of the nuclear hormone receptor superfamily that regulates target gene transcription to ensure appropriate control of calcium homeostasis and bone development. Here, we provide several lines of evidence that show that the VDR gene is a target for transcriptional regulation by Osx in osteoblasts. For example, calvaria obtained from Osx-null embryos displayed dramatic reductions in VDR expression compared to wild-type calvaria. Stable overexpression of Osx stimulated VDR expression in C2C12 mesenchymal cells. Inhibition of Osx expression by siRNA led to downregulation of VDR. In contrast, Osx levels remained unchanged in osteoblasts in VDR-null mice. Mechanistic approaches using transient transfection assays showed that Osx directly activated a 1 kb fragment of the VDR promoter in a dose-dependent manner. To define the region of the VDR promoter that was responsive to Osx, a series of VDR promoter deletion mutants were examined and the minimal Osx-responsive region was refined to the proximal 120 bp of the VDR promoter. Additional point mutants were used to identify two GC-rich regions that were responsible for VDR promoter activation by Osx. Chromatin immunoprecipitation assays demonstrated that endogenous Osx was associated with the native VDR promoter in primary osteoblasts in vivo. Cumulatively, these data strongly support a direct regulatory role for Osx in VDR gene expression. They further provide new insight into potential mechanisms and pathways that Osx controls in osteoblasts and during the process of osteoblastic cell differentiation.

Genome-wide function of H2B ubiquitylation in promoter and genic regions.

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Batta, Kiran; Zhang, Zhenhai; Yen, Kuangyu; Goffman, David B; Pugh, B Franklin

2011-11-01

Nucleosomal organization in and around genes may contribute substantially to transcriptional regulation. The contribution of histone modifications to genome-wide nucleosomal organization has not been systematically evaluated. In the present study, we examine the role of H2BK123 ubiquitylation, a key regulator of several histone modifications, on nucleosomal organization at promoter, genic, and transcription termination regions in Saccharomyces cerevisiae. Using high-resolution MNase chromatin immunoprecipitation and sequencing (ChIP-seq), we map nucleosome positioning and occupancy in mutants of the H2BK123 ubiquitylation pathway. We found that H2B ubiquitylation-mediated nucleosome formation and/or stability inhibits the assembly of the transcription machinery at normally quiescent promoters, whereas ubiquitylation within highly active gene bodies promotes transcription elongation. This regulation does not proceed through ubiquitylation-regulated histone marks at H3K4, K36, and K79. Our findings suggest that mechanistically similar functions of H2B ubiquitylation (nucleosome assembly) elicit different functional outcomes on genes depending on its positional context in promoters (repressive) versus transcribed regions (activating).

Tumor-extrinsic discoidin domain receptor 1 promotes mammary tumor growth by regulating adipose stromal interleukin 6 production in mice.

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Sun, Xiujie; Gupta, Kshama; Wu, Bogang; Zhang, Deyi; Yuan, Bin; Zhang, Xiaowen; Chiang, Huai-Chin; Zhang, Chi; Curiel, Tyler J; Bendeck, Michelle P; Hursting, Stephen; Hu, Yanfen; Li, Rong

2018-02-23

Discoidin domain receptor 1 (DDR1) is a collagen receptor that mediates cell communication with the extracellular matrix (ECM). Aberrant expression and activity of DDR1 in tumor cells are known to promote tumor growth. Although elevated DDR1 levels in the stroma of breast tumors are associated with poor patient outcome, a causal role for tumor-extrinsic DDR1 in cancer promotion remains unclear. Here we report that murine mammary tumor cells transplanted to syngeneic recipient mice in which Ddr1 has been knocked out (KO) grow less robustly than in WT mice. We also found that the tumor-associated stroma in Ddr1- KO mice exhibits reduced collagen deposition compared with the WT controls, supporting a role for stromal DDR1 in ECM remodeling of the tumor microenvironment. Furthermore, the stromal-vascular fraction (SVF) of Ddr1 knockout adipose tissue, which contains committed adipose stem/progenitor cells and preadipocytes, was impaired in its ability to stimulate tumor cell migration and invasion. Cytokine array-based screening identified interleukin 6 (IL-6) as a cytokine secreted by the SVF in a DDR1-dependent manner. SVF-produced IL-6 is important for SVF-stimulated tumor cell invasion in vitro , and, using antibody-based neutralization, we show that tumor promotion by IL-6 in vivo requires DDR1. In conclusion, our work demonstrates a previously unrecognized function of DDR1 in promoting tumor growth. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Surface Display of the Receptor-Binding Region of the Lactobacillus brevis S-Layer Protein in Lactococcus lactis Provides Nonadhesive Lactococci with the Ability To Adhere to Intestinal Epithelial Cells

OpenAIRE

Åvall-Jääskeläinen, Silja; Lindholm, Agneta; Palva, Airi

2003-01-01

Lactobacillus brevis is a promising lactic acid bacterium for use as a probiotic dietary adjunct and a vaccine vector. The N-terminal region of the S-layer protein (SlpA) of L. brevis ATCC 8287 was recently shown to mediate adhesion to various human cell lines in vitro. In this study, a surface display cassette was constructed on the basis of this SlpA receptor-binding domain, a proteinase spacer, and an autolysin anchor. The cassette was expressed under control of the nisA promoter in Lactoc...

CINPA1 Is an Inhibitor of Constitutive Androstane Receptor That Does Not Activate Pregnane X Receptor

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Cherian, Milu T; Lin, Wenwei; Wu, Jing

2015-01-01

Constitutive androstane receptor (CAR) and pregnane X receptor (PXR) are xenobiotic sensors that enhance the detoxification and elimination of xenobiotics and endobiotics by modulating the expression of genes encoding drug-metabolizing enzymes and transporters. Elevated levels of drug-metabolizing enzymes and efflux transporters, resulting from CAR activation in various cancers, promote the elimination of chemotherapeutic agents, leading to reduced therapeutic effectiveness and acquired drug resistance. CAR inhibitors, in combination with existing chemotherapeutics, could therefore be used to attenuate multidrug resistance in cancers. Interestingly, all previously reported CAR inverse-agonists are also activators of PXR, rendering them mechanistically counterproductive in tissues where both these xenobiotic receptors are present and active. We used a directed high-throughput screening approach, followed by subsequent mechanistic studies, to identify novel, potent, and specific small-molecule CAR inhibitors that do not activate PXR. We describe here one such inhibitor, CINPA1 (CAR inhibitor not PXR activator 1), capable of reducing CAR-mediated transcription with an IC50 of ∼70 nM. CINPA1 1) is a specific xenobiotic receptor inhibitor and has no cytotoxic effects up to 30 µM; 2) inhibits CAR-mediated gene expression in primary human hepatocytes, where CAR is endogenously expressed; 3) does not alter the protein levels or subcellular localization of CAR; 4) increases corepressor and reduces coactivator interaction with the CAR ligand-binding domain in mammalian two-hybrid assays; and 5) disrupts CAR binding to the promoter regions of target genes in chromatin immunoprecipitation assays. CINPA1 could be used as a novel molecular tool for understanding CAR function. PMID:25762023

The promotion of regional integration of electricity markets: Lessons for developing countries

International Nuclear Information System (INIS)

Oseni, Musiliu O.; Pollitt, Michael G.

2016-01-01

This paper focuses on how to promote regional cooperation in electricity. We begin by discussing the theory of international trade cooperation in electricity, with a view to discussing what preconditions might be important in facilitating wide area trading across national borders. We then develop lessons based on the comparison of four case studies. These include three regional developing country power pools – the Southern African Power pool (SAPP), West African Power pool (WAPP) and the Central American Power Market (MER). We contrast these with Northern Europe's Nord Pool. These cases highlight both the potential and difficulty of having cross-jurisdictional power pools. In the light of the theory and evidence we present, we draw key lessons in the areas of: preconditions for trading; necessary institutional arrangements; practicalities of timetabling; reasons to be hopeful about future prospects. - Highlights: • This paper focuses on how to promote regional electricity cooperation. • We develop lessons based on comparison of four international case studies. • The cases highlight both the potential and difficulty of power pools. • We identify preconditions, institutional arrangements and timetabling. • We conclude that the future prospects for regional power pools are good.

Multiple serotonin receptors: regional distribution and effect of raphe lesions

International Nuclear Information System (INIS)

Blackshear, M.A.; Sanders-Bush, E.; Steranka, L.R.

1981-01-01

These studies confirm and extend the recent work suggesting that [ 3 H]lysergic acid diethylamide (LSD) labels two distinct binding sites in rat brain resembling serotonin (5HT) receptors. Although Scatchard analyses of [ 3 H]LSD binding to membranes prepared from cortex/hippocampus were linear, the heterogeneity of the [ 3 H]LSD binding sites was clearly demonstrated in displacement studies. The displacement curves for both 5HT and spiperone were bisigmoidal with the concentration required to saturate the high affinity components nearly 3 orders of magnitude lower than the concentrations necessary to saturate the low affinity components. Additivity studies suggested that the sites with high affinity for 5HT and spiperone are different, independent sites. These sites are referred to as 5HT 1 and 5HT 2 respectively. Regional analyses showed, that in the frontal cortex, the density of the 5HT 2 site was slightly greater than the 5HT 1 site whereas the 5HT 1 site was predominant in all other brain areas, including the spinal cord. The pharmacological properties of the two sites have features in common with 5HT receptors; however, electrolytic lesions of the midbrain raphe nuclei did not change the densities or binding constants of the two apparent 5HT receptor subtypes, even though the number of high affinity 5HT uptake sites was markedly reduced. (Auth.)

Differences in Signal Activation by LH and hCG are Mediated by the LH/CG Receptor's Extracellular Hinge Region.

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Grzesik, Paul; Kreuchwig, Annika; Rutz, Claudia; Furkert, Jens; Wiesner, Burkhard; Schuelein, Ralf; Kleinau, Gunnar; Gromoll, Joerg; Krause, Gerd

2015-01-01

The human lutropin (hLH)/choriogonadotropin (hCG) receptor (LHCGR) can be activated by binding two slightly different gonadotropic glycoprotein hormones, choriogonadotropin (CG) - secreted by the placenta, and lutropin (LH) - produced by the pituitary. They induce different signaling profiles at the LHCGR. This cannot be explained by binding to the receptor's leucine-rich-repeat domain (LRRD), as this binding is similar for the two hormones. We therefore speculate that there are previously unknown differences in the hormone/receptor interaction at the extracellular hinge region, which might help to understand functional differences between the two hormones. We have therefore performed a detailed study of the binding and action of LH and CG at the LHCGR hinge region. We focused on a primate-specific additional exon in the hinge region, which is located between LRRD and the serpentine domain. The segment of the hinge region encoded by exon10 was previously reported to be only relevant to hLH signaling, as the exon10-deletion receptor exhibits decreased hLH signaling, but unchanged hCG signaling. We designed an advanced homology model of the hormone/LHCGR complex, followed by experimental characterization of relevant fragments in the hinge region. In addition, we examined predictions of a helical exon10-encoded conformation by block-wise polyalanine (helix supporting) mutations. These helix preserving modifications showed no effect on hormone-induced signaling. However, introduction of a structure-disturbing double-proline mutant LHCGR-Q303P/E305P within the exon10-helix has, in contrast to exon10-deletion, no impact on hLH, but only on hCG signaling. This opposite effect on signaling by hLH and hCG can be explained by distinct sites of hormone interaction in the hinge region. In conclusion, our analysis provides details of the differences between hLH- and hCG-induced signaling that are mainly determined in the L2-beta loop of the hormones and in the hinge

Low Density Lipoprotein Receptor Class A Repeats Are O-Glycosylated in Linker Regions

DEFF Research Database (Denmark)

Pedersen, Nis Borbye; Wang, Shengjun; Narimatsu, Yoshiki

2014-01-01

, which in wild-type CHO cells is glycosylated with the typical sialylated core 1 structure. The glycosites in linker regions of LDLR class A repeats are conserved in LDLR from man to Xenopus and found in other homologous receptors. O-Glycosylation is controlled by a large family of polypeptide Gal...

Comparison of benzodiazepine receptor and regional cerebral blood flow imagings of epileptiform foci in hippocampal kindled rabbits

International Nuclear Information System (INIS)

Kurokawa, Kenzo

1993-01-01

To compare the benzodiazepine (Bz) receptor imaging and regional cerebral blood flow (rCBF) imaging in the detection of epileptic foci, the distribution pattern of the Bz receptor and rCBF in hippocampal kindled rabbits was examined by a double tracer autoradiography using ethyl 7-[ 125 I]-5,6-dihydro-5-methyl-6-oxo-4H-imidazo [1, 5-a][1,4] benzodiazepine-3-carboxylate ( 125 I-Ro 16-0154) and 99m Tc-hexamethyl-propyleneamine oxime ( 99m Tc-HMPAO). In visual and quantitative analyses, 125 I-Ro 16-0154 accumulation in brain slices extracted after the completion of the kindling was markedly and extensively decreased in the kindled CA1 region mimicking a primary epileptic focus. 125 I-Ro 16-0154 accumulation was moderately decreased in the ipsilateral temporal lobe, dentate gyrus, CA2, CA4, and bilateral CA3 regions, regarded as the propagated sites of seizure discharges. 99m Tc-HMPAO accumulation was found to be decreased in the ipsilateral CA1, frontal, temporal and dentate gyri. However, the decrease was much more slight and less extensive than that in 125 I-Ro 16-0154 accumulation. These results suggest that Bz receptor imaging is much more sensitive in the detection of epileptic foci than rCBF imaging, and therefore that Bz receptor imaging is useful in clinical epilepsy. (author)

High fructose consumption induces DNA methylation at PPARα and CPT1A promoter regions in the rat liver

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Ohashi, Koji [Department of Clinical Biochemistry, Fujita Health University School of Health Sciences, Toyoake (Japan); Munetsuna, Eiji [Department of Biochemistry, Fujita Health University School of Medicine, Toyoake (Japan); Yamada, Hiroya, E-mail: hyamada@fujita-hu.ac.jp [Department of Hygiene, Fujita Health University School of Medicine, Toyoake (Japan); Ando, Yoshitaka [Department of Joint Research Laboratory of Clinical Medicine, Fujita Health University Hospital, Toyoake (Japan); Yamazaki, Mirai; Taromaru, Nao; Nagura, Ayuri; Ishikawa, Hiroaki [Department of Clinical Biochemistry, Fujita Health University School of Health Sciences, Toyoake (Japan); Suzuki, Koji [Department of Public Health, Fujita Health University School of Health Sciences, Toyoake (Japan); Teradaira, Ryoji [Department of Clinical Biochemistry, Fujita Health University School of Health Sciences, Toyoake (Japan); Hashimoto, Shuji [Department of Hygiene, Fujita Health University School of Medicine, Toyoake (Japan)

2015-12-04

DNA methylation status is affected by environmental factors, including nutrition. Fructose consumption is considered a risk factor for the conditions that make up metabolic syndrome such as dyslipidemia. However, the pathogenetic mechanism by which fructose consumption leads to metabolic syndrome is unclear. Based on observations that epigenetic modifications are closely related to induction of metabolic syndrome, we hypothesized that fructose-induced metabolic syndrome is caused by epigenetic alterations. Male SD rats were designated to receive water or 20% fructose solution for 14 weeks. mRNA levels for peroxisome proliferator-activated receptor alpha (PPARα) and carnitine palmitoyltransferase 1A (CPT1A) was analyzed using Real-time PCR. Restriction digestion and real-time PCR (qAMP) was used for the analysis of DNA methylation status. Hepatic lipid accumulation was also observed by fructose intake. Fructose feeding also significantly decreased mRNA levels for PPARα and CPT1A. qAMP analysis demonstrated the hypermethylation of promoter regions of PPARα and CTP1A genes. Fructose-mediated attenuated gene expression may be mediated by alterations of DNA methylation status, and pathogenesis of metabolic syndrome induced by fructose relates to DNA methylation status. - Highlights: • No general consensus has been reached regarding the molecular mechanisms of the pathogenesis of fructose-induced diseases. • Significant increase in hepatic total methylation level was observed after fructose-supplemented feeding. • Fructose feeding significantly decreased mRNA levels for PPARα and CPT1A. • qAMP analysis demonstrated the hypermethylation of promoter regions of PPARα and CTP1A genes. • Fructose-mediated attenuated gene expression may be mediated by alterations of DNA methylation status in rat liver.

High fructose consumption induces DNA methylation at PPARα and CPT1A promoter regions in the rat liver

International Nuclear Information System (INIS)

Ohashi, Koji; Munetsuna, Eiji; Yamada, Hiroya; Ando, Yoshitaka; Yamazaki, Mirai; Taromaru, Nao; Nagura, Ayuri; Ishikawa, Hiroaki; Suzuki, Koji; Teradaira, Ryoji; Hashimoto, Shuji

2015-01-01

DNA methylation status is affected by environmental factors, including nutrition. Fructose consumption is considered a risk factor for the conditions that make up metabolic syndrome such as dyslipidemia. However, the pathogenetic mechanism by which fructose consumption leads to metabolic syndrome is unclear. Based on observations that epigenetic modifications are closely related to induction of metabolic syndrome, we hypothesized that fructose-induced metabolic syndrome is caused by epigenetic alterations. Male SD rats were designated to receive water or 20% fructose solution for 14 weeks. mRNA levels for peroxisome proliferator-activated receptor alpha (PPARα) and carnitine palmitoyltransferase 1A (CPT1A) was analyzed using Real-time PCR. Restriction digestion and real-time PCR (qAMP) was used for the analysis of DNA methylation status. Hepatic lipid accumulation was also observed by fructose intake. Fructose feeding also significantly decreased mRNA levels for PPARα and CPT1A. qAMP analysis demonstrated the hypermethylation of promoter regions of PPARα and CTP1A genes. Fructose-mediated attenuated gene expression may be mediated by alterations of DNA methylation status, and pathogenesis of metabolic syndrome induced by fructose relates to DNA methylation status. - Highlights: • No general consensus has been reached regarding the molecular mechanisms of the pathogenesis of fructose-induced diseases. • Significant increase in hepatic total methylation level was observed after fructose-supplemented feeding. • Fructose feeding significantly decreased mRNA levels for PPARα and CPT1A. • qAMP analysis demonstrated the hypermethylation of promoter regions of PPARα and CTP1A genes. • Fructose-mediated attenuated gene expression may be mediated by alterations of DNA methylation status in rat liver.

The Innate Immune Receptor NLRX1 Functions as a Tumor Suppressor by Reducing Colon Tumorigenesis and Key Tumor-Promoting Signals

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A. Alicia Koblansky

2016-03-01

Full Text Available NOD-like receptor (NLR proteins are intracellular innate immune sensors/receptors that regulate immunity. This work shows that NLRX1 serves as a tumor suppressor in colitis-associated cancer (CAC and sporadic colon cancer by keeping key tumor promoting pathways in check. Nlrx1−/− mice were highly susceptible to CAC, showing increases in key cancer-promoting pathways including nuclear factor κB (NF-κB, mitogen-activated protein kinase (MAPK, signal transducer and activator of transcription 3 (STAT3, and interleukin 6 (IL-6. The tumor-suppressive function of NLRX1 originated primarily from the non-hematopoietic compartment. This prompted an analysis of NLRX1 function in the Apcmin/+ genetic model of sporadic gastrointestinal cancer. NLRX1 attenuated Apcmin/+ colon tumorigenesis, cellular proliferation, NF-κB, MAPK, STAT3 activation, and IL-6 levels. Application of anti-interleukin 6 receptor (IL6R antibody therapy reduced tumor burden, increased survival, and reduced STAT3 activation in Nlrx1−/−Apcmin/+ mice. As an important clinical correlate, human colon cancer samples expressed lower levels of NLRX1 than healthy controls in multiple patient cohorts. These data implicate anti-IL6R as a potential personalized therapy for colon cancers with reduced NLRX1.

Study on the binding sites of radiosensitivity associated transcription factor in the promoter region of Ier5 gene

International Nuclear Information System (INIS)

Cui Wei; Yin Lingling; Dong Lingyue

2012-01-01

Objective: To clarify the mechanism of immediate early response gene 5 (Ier5) transcription induced by radiation. Methods: Deletant construction, site-specific mutagenesis,electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) were used to forecast the promoter region, binding sites and transcription factors of Ier5 gene in HeLa cells. Results: The promoter region of Ier5 gene might be in the region of Ier5 -8 deletant (-408 - -238 bp). The Ier5 gene had two transcription factors of GCF and NFI, and GCF had two binding sites located in the region of -388 - -382 bp and -274 - -270 bp of Ier5 promoter. The binding site of NFI was located in -362 - -357 bp of Ier5 promoter. GCF could inhibit the expression of Ier5 gene and this inhibition was diminished when the radiation dose increased. In contrast, NFI increased the expression of Ier5. Conclusions: The most possible region of Ier5 promoter is from -408 to -238 bp which has two binding sites for the radiosensitivity transcription factors of GCF and NFI that could negatively and positively regulate the expression of Ier5 respectively. (authors)

Liver X receptor α and farnesoid X receptor are major transcriptional regulators of OATP1B1.

Science.gov (United States)

Meyer Zu Schwabedissen, Henriette E; Böttcher, Kerstin; Chaudhry, Amarjit; Kroemer, Heyo K; Schuetz, Erin G; Kim, Richard B

2010-11-01

Organic anion transporting polypeptide 1B1 (OATP1B1) is a liver-enriched transporter involved in the hepatocellular uptake of many endogenous molecules and several structurally divergent drugs in clinical use. Although OATP1B1 coding region polymorphisms are known to make an impact on substrate drug disposition in humans, little is known regarding the mechanisms underlying the transcriptional regulation of this transporter. In this study, we note that messenger RNA (mRNA) expression of OATP1B1 in a large human liver bank exhibited marked interindividual variability that was not associated with coding region polymorphisms. Accordingly, we hypothesized that such variability in expression is reflective of nuclear receptor-mediated transcriptional regulation of this transporter. We tested prototypical ligands for the nuclear receptors pregnane X receptor (PXR), constitutive androstane receptor (CAR), liver X receptor (LXR) α, and farnesoid X receptor (FXR) in a human hepatoma-derived cell line and noted induction of OATP1B1 mRNA when the cells were treated with LXRα or FXR ligands. To confirm a direct role for LXRα and FXR to OATP1B1 expression, we performed detailed promoter analysis and cell-based reporter gene assays resulting in the identification of two functional FXR response elements and one LXRα response element. The direct interaction between nuclear receptors with the identified response elements was assessed using chromatin immunoprecipitation assays. Using isolated primary human hepatocytes, we show that LXRα or FXR agonists, but not PXR or CAR agonists, are capable of OATP1B1 induction. We note that OATP1B1 transcriptional regulation is under dual nuclear receptor control through the oxysterol sensing LXRα and the bile acid sensor FXR. Accordingly, the interplay between OATP1B1 and nuclear receptors may play an important and heretofore unrecognized role during cholestasis, drug-induced liver injury, and OATP1B1 induction-related drug interactions.

DNA Methylation Analysis of BRD1 Promoter Regions and the Schizophrenia rs138880 Risk Allele.

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Mads Dyrvig

Full Text Available The bromodomain containing 1 gene, BRD1 is essential for embryogenesis and CNS development. It encodes a protein that participates in histone modifying complexes and thereby regulates the expression of a large number of genes. Genetic variants in the BRD1 locus show association with schizophrenia and bipolar disorder and risk alleles in the promoter region correlate with reduced BRD1 expression. Insights into the transcriptional regulation of BRD1 and the pathogenic mechanisms associated with BRD1 risk variants, however, remain sparse. By studying transcripts in human HeLa and SH-SY5Y cells we provide evidence for differences in relative expression of BRD1 transcripts with three alternative 5' UTRs (exon 1C, 1B, and 1A. We further show that expression of these transcript variants covaries negatively with DNA methylation proportions in their upstream promoter regions suggesting that promoter usage might be regulated by DNA methylation. In line with findings that the risk allele of the rs138880 SNP in the BRD1 promoter region correlates with reduced BRD1 expression, we find that it is also associated with moderate regional BRD1 promoter hypermethylation in both adipose tissue and blood. Importantly, we demonstrate by inspecting available DNA methylation and expression data that these regions undergo changes in methylation during fetal brain development and that differences in their methylation proportions in fetal compared to postnatal frontal cortex correlate significantly with BRD1 expression. These findings suggest that BRD1 may be dysregulated in both the developing and mature brain of risk allele carriers. Finally, we demonstrate that commonly used mood stabilizers Lithium, Valproate, and Carbamazepine affect the expression of BRD1 in SH-SY5Y cells. Altogether this study indicates a link between genetic risk and epigenetic dysregulation of BRD1 which raises interesting perspectives for targeting the mechanisms pharmacologically.

Regulation of retinoid X receptor gamma expression by fed state in mouse liver

International Nuclear Information System (INIS)

Park, Sangkyu; Lee, Yoo Jeong; Ko, Eun Hee; Kim, Jae-woo

2015-01-01

Glucose metabolism is balanced by glycolysis and gluconeogenesis with precise control in the liver. The expression of genes related to glucose metabolism is regulated primarily by glucose and insulin at transcriptional level. Nuclear receptors play important roles in regulating the gene expression of glucose metabolism at transcriptional level. Some of these nuclear receptors form heterodimers with RXRs to bind to their specific regulatory elements on the target promoters. To date, three isotypes of RXRs have been identified; RXRα, RXRβ and RXRγ. However, their involvement in the interactions with other nuclear receptors in the liver remains unclear. In this study, we found RXRγ is rapidly induced after feeding in the mouse liver, indicating a potential role of RXRγ in controlling glucose or lipid metabolism in the fasting–feeding cycle. In addition, RXRγ expression was upregulated by glucose in primary hepatocytes. This implies that glucose metabolism governed by RXRγ in conjunction with other nuclear receptors. The luciferase reporter assay showed that RXRγ as well as RXRα increased SREBP-1c promoter activity in hepatocytes. These results suggest that RXRγ may play an important role in tight control of glucose metabolism in the fasting–feeding cycle. - Highlights: • Refeeding increases the RXRγ expression level in mouse liver. • RXRγ expression is induced by high glucose condition in primary hepatocytes. • RXRγ and LXRα have synergistic effect on SREBP-1c promoter activity. • RXRγ binds to LXRE(-299/-280) located within SREBP-1c promoter region and interacts with LXRα

Regulation of retinoid X receptor gamma expression by fed state in mouse liver

Energy Technology Data Exchange (ETDEWEB)

Park, Sangkyu, E-mail: 49park@cku.ac.kr [Department of Biochemistry, College of Medicine, Catholic Kwandong University, Gangneung 210-701 (Korea, Republic of); Lee, Yoo Jeong [Division of Metabolic Disease, Center for Biomedical Sciences, National Institute of Health Korea, Osong 361-709 (Korea, Republic of); Ko, Eun Hee [Department of Biochemistry and Molecular Biology, Integrated Genomic Research Center for Metabolic Regulation, Institute of Genetic Science, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Kim, Jae-woo [Department of Biochemistry and Molecular Biology, Integrated Genomic Research Center for Metabolic Regulation, Institute of Genetic Science, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Brain Korea 21 PLUS Project for Medical Science, Yonsei University, Seoul 120-752 (Korea, Republic of)

2015-02-27

Glucose metabolism is balanced by glycolysis and gluconeogenesis with precise control in the liver. The expression of genes related to glucose metabolism is regulated primarily by glucose and insulin at transcriptional level. Nuclear receptors play important roles in regulating the gene expression of glucose metabolism at transcriptional level. Some of these nuclear receptors form heterodimers with RXRs to bind to their specific regulatory elements on the target promoters. To date, three isotypes of RXRs have been identified; RXRα, RXRβ and RXRγ. However, their involvement in the interactions with other nuclear receptors in the liver remains unclear. In this study, we found RXRγ is rapidly induced after feeding in the mouse liver, indicating a potential role of RXRγ in controlling glucose or lipid metabolism in the fasting–feeding cycle. In addition, RXRγ expression was upregulated by glucose in primary hepatocytes. This implies that glucose metabolism governed by RXRγ in conjunction with other nuclear receptors. The luciferase reporter assay showed that RXRγ as well as RXRα increased SREBP-1c promoter activity in hepatocytes. These results suggest that RXRγ may play an important role in tight control of glucose metabolism in the fasting–feeding cycle. - Highlights: • Refeeding increases the RXRγ expression level in mouse liver. • RXRγ expression is induced by high glucose condition in primary hepatocytes. • RXRγ and LXRα have synergistic effect on SREBP-1c promoter activity. • RXRγ binds to LXRE(-299/-280) located within SREBP-1c promoter region and interacts with LXRα.

CISH promoter polymorphism effects on T cell cytokine receptor signaling and type 1 diabetes susceptibility.

Science.gov (United States)

Seyfarth, Julia; Ahlert, Heinz; Rosenbauer, Joachim; Baechle, Christina; Roden, Michael; Holl, Reinhard W; Mayatepek, Ertan; Meissner, Thomas; Jacobsen, Marc

2018-02-06

Impaired regulatory T cell immunity plays a central role in the development of type 1 diabetes (T1D). Interleukin-2 receptor (IL-2R) signaling is essential for regulatory T cells (T REG ), and cytokine-inducible SH2-containing protein (CIS) regulates IL-2R signaling as a feedback inhibitor. Previous studies identified association of CISH promoter region single nucleotide polymorphisms (SNPs) with susceptibility to infectious diseases. Here we analyzed allele frequencies of three CISH SNPs (i.e., rs809451, rs414171, rs2239751) in a study of T1D patients (n = 260, onset age  10 years). Minor allele frequencies were compared to a control cohort of the 1000 Genomes Project. Assigned haplotypes were determined for effects on T1D manifestation and severity. Finally, the CISH haplotype influence on cytokine signaling and function was explored in T cells from healthy donors. We detected similar minor allele frequencies between T1D patients and the control cohort. T1D onset age, residual serum C-peptide level, and insulin requirement were comparable between different haplotypes. Only minor differences between the haplotypes were found for in vitro cytokine (i.e., IL-2, IL-7)-induced CIS mRNA expression. STAT5 phosphorylation was induced by IL-2 or IL-7, but no differences were found between the haplotypes. T REG purified from healthy donors with the two most common haplotypes showed similar capacity to inhibit heterologous effector T cells. This study provides no evidence for an association of CISH promoter SNPs with susceptibility to T1D or severity of disease. In contrast to previous studies, no influence of different haplotypes on CIS mRNA expression or T cell-mediated functions was found.

The Orphan Nuclear Receptor TR4 Is a Vitamin A-activated Nuclear Receptor

Energy Technology Data Exchange (ETDEWEB)

Zhou, X. Edward; Suino-Powell, Kelly M.; Xu, Yong; Chan, Cee-Wah; Tanabe, Osamu; Kruse, Schoen W.; Reynolds, Ross; Engel, James Douglas; Xu, H. Eric (Michigan-Med); (Van Andel)

2015-11-30

Testicular receptors 2 and 4 (TR2/4) constitute a subgroup of orphan nuclear receptors that play important roles in spermatogenesis, lipid and lipoprotein regulation, and the development of the central nervous system. Currently, little is known about the structural features and the ligand regulation of these receptors. Here we report the crystal structure of the ligand-free TR4 ligand binding domain, which reveals an autorepressed conformation. The ligand binding pocket of TR4 is filled by the C-terminal half of helix 10, and the cofactor binding site is occupied by the AF-2 helix, thus preventing ligand-independent activation of the receptor. However, TR4 exhibits constitutive transcriptional activity on multiple promoters, which can be further potentiated by nuclear receptor coactivators. Mutations designed to disrupt cofactor binding, dimerization, or ligand binding substantially reduce the transcriptional activity of this receptor. Importantly, both retinol and retinoic acid are able to promote TR4 to recruit coactivators and to activate a TR4-regulated reporter. These findings demonstrate that TR4 is a ligand-regulated nuclear receptor and suggest that retinoids might have a much wider regulatory role via activation of orphan receptors such as TR4.

Kinetics of activation of the P4 promoter of pBR322 by the Escherichia coli cyclic AMP receptor protein.

Science.gov (United States)

Hoggett, J G; Brierley, I

1992-11-01

The activation of transcription initiation from the P4 promoter of pBR322 by the Escherichia coli cyclic AMP receptor protein (CRP) has been investigated using a fluorescence abortive initiation assay. The effect of the cyclic-AMP/CRP complex on the linear P4 promoter was to increase the initial binding (KB) of RNA polymerase to the promoter by about a factor of 10, but the rate of isomerization of closed to open complex (kf) was unaffected. One molecule of CRP per promoter was required for activation, and the concentration of cyclic AMP producing half-maximal stimulation was about 7-8 microM. Supercoiling caused a 2-3-fold increase in the rate of isomerization of the CRP-activated promoter, but weakened the initial binding of polymerase by about one order of magnitude. The unactivated supercoiled promoter was too weak to allow reliable assessment of kinetic parameters against the high background rate originating from the rest of the plasmid.

Surface display of the receptor-binding region of the Lactobacillus brevis S-layer protein in Lactococcus lactis provides nonadhesive lactococci with the ability to adhere to intestinal epithelial cells.

Science.gov (United States)

Avall-Jääskeläinen, Silja; Lindholm, Agneta; Palva, Airi

2003-04-01

Lactobacillus brevis is a promising lactic acid bacterium for use as a probiotic dietary adjunct and a vaccine vector. The N-terminal region of the S-layer protein (SlpA) of L. brevis ATCC 8287 was recently shown to mediate adhesion to various human cell lines in vitro. In this study, a surface display cassette was constructed on the basis of this SlpA receptor-binding domain, a proteinase spacer, and an autolysin anchor. The cassette was expressed under control of the nisA promoter in Lactococcus lactis NZ9000. Western blot assay of lactococcal cell wall extracts with anti-SlpA antibodies confirmed that the SlpA adhesion domain of the fusion protein was expressed and located within the cell wall layer. Whole-cell enzyme-linked immunosorbent assay and immunofluorescence microscopy verified that the SlpA adhesion-mediating region was accessible on the lactococcal cell surface. In vitro adhesion assays with the human intestinal epithelial cell line Intestine 407 indicated that the recombinant lactococcal cells had gained an ability to adhere to Intestine 407 cells significantly greater than that of wild-type L. lactis NZ9000. Serum inhibition assay further confirmed that adhesion of recombinant lactococci to Intestine 407 cells was indeed mediated by the N terminus-encoding part of the slpA gene. The ability of the receptor-binding region of SlpA to adhere to fibronectin was also confirmed with this lactococcal surface display system. These results show that, with the aid of the receptor-binding region of the L. brevis SlpA protein, the ability to adhere to gut epithelial cells can indeed be transferred to another, nonadhesive, lactic acid bacterium.

Haloperidol, a sigma receptor 1 antagonist, promotes ferroptosis in hepatocellular carcinoma cells.

Science.gov (United States)

Bai, Tao; Wang, Shuai; Zhao, Yipu; Zhu, Rongtao; Wang, Weijie; Sun, Yuling

2017-09-30

Ferroptosis is a novel form of cell death, which is characterized by accumulation of reactive oxygen species (ROS). Sigma 1 receptor (S1R) has been suggested to function in oxidative stress metabolism. Both erastin and sorafenib significantly induced S1R protein expression. Haloperidol strongly promoted erastin- and sorafenib-induced cell death, which was blocked by ferrostatin-1 but not ZVAD-FMK or necrosulfonamide. During ferroptosis, haloperidol substantially increased the cellular levels of Fe 2+ , GSH and lipid peroxidation. Furthermore, several ferroptosis-related protein targets were up-regulated in the absence of haloperidol. Thus, Our study identified an association between haloperidol and ferroptosis for the first time. Our analyses of a combination of drugs may provide a novel strategy of hepatocellular carcinoma (HCC) therapy. Copyright © 2017 Elsevier Inc. All rights reserved.

A study of the frequency of methylation of gene promoter regions in ...

Indian Academy of Sciences (India)

2013-04-02

Apr 2, 2013 ... colorectal cancer in the Taiwanese population. CHANG-CHIEH WU1 ... hypermethylation of promoter-region CpG islands is an important ... mismatch repair gene MLH1 plays an important role in dele- ..... Asia Pac. J. Clin.

Surface Expression of TGF-β Docking Receptor GARP Promotes Oncogenesis and Immune Tolerance in Breast Cancer

OpenAIRE

Metelli, Alessandra; Wu, Bill X; Fugle, Caroline W; Rachidi, Saleh; Sun, Shaoli; Zhang, Yongliang; Wu, Jennifer; Tomlinson, Stephen; Howe, Philip; Yang, Yi; Garrett-Mayer, Elizabeth; Liu, Bei; Li, Zihai

2016-01-01

GARP encoded by the Lrrc32 gene is the cell surface docking receptor for latent TGF-β which is expressed naturally by platelets and regulatory T cells. Although Lrrc32 is amplified frequently in breast cancer, the expression and relevant functions of GARP in cancer have not been explored. Here we report that GARP exerts oncogenic effects, promoting immune tolerance by enriching and activating latent TGF-β in the tumor microenvironment. We found that human breast, lung and colon cancers expres...

Diosgenin promotes oligodendrocyte progenitor cell differentiation through estrogen receptor-mediated ERK1/2 activation to accelerate remyelination.

Science.gov (United States)

Xiao, Lin; Guo, Dazhi; Hu, Chun; Shen, Weiran; Shan, Lei; Li, Cui; Liu, Xiuyun; Yang, Wenjing; Zhang, Weidong; He, Cheng

2012-07-01

Differentiation of oligodendrocyte progenitor cells (OPCs) into mature oligodendrocytes is a prerequisite for remyelination after demyelination, and impairment of this process is suggested to be a major reason for remyelination failure. Diosgenin, a plant-derived steroid, has been implicated for therapeutic use in many diseases, but little is known about its effect on the central nervous system. In this study, using a purified rat OPC culture model, we show that diosgenin significantly and specifically promotes OPC differentiation without affecting the viability, proliferation, or migration of OPC. Interestingly, the effect of diosgenin can be blocked by estrogen receptor (ER) antagonist ICI 182780 but not by glucocorticoid and progesterone receptor antagonist RU38486, nor by mineralocorticoid receptor antagonist spirolactone. Moreover, it is revealed that both ER-alpha and ER-beta are expressed in OPC, and diosgenin can activate the extracellular signal-regulated kinase 1/2 (ERK1/2) in OPC via ER. The pro-differentiation effect of diosgenin can also be obstructed by the ERK inhibitor PD98059. Furthermore, in the cuprizone-induced demyelination model, it is demonstrated that diosgenin administration significantly accelerates/enhances remyelination as detected by Luxol fast blue stain, MBP immunohistochemistry and real time RT-PCR. Diosgenin also increases the number of mature oligodendrocytes in the corpus callosum while it does not affect the number of OPCs. Taking together, our results suggest that diosgenin promotes the differentiation of OPC into mature oligodendrocyte through an ER-mediated ERK1/2 activation pathway to accelerate remyelination, which implicates a novel therapeutic usage of this steroidal natural product in demyelinating diseases such as multiple sclerosis (MS). Copyright © 2012 Wiley Periodicals, Inc.

Surface Expression of TGFβ Docking Receptor GARP Promotes Oncogenesis and Immune Tolerance in Breast Cancer.

Science.gov (United States)

Metelli, Alessandra; Wu, Bill X; Fugle, Caroline W; Rachidi, Saleh; Sun, Shaoli; Zhang, Yongliang; Wu, Jennifer; Tomlinson, Stephen; Howe, Philip H; Yang, Yi; Garrett-Mayer, Elizabeth; Liu, Bei; Li, Zihai

2016-12-15

GARP encoded by the Lrrc32 gene is the cell surface docking receptor for latent TGFβ, which is expressed naturally by platelets and regulatory T cells (Treg). Although Lrrc32 is amplified frequently in breast cancer, the expression and relevant functions of GARP in cancer have not been explored. Here, we report that GARP exerts oncogenic effects, promoting immune tolerance by enriching and activating latent TGFβ in the tumor microenvironment. We found that human breast, lung, and colon cancers expressed GARP aberrantly. In genetic studies in normal mammary gland epithelial and carcinoma cells, GARP expression increased TGFβ bioactivity and promoted malignant transformation in immunodeficient mice. In breast carcinoma-bearing mice that were immunocompetent, GARP overexpression promoted Foxp3 + Treg activity, which in turn contributed to enhancing cancer progression and metastasis. Notably, administration of a GARP-specific mAb limited metastasis in an orthotopic model of human breast cancer. Overall, these results define the oncogenic effects of the GARP-TGFβ axis in the tumor microenvironment and suggest mechanisms that might be exploited for diagnostic and therapeutic purposes. Cancer Res; 76(24); 7106-17. ©2016 AACR. ©2016 American Association for Cancer Research.

CINPA1 is an inhibitor of constitutive androstane receptor that does not activate pregnane X receptor.

Science.gov (United States)

Cherian, Milu T; Lin, Wenwei; Wu, Jing; Chen, Taosheng

2015-05-01

Constitutive androstane receptor (CAR) and pregnane X receptor (PXR) are xenobiotic sensors that enhance the detoxification and elimination of xenobiotics and endobiotics by modulating the expression of genes encoding drug-metabolizing enzymes and transporters. Elevated levels of drug-metabolizing enzymes and efflux transporters, resulting from CAR activation in various cancers, promote the elimination of chemotherapeutic agents, leading to reduced therapeutic effectiveness and acquired drug resistance. CAR inhibitors, in combination with existing chemotherapeutics, could therefore be used to attenuate multidrug resistance in cancers. Interestingly, all previously reported CAR inverse-agonists are also activators of PXR, rendering them mechanistically counterproductive in tissues where both these xenobiotic receptors are present and active. We used a directed high-throughput screening approach, followed by subsequent mechanistic studies, to identify novel, potent, and specific small-molecule CAR inhibitors that do not activate PXR. We describe here one such inhibitor, CINPA1 (CAR inhibitor not PXR activator 1), capable of reducing CAR-mediated transcription with an IC50 of ∼70 nM. CINPA1 1) is a specific xenobiotic receptor inhibitor and has no cytotoxic effects up to 30 µM; 2) inhibits CAR-mediated gene expression in primary human hepatocytes, where CAR is endogenously expressed; 3) does not alter the protein levels or subcellular localization of CAR; 4) increases corepressor and reduces coactivator interaction with the CAR ligand-binding domain in mammalian two-hybrid assays; and 5) disrupts CAR binding to the promoter regions of target genes in chromatin immunoprecipitation assays. CINPA1 could be used as a novel molecular tool for understanding CAR function. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

Interactions between the cyclic AMP receptor protein and the alpha subunit of RNA polymerase at the Escherichia coli galactose operon P1 promoter.

Science.gov (United States)

Attey, A; Belyaeva, T; Savery, N; Hoggett, J; Fujita, N; Ishihama, A; Busby, S

1994-10-25

DNAase I footprinting has been used to study open complexes between Escherichia coli RNA polymerase and the galactose operon P1 promoter, both in the absence and the presence of CRP (the cyclic AMP receptor protein, a transcription activator). From the effects of deletion of the C-terminal part of the RNA polymerase alpha subunit, we deduce that alpha binds at the upstream end of both the binary RNA polymerase-galP1 and ternary RNA polymerase-CRP-galP1 complexes. Disruption of the alpha-upstream contact suppresses open complex formation at galP1 at lower temperatures. In ternary RNA polymerase-CRP-galP1 complexes, alpha appears to make direct contact with Activating Region 1 in CRP. DNAase I footprinting has been used to detect and quantify interactions between purified alpha and CRP bound at galP1.

The C'-terminal interaction domain of the thyroid hormone receptor confers the ability of the DNA site to dictate positive or negative transcriptional activity

International Nuclear Information System (INIS)

Holloway, J.M.; Glass, C.K.; Adler, S.; Nelson, C.A.; Rosenfeld, M.G.

1990-01-01

To investigate mechanisms responsible for positive and negative transcriptional control, the authors have utilized two types of promoters that are diffferentially regulated by thyroid hormone (T 3 ) receptors. Promoters containing the palindromic T 3 response element TCAGGTCA TGACCTGA are positively regulated by the T 3 receptor after the administration of T 3 , whereas otherwise identical promoters containing the estrogen response element TCAGGTCA CTG TGACCTGA can be regulated negatively; converse effects are observed with the estrogen receptor. They describe evidence that the transcriptional inhibitory effects of the T 3 or estrogen receptors on the estrogen or T 3 response elements, respectively, are imposed by amino acid sequences in the C'-terminal region that colocalize with dimerization and hormone-binding domains and that these sequences can transfer inhibitory functions to other classes of transcription factors. Removal of the C'-terminal dimerization and hormone-binding domains of either the αT 3 or estrogen receptors permits each receptor to act constitutively to enhance transcription on both T 3 and estrogen response elements. It is, therefore, suggested that protein-protein interactions between receptor C' termini limit the subset of DNA binding sites on which transcriptional activation occurs

CDK11{sup p58} represses vitamin D receptor-mediated transcriptional activation through promoting its ubiquitin-proteasome degradation

Energy Technology Data Exchange (ETDEWEB)

Chi, Yayun; Hong, Yi; Zong, Hongliang; Wang, Yanlin; Zou, Weiying; Yang, Junwu; Kong, Xiangfei; Yun, Xiaojing [Gene Research Center, Shanghai Medical College and Institutes of Biomedical, Shanghai 200032 (China); Gu, Jianxin, E-mail: jxgu@shmu.edu.cn [Gene Research Center, Shanghai Medical College and Institutes of Biomedical, Shanghai 200032 (China)

2009-08-28

Vitamin D receptor (VDR) is a member of the nuclear receptor superfamily and regulates transcription of target genes. In this study, we identified CDK11{sup p58} as a novel protein involved in the regulation of VDR. CDK11{sup p58}, a member of the large family of p34cdc2-related kinases, is associated with cell cycle progression, tumorigenesis, and apoptotic signaling. Our study demonstrated that CDK11{sup p58} interacted with VDR and repressed VDR-dependent transcriptional activation. Furthermore, overexpression of CDK11{sup p58} decreased the stability of VDR through promoting its ubiquitin-proteasome-mediated degradation. Taken together, these results suggest that CDK11{sup p58} is involved in the negative regulation of VDR.

Estrogen and estrogen receptor alpha promotes malignancy and osteoblastic tumorigenesis in prostate cancer.

Science.gov (United States)

Mishra, Sweta; Tai, Qin; Gu, Xiang; Schmitz, James; Poullard, Ashley; Fajardo, Roberto J; Mahalingam, Devalingam; Chen, Xiaodong; Zhu, Xueqiong; Sun, Lu-Zhe

2015-12-29

The role of estrogen signaling in regulating prostate tumorigenesis is relatively underexplored. Although, an increasing body of evidence has linked estrogen receptor beta (ERß) to prostate cancer, the function of estrogen receptor alpha (ERα) in prostate cancer is not very well studied. We have discovered a novel role of ERα in the pathogenesis of prostate tumors. Here, we show that prostate cancer cells express ERα and estrogen induces oncogenic properties in prostate cancer cells through ERα. Importantly, ERα knockdown in the human prostate cancer PacMetUT1 cells as well as pharmacological inhibition of ERα with ICI 182,780 inhibited osteoblastic lesion formation and lung metastasis in vivo. Co-culture of pre-osteoblasts with cancer cells showed a significant induction of osteogenic markers in the pre-osteoblasts, which was attenuated by knockdown of ERα in cancer cells suggesting that estrogen/ERα signaling promotes crosstalk between cancer and osteoblastic progenitors to stimulate osteoblastic tumorigenesis. These results suggest that ERα expression in prostate cancer cells is essential for osteoblastic lesion formation and lung metastasis. Thus, inhibition of ERα signaling in prostate cancer cells may be a novel therapeutic strategy to inhibit the osteoblastic lesion development as well as lung metastasis in patients with advanced prostate cancer.

IGF-II Promotes Stemness of Neural Restricted Precursors

Science.gov (United States)

Ziegler, Amber N.; Schneider, Joel S.; Qin, Mei; Tyler, William A.; Pintar, John E.; Fraidenraich, Diego; Wood, Teresa L.; Levison, Steven W.

2016-01-01

Insulin-like growth factor (IGF)-I and IGF-II regulate brain development and growth through the IGF type 1 receptor (IGF-1R). Less appreciated is that IGF-II, but not IGF-I, activates a splice variant of the insulin receptor (IR) known as IR-A. We hypothesized that IGF-II exerts distinct effects from IGF-I on neural stem/progenitor cells (NSPs) via its interaction with IR-A. Immunofluorescence revealed high IGF-II in the medial region of the subventricular zone (SVZ) comprising the neural stem cell niche, with IGF-II mRNA predominant in the adjacent choroid plexus. The IGF-1R and the IR isoforms were differentially expressed with IR-A predominant in the medial SVZ, whereas the IGF-1R was more abundant laterally. Similarly, IR-A was more highly expressed by NSPs, whereas the IGF-1R was more highly expressed by lineage restricted cells. In vitro, IGF-II was more potent in promoting NSP expansion than either IGF-I or standard growth medium. Limiting dilution and differentiation assays revealed that IGF-II was superior to IGF-I in promoting stemness. In vivo, NSPs propagated in IGF-II migrated to and took up residence in periventricular niches while IGF-I-treated NSPs predominantly colonized white matter. Knockdown of IR or IGF-1R using shRNAs supported the conclusion that the IGF-1R promotes progenitor proliferation, whereas the IR is important for self-renewal. Q-PCR revealed that IGF-II increased Oct4, Sox1, and FABP7 mRNA levels in NSPs. Our data support the conclusion that IGF-II promotes the self-renewal of neural stem/progenitors via the IR. By contrast, IGF-1R functions as a mitogenic receptor to increase precursor abundance. PMID:22593020

Role of regional policies in promoting networking and innovation activity of firms

OpenAIRE

Kirsi Mukkala; Jari Ritsilä

2004-01-01

The success of firms and regions is increasingly defined by their innovation and learning capabilities. It has been emphasized in several studies that a local operational environment may have a positive impact on innovation activity of firms. From policy point of view, the relationship between firms and their local environment is an important research topic. The purpose of this paper is to explore whether there is a demand for regional policy makers in promoting innovative and networking acti...

Effect of TNFα on activities of different promoters of human apolipoprotein A-I gene

International Nuclear Information System (INIS)

Orlov, Sergey V.; Mogilenko, Denis A.; Shavva, Vladimir S.; Dizhe, Ella B.; Ignatovich, Irina A.; Perevozchikov, Andrej P.

2010-01-01

Research highlights: → TNFα stimulates the distal alternative promoter of human apoA-I gene. → TNFα acts by weakening of promoter competition within apoA-I gene (promoter switching). → MEK1/2 and nuclear receptors PPARα and LXRs take part in apoA-I promoter switching. -- Abstract: Human apolipoprotein A-I (ApoA-I) is a major structural and functional protein component of high-density lipoproteins. The expression of the apolipoprotein A-I gene (apoA-I) in hepatocytes is repressed by pro-inflammatory cytokines such as IL-1β and TNFα. Recently, two novel additional (alternative) promoters for human apoA-I gene have been identified. Nothing is known about the role of alternative promoters in TNFα-mediated downregulation of apoA-I gene. In this article we report for the first time about the different effects of TNFα on two alternative promoters of human apoA-I gene. Stimulation of HepG2 cells by TNFα leads to activation of the distal alternative apoA-I promoter and downregulation of the proximal alternative and the canonical apoA-I promoters. This effect is mediated by weakening of the promoter competition within human apoA-I 5'-regulatory region (apoA-I promoter switching) in the cells treated by TNFα. The MEK1/2-ERK1/2 cascade and nuclear receptors PPARα and LXRs are important for TNFα-mediated apoA-I promoter switching.

Thyrotropin-luteinizing hormone/chorionic gonadotropin receptor extracellular domain chimeras as probes for thyrotropin receptor function

International Nuclear Information System (INIS)

Nagayama, Yuji; Wadsworth, H.L.; Chazenbalk, G.D.; Russo, D.; Seto, Pui; Rapoport, B.

1991-01-01

To define the sites in the extracellular domain of the human thyrotropin (TSH) receptor that are involved in TSH binding and signal transduction the authors constructed chimeric thyrotropin-luteinizing hormone/chorionic gonadotropin (TSH-LH/CG) receptors. The extracellular domain of the human TSH receptor was divided into five regions that were replaced, either singly or in various combinations, with homologous regions of the rat LH/CG receptor. The chimeric receptors were stably expressed in Chinese hamster ovary cells. The data obtained suggest that the carboxyl region of the extracellular domain (amino acid residues 261-418) and particularly the middle region (residues 171-260) play a role in signal transduction. The possibility is also raised of an interaction between the amino and carboxyl regions of the extracellular domain in the process of signal transduction. In summary, these studies suggest that the middle region and carboxyl half of the extracellular domain of the TSH receptor are involved in signal transduction and that the TSH-binding region is likely to span the entire extracellular domain, with multiple discontinuous contact sites

Histone Deacetylase 7 Promotes Toll-like Receptor 4-dependent Proinflammatory Gene Expression in Macrophages*

Science.gov (United States)

Shakespear, Melanie R.; Hohenhaus, Daniel M.; Kelly, Greg M.; Kamal, Nabilah A.; Gupta, Praveer; Labzin, Larisa I.; Schroder, Kate; Garceau, Valerie; Barbero, Sheila; Iyer, Abishek; Hume, David A.; Reid, Robert C.; Irvine, Katharine M.; Fairlie, David P.; Sweet, Matthew J.

2013-01-01

Broad-spectrum inhibitors of histone deacetylases (HDACs) constrain Toll-like receptor (TLR)-inducible production of key proinflammatory mediators. Here we investigated HDAC-dependent inflammatory responses in mouse macrophages. Of the classical Hdacs, Hdac7 was expressed at elevated levels in inflammatory macrophages (thioglycollate-elicited peritoneal macrophages) as compared with bone marrow-derived macrophages and the RAW264 cell line. Overexpression of a specific, alternatively spliced isoform of Hdac7 lacking the N-terminal 22 amino acids (Hdac7-u), but not the Refseq Hdac7 (Hdac7-s), promoted LPS-inducible expression of Hdac-dependent genes (Edn1, Il-12p40, and Il-6) in RAW264 cells. A novel class IIa-selective HDAC inhibitor reduced recombinant human HDAC7 enzyme activity as well as TLR-induced production of inflammatory mediators in thioglycollate-elicited peritoneal macrophages. Both LPS and Hdac7-u up-regulated the activity of the Edn1 promoter in an HDAC-dependent fashion in RAW264 cells. A hypoxia-inducible factor (HIF) 1 binding site in this promoter was required for HDAC-dependent TLR-inducible promoter activity and for Hdac7- and HIF-1α-mediated trans-activation. Coimmunoprecipitation assays showed that both Hdac7-u and Hdac7-s interacted with HIF-1α, whereas only Hdac7-s interacted with the transcriptional repressor CtBP1. Thus, Hdac7-u positively regulates HIF-1α-dependent TLR signaling in macrophages, whereas an interaction with CtBP1 likely prevents Hdac7-s from exerting this effect. Hdac7 may represent a potential inflammatory disease target. PMID:23853092

Histone deacetylase 7 promotes Toll-like receptor 4-dependent proinflammatory gene expression in macrophages.

Science.gov (United States)

Shakespear, Melanie R; Hohenhaus, Daniel M; Kelly, Greg M; Kamal, Nabilah A; Gupta, Praveer; Labzin, Larisa I; Schroder, Kate; Garceau, Valerie; Barbero, Sheila; Iyer, Abishek; Hume, David A; Reid, Robert C; Irvine, Katharine M; Fairlie, David P; Sweet, Matthew J

2013-08-30

Broad-spectrum inhibitors of histone deacetylases (HDACs) constrain Toll-like receptor (TLR)-inducible production of key proinflammatory mediators. Here we investigated HDAC-dependent inflammatory responses in mouse macrophages. Of the classical Hdacs, Hdac7 was expressed at elevated levels in inflammatory macrophages (thioglycollate-elicited peritoneal macrophages) as compared with bone marrow-derived macrophages and the RAW264 cell line. Overexpression of a specific, alternatively spliced isoform of Hdac7 lacking the N-terminal 22 amino acids (Hdac7-u), but not the Refseq Hdac7 (Hdac7-s), promoted LPS-inducible expression of Hdac-dependent genes (Edn1, Il-12p40, and Il-6) in RAW264 cells. A novel class IIa-selective HDAC inhibitor reduced recombinant human HDAC7 enzyme activity as well as TLR-induced production of inflammatory mediators in thioglycollate-elicited peritoneal macrophages. Both LPS and Hdac7-u up-regulated the activity of the Edn1 promoter in an HDAC-dependent fashion in RAW264 cells. A hypoxia-inducible factor (HIF) 1 binding site in this promoter was required for HDAC-dependent TLR-inducible promoter activity and for Hdac7- and HIF-1α-mediated trans-activation. Coimmunoprecipitation assays showed that both Hdac7-u and Hdac7-s interacted with HIF-1α, whereas only Hdac7-s interacted with the transcriptional repressor CtBP1. Thus, Hdac7-u positively regulates HIF-1α-dependent TLR signaling in macrophages, whereas an interaction with CtBP1 likely prevents Hdac7-s from exerting this effect. Hdac7 may represent a potential inflammatory disease target.

Promoting regional mobility

DEFF Research Database (Denmark)

Jensen, Anne

Pricing of transport has been part of EU's common transport policy since this gained momentum in the early 1990s. Since then, it has been closely connected to the trans-European transport network (TEN-T) and to rising demands of efficient mobility systems at a local, regional and Community scale....... Development of pricing policies is contested at Community level and has taken place in a clash between different policy rationalities. Significantly though, the effects of the pricing policies are closely related to regional mobility systems, e.g. through financing large trans-border infrastructure projects...... and establishing common technical charging systems thus changing the conditions for regional mobility. This paper explores how policies of infrastructure pricing shape new ways of governing mobility which influences trans-border, regional policy-making. The key findings are that there is a tendency to include...

Defining structural and functional dimensions of the extracellular thyrotropin receptor region.

Science.gov (United States)

Kleinau, Gunnar; Mueller, Sandra; Jaeschke, Holger; Grzesik, Paul; Neumann, Susanne; Diehl, Anne; Paschke, Ralf; Krause, Gerd

2011-06-24

The extracellular region of the thyrotropin receptor (TSHR) can be subdivided into the leucine-rich repeat domain (LRRD) and the hinge region. Both the LRRD and the hinge region interact with thyrotropin (TSH) or autoantibodies. Structural data for the TSHR LRRD were previously determined by crystallization (amino acids Glu(30)-Thr(257), 10 repeats), but the structure of the hinge region is still undefined. Of note, the amino acid sequence (Trp(258)-Tyr(279)) following the crystallized LRRD comprises a pattern typical for leucine-rich repeats with conserved hydrophobic side chains stabilizing the repeat fold. Moreover, functional data for amino acids between the LRRD and the transmembrane domain were fragmentary. We therefore investigated systematically these TSHR regions by mutagenesis to reveal insights into their functional contribution and potential structural features. We found that mutations of conserved hydrophobic residues between Thr(257) and Tyr(279) cause TSHR misfold, which supports a structural fold of this peptide, probably as an additional leucine-rich repeat. Furthermore, we identified several new mutations of hydrophilic amino acids in the entire hinge region leading to partial TSHR inactivation, indicating that these positions are important for intramolecular signal transduction. In summary, we provide new information regarding the structural features and functionalities of extracellular TSHR regions. Based on these insights and in context with previous results, we suggest an extracellular activation mechanism that supports an intramolecular agonistic unit as a central switch for activating effects at the extracellular region toward the serpentine domain.

Functional effects of polymorphisms on glucocorticoid receptor modulation of human anxiogenic substance-P gene promoter activity in primary amygdala neurones.

Science.gov (United States)

Hay, Colin W; Shanley, Lynne; Davidson, Scott; Cowie, Philip; Lear, Marissa; McGuffin, Peter; Riedel, Gernot; McEwan, Iain J; MacKenzie, Alasdair

2014-09-01

Expression or introduction of the neuropeptide substance-P (SP; encoded by the TAC1 gene in humans and Tac1 in rodents) in the amygdala induces anxiety related behaviour in rodents. In addition, pharmacological antagonism of the main receptor of SP in humans; NK1, is anxiolytic. In the current study, we show that the Tac1 locus is up-regulated in primary rat amygdala neurones in response to activation of the glucocorticoid receptor (GR); a classic component of the stress response. Using a combination of bioinformatics, electrophoretic mobility shift assays (EMSA) and reporter plasmid magnetofection into rat primary amygdala neurones we identified a highly conserved GR response sequence (2GR) in the human TAC1 promoter that binds GR in response to dexamethasone (Dex) or forskolin. We also identified a second GR binding site in the human promoter that was polymorphic and whose T-allele is only found in Japanese and Chinese populations. We present evidence that the T-allele of SNPGR increases the activity of the TAC1 promoter through de-sequestration or de-repression of 2GR. The identification of Dex/forskolin response elements in the TAC1 promoter in amygdala neurones suggests a possible link in the chain of molecular events connecting GR activation and anxiety. In addition, the discovery of a SNP which can alter this response may have implications for our understanding of the role of regulatory variation in susceptibility to stress in specific populations. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

The G-protein coupled chemoattractant receptor FPR2 promotes malignant phenotype of human colon cancer cells

Science.gov (United States)

Xiang, Yi; Yao, Xiaohong; Chen, Keqiang; Wang, Xiafei; Zhou, Jiamin; Gong, Wanghua; Yoshimura, Teizo; Huang, Jiaqiang; Wang, Rongquan; Wu, Yuzhang; Shi, Guochao; Bian, Xiuwu; Wang, Jiming

2016-01-01

The G-protein coupled chemoattractant receptor formylpeptide receptor-2 (FPR2 in human, Fpr2 in mice) is expressed by mouse colon epithelial cells and plays a critical role in mediating mucosal homeostasis and inflammatory responses. However, the biological role of FPR2 in human colon is unclear. Our investigation revealed that a considerable number of human colon cancer cell lines expressed FPR2 and its ligands promoted cell migration and proliferation. Human colon cancer cell lines expressing high levels of FPR2 also formed more rapidly growing tumors in immunocompromised mice as compared with cell lines expressing lower levels of FPR2. Knocking down of FPR2 from colon cancer cell lines highly expressing FPR2 reduced their tumorigenicity. Clinically, FPR2 is more highly expressed in progressive colon cancer, associated with poorer patient prognosis. These results suggest that FPR2 can be high-jacked by colon cancer cells for their growth advantage, thus becoming a potential target for therapeutic development. PMID:27904774

Estrogen regulation of chicken riboflavin carrier protein gene is mediated by ERE half sites without direct binding of estrogen receptor.

Science.gov (United States)

Bahadur, Urvashi; Ganjam, Goutham K; Vasudevan, Nandini; Kondaiah, Paturu

2005-02-28

Estrogen is an important steroid hormone that mediates most of its effects on regulation of gene expression by binding to intracellular receptors. The consensus estrogen response element (ERE) is a 13bp palindromic inverted repeat with a three nucleotide spacer. However, several reports suggest that many estrogen target genes are regulated by diverse elements, such as imperfect EREs and ERE half sites (ERE 1/2), which are either the proximal or the distal half of the palindrome. To gain more insight into ERE half site-mediated gene regulation, we used a region from the estrogen-regulated chicken riboflavin carrier protein (RCP) gene promoter that contains ERE half sites. Using moxestrol, an analogue of estrogen and transient transfection of deletion and mutation containing RCP promoter/reporter constructs in chicken hepatoma (LMH2A) cells, we identified an estrogen response unit (ERU) composed of two consensus ERE 1/2 sites and one non-consensus ERE 1/2 site. Mutation of any of these sites within this ERU abolishes moxestrol response. Further, the ERU is able to confer moxestrol responsiveness to a heterologous promoter. Interestingly, RCP promoter is regulated by moxestrol in estrogen responsive human MCF-7 cells, but not in other cell lines such as NIH3T3 and HepG2 despite estrogen receptor-alpha (ER-alpha) co transfection. Electrophoretic mobility shift assays (EMSAs) with promoter regions encompassing the half sites and nuclear extracts from LMH2A cells show the presence of a moxestrol-induced complex that is abolished by a polyclonal anti-ERalpha antibody. Surprisingly, estrogen receptor cannot bind to these promoter elements in isolation. Thus, there appears to be a definite requirement for some other factor(s) in addition to estrogen receptor, for the generation of a suitable response of this promoter to estrogen. Our studies therefore suggest a novel mechanism of gene regulation by estrogen, involving ERE half sites without direct binding of ER to the

Region-specific proteolysis differentially regulates type 1 inositol 1,4,5-trisphosphate receptor activity.

Science.gov (United States)

Wang, Liwei; Wagner, Larry E; Alzayady, Kamil J; Yule, David I

2017-07-14

The inositol 1,4,5 trisphosphate receptor (IP 3 R) is an intracellular Ca 2+ release channel expressed predominately on the membranes of the endoplasmic reticulum. IP 3 R1 can be cleaved by caspase or calpain into at least two receptor fragments. However, the functional consequences of receptor fragmentation are poorly understood. Our previous work has demonstrated that IP 3 R1 channels, formed following either enzymatic fragmentation or expression of the corresponding complementary polypeptide chains, retain tetrameric architecture and are still activated by IP 3 binding despite the loss of peptide continuity. In this study, we demonstrate that region-specific receptor fragmentation modifies channel regulation. Specifically, the agonist-evoked temporal Ca 2+ release profile and protein kinase A modulation of Ca 2+ release are markedly altered. Moreover, we also demonstrate that activation of fragmented IP 3 R1 can result in a distinct functional outcome. Our work suggests that proteolysis of IP 3 R1 may represent a novel form of modulation of IP 3 R1 channel function and increases the repertoire of Ca 2+ signals achievable through this channel. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Nucleopolis for promoting the nuclear excellence of the Normandy region

International Nuclear Information System (INIS)

2016-01-01

Nucleopolis is the Norman economic pole dedicated to nuclear energy, nuclear medicine and nuclear safety, it gathers about 70 enterprises whatever their sizes, research laboratories and teaching or training units. Nucleopolis was founded in 2009 with the economic development of the region as a unique purpose. Nucleopolis will ease the access of its members to local, national and international markets through actions of networking and by promoting innovations and skill development. Nucleopolis proposes to its members a series of services around 4 departments: Nucleo'Network to promote networking between the members themselves and between the members and major contractors; Nucleo'Business to propose assistance in national and international business; Nucleo'Competence to propose adequate training to its members to upgrade their skills and Nucleo'Innovation to foster collaborative work between its members on innovative projects. (A.C.)

Restoration of CpG Methylation in The Egf Promoter Region during Rat Liver Regeneration

Science.gov (United States)

Deming, Li; Ziwei, Li; Xueqiang, Guo; Cunshuan, Xu

2015-01-01

Epidermal growth factor (EGF) is an important factor for healing after tissue damage in diverse experimental models. It plays an important role in liver regeneration (LR). The objective of this experiment is to investigate the methylation variation of 10 CpG sites in the Egf promoter region and their relevance to Egf expression during rat liver regenera- tion. As a follow up of our previous study, rat liver tissue was collected after rat 2/3 partial hepatectomy (PH) during the re-organization phase (from days 14 to days 28). Liver DNA was extracted and modified by sodium bisulfate. The methylation status of 10 CpG sites in Egf promoter region was determined using bisulfite sequencing polymerase chain reaction (PCR), as BSP method. The results showed that 3 (sites 3, 4 and 9) out of 10 CpG sites have strikingly methylation changes during the re-organization phase compared to the regeneration phase (from 2 hours to 168 hours, P=0.002, 0.048 and 0.018, respectively). Our results showed that methylation modification of CpGs in the Egf promoter region could be restored to the status before PH operation and changes of methylation didn’t affect Egf mRNA expression during the re-organization phase. PMID:26464832

The axonal guidance receptor neogenin promotes acute inflammation.

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Klemens König

Full Text Available Neuronal guidance proteins (NGP were originally described in the context of axonal growth and migration. Yet recent work has demonstrated that NGPs also serve as guidance cues for immune competent cells. A crucial target receptor for NGPs during embryonic development is the neogenin receptor, however its role during acute inflammation is unknown. We report here that neogenin is abundantly expressed outside the nervous system and that animals with endogenous repression of neogenin (Neo1(-/- demonstrate attenuated changes of acute inflammation. Studies using functional inhibition of neogenin resulted in a significant attenuation of inflammatory peritonitis. In studies employing bone marrow chimeric animals we found the hematopoietic presence of Neo1(-/- to be responsible for the attenuated inflammatory response. Taken together our studies suggest that the guidance receptor neogenin holds crucial importance for the propagation of an acute inflammatory response and further define mechanisms shared between the nervous and the immune system.

Promoted neuronal differentiation after activation of alpha4/beta2 nicotinic acetylcholine receptors in undifferentiated neural progenitors.

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Takeshi Takarada

Full Text Available BACKGROUND: Neural progenitor is a generic term used for undifferentiated cell populations of neural stem, neuronal progenitor and glial progenitor cells with abilities for proliferation and differentiation. We have shown functional expression of ionotropic N-methyl-D-aspartate (NMDA and gamma-aminobutyrate type-A receptors endowed to positively and negatively regulate subsequent neuronal differentiation in undifferentiated neural progenitors, respectively. In this study, we attempted to evaluate the possible functional expression of nicotinic acetylcholine receptor (nAChR by undifferentiated neural progenitors prepared from neocortex of embryonic rodent brains. METHODOLOGY/PRINCIPAL FINDINGS: Reverse transcription polymerase chain reaction analysis revealed mRNA expression of particular nAChR subunits in undifferentiated rat and mouse progenitors prepared before and after the culture with epidermal growth factor under floating conditions. Sustained exposure to nicotine significantly inhibited the formation of neurospheres composed of clustered proliferating cells and 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide reduction activity at a concentration range of 1 µM to 1 mM without affecting cell survival. In these rodent progenitors previously exposed to nicotine, marked promotion was invariably seen for subsequent differentiation into cells immunoreactive for a neuronal marker protein following the culture of dispersed cells under adherent conditions. Both effects of nicotine were significantly prevented by the heteromeric α4β2 nAChR subtype antagonists dihydro-β-erythroidine and 4-(5-ethoxy-3-pyridinyl-N-methyl-(3E-3-buten-1-amine, but not by the homomeric α7 nAChR subtype antagonist methyllycaconitine, in murine progenitors. Sustained exposure to nicotine preferentially increased the expression of Math1 among different basic helix-loop-helix proneural genes examined. In undifferentiated progenitors from embryonic mice

Nuclear factor ETF specifically stimulates transcription from promoters without a TATA box.

Science.gov (United States)

Kageyama, R; Merlino, G T; Pastan, I

1989-09-15

Transcription factor ETF stimulates the expression of the epidermal growth factor receptor (EGFR) gene which does not have a TATA box in the promoter region. Here, we show that ETF recognizes various GC-rich sequences including stretches of deoxycytidine or deoxyguanosine residues and GC boxes with similar affinities. ETF also binds to TATA boxes but with a lower affinity. ETF stimulated in vitro transcription from several promoters without TATA boxes but had little or no effect on TATA box-containing promoters even though they had strong ETF-binding sites. These inactive ETF-binding sites became functional when placed upstream of the EGFR promoter whose own ETF-binding sites were removed. Furthermore, when a TATA box was introduced into the EGFR promoter, the responsiveness to ETF was abolished. These results indicate that ETF is a specific transcription factor for promoters which do not contain TATA elements.

Surface Expression of TGF-β Docking Receptor GARP Promotes Oncogenesis and Immune Tolerance in Breast Cancer

Science.gov (United States)

Metelli, Alessandra; Wu, Bill X; Fugle, Caroline W; Rachidi, Saleh; Sun, Shaoli; Zhang, Yongliang; Wu, Jennifer; Tomlinson, Stephen; Howe, Philip; Yang, Yi; Garrett-Mayer, Elizabeth; Liu, Bei; Li, Zihai

2016-01-01

GARP encoded by the Lrrc32 gene is the cell surface docking receptor for latent TGF-β which is expressed naturally by platelets and regulatory T cells. Although Lrrc32 is amplified frequently in breast cancer, the expression and relevant functions of GARP in cancer have not been explored. Here we report that GARP exerts oncogenic effects, promoting immune tolerance by enriching and activating latent TGF-β in the tumor microenvironment. We found that human breast, lung and colon cancers expressed GARP aberrantly. In genetic studies in normal mammary gland epithelial and carcinoma cells, GARP expression increased TGF-β bioactivity and promoted malignant transformation in immune deficient mice. In breast carcinoma-bearing mice that were immune competent, GARP overexpression promoted Foxp3+ regulatory T cell activity, which in turn contributed to enhancing cancer progression and metastasis. Notably, administration of a panel of GARP-specific monoclonal antibodies limited metastasis in an orthotopic model of human breast cancer. Overall, these results define the oncogenic effects of the GARP-TGF-β axis in the tumor microenvironment and suggest mechanisms that might be exploited for diagnostic and therapeutic purposes. PMID:27913437

Ritonavir binds to and downregulates estrogen receptors: Molecular mechanism of promoting early atherosclerosis

Energy Technology Data Exchange (ETDEWEB)

Xiang, Jin [Ministry of Education Laboratory of Combinatorial Biosynthesis and Drug Discovery, School of Pharmaceutical Science, Wuhan University, Wuhan 430071 (China); Wang, Ying [Department of Pathophysiology, School of Medicine, Wuhan University, Wuhan 430071 (China); Su, Ke [Department of Nephrology, Renmin Hospital of Wuhan University, Wuhan 430060 (China); Liu, Min [Ministry of Education Laboratory of Combinatorial Biosynthesis and Drug Discovery, School of Pharmaceutical Science, Wuhan University, Wuhan 430071 (China); Hu, Peng-Chao [Department of Pathophysiology, School of Medicine, Wuhan University, Wuhan 430071 (China); Ma, Tian; Li, Jia-Xi [Ministry of Education Laboratory of Combinatorial Biosynthesis and Drug Discovery, School of Pharmaceutical Science, Wuhan University, Wuhan 430071 (China); Wei, Lei [Department of Pathophysiology, School of Medicine, Wuhan University, Wuhan 430071 (China); Zheng, Zhongliang, E-mail: biochem@whu.edu.cn [State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072 (China); Yang, Fang, E-mail: fang-yang@whu.edu.cn [Department of Physiology, School of Medicine, Wuhan University, Wuhan 430071 (China)

2014-10-01

Estrogenic actions are closely related to cardiovascular disease. Ritonavir (RTV), a human immunodeficiency virus (HIV) protease inhibitor, induces atherosclerosis in an estrogen-related manner. However, how RTV induce pathological phenotypes through estrogen pathway remains unclear. In this study, we found that RTV increases thickness of coronary artery walls of Sprague Dawley rats and plasma free fatty acids (FFA) levels. In addition, RTV could induce foam cell formation, downregulate both estrogen receptor α (ERα) and ERβ expression, upregulate G protein-coupled estrogen receptor (GPER) expression, and all of them could be partially blocked by 17β-estradiol (E2), suggesting RTV acts as an antagonist for E2. Computational modeling shows a similar interaction with ERα between RTV and 2-aryl indoles, which are highly subtype-selective ligands for ERα. We also found that RTV directly bound to ERα and selectively inhibited the nuclear localization of ERα, and residue Leu536 in the hydrophobic core of ligand binding domain (LBD) was essential for the interaction with RTV. In addition, RTV did not change the secondary structure of ERα-LBD like E2, which explained how ERα lost the capacity of nuclear translocation under the treatment of RTV. All of the evidences suggest that ritonavir acts as an antagonist for 17β-estradiol in regulating α subtype estrogen receptor function and early events of atherosclerosis. - Graphical abstract: RTV directly binds to ERα and Leu536 in the hydrophobic core of ligand binding domain is essential for the interaction. - Highlights: • RTV increases the thickness of rat coronary artery wall and foam cell formation. • RTV downregulates the expression of ERα and ERβ. • RTV inhibits ERα promoter activity. • RTV directly binds to ERα and the key amino acid is Leu536. • RTV inhibits the nuclear translocation of ERα and GPER.

Ritonavir binds to and downregulates estrogen receptors: Molecular mechanism of promoting early atherosclerosis

International Nuclear Information System (INIS)

Xiang, Jin; Wang, Ying; Su, Ke; Liu, Min; Hu, Peng-Chao; Ma, Tian; Li, Jia-Xi; Wei, Lei; Zheng, Zhongliang; Yang, Fang

2014-01-01

Estrogenic actions are closely related to cardiovascular disease. Ritonavir (RTV), a human immunodeficiency virus (HIV) protease inhibitor, induces atherosclerosis in an estrogen-related manner. However, how RTV induce pathological phenotypes through estrogen pathway remains unclear. In this study, we found that RTV increases thickness of coronary artery walls of Sprague Dawley rats and plasma free fatty acids (FFA) levels. In addition, RTV could induce foam cell formation, downregulate both estrogen receptor α (ERα) and ERβ expression, upregulate G protein-coupled estrogen receptor (GPER) expression, and all of them could be partially blocked by 17β-estradiol (E2), suggesting RTV acts as an antagonist for E2. Computational modeling shows a similar interaction with ERα between RTV and 2-aryl indoles, which are highly subtype-selective ligands for ERα. We also found that RTV directly bound to ERα and selectively inhibited the nuclear localization of ERα, and residue Leu536 in the hydrophobic core of ligand binding domain (LBD) was essential for the interaction with RTV. In addition, RTV did not change the secondary structure of ERα-LBD like E2, which explained how ERα lost the capacity of nuclear translocation under the treatment of RTV. All of the evidences suggest that ritonavir acts as an antagonist for 17β-estradiol in regulating α subtype estrogen receptor function and early events of atherosclerosis. - Graphical abstract: RTV directly binds to ERα and Leu536 in the hydrophobic core of ligand binding domain is essential for the interaction. - Highlights: • RTV increases the thickness of rat coronary artery wall and foam cell formation. • RTV downregulates the expression of ERα and ERβ. • RTV inhibits ERα promoter activity. • RTV directly binds to ERα and the key amino acid is Leu536. • RTV inhibits the nuclear translocation of ERα and GPER

Identification and characterization of a liver stage-specific promoter region of the malaria parasite Plasmodium.

Directory of Open Access Journals (Sweden)

Susanne Helm

Full Text Available During the blood meal of a Plasmodium-infected mosquito, 10 to 100 parasites are inoculated into the skin and a proportion of these migrate via the bloodstream to the liver where they infect hepatocytes. The Plasmodium liver stage, despite its clinical silence, represents a highly promising target for antimalarial drug and vaccine approaches. Successfully invaded parasites undergo a massive proliferation in hepatocytes, producing thousands of merozoites that are transported into a blood vessel to infect red blood cells. To successfully develop from the liver stage into infective merozoites, a tight regulation of gene expression is needed. Although this is a very interesting aspect in the biology of Plasmodium, little is known about gene regulation in Plasmodium parasites in general and in the liver stage in particular. We have functionally analyzed a novel promoter region of the rodent parasite Plasmodium berghei that is exclusively active during the liver stage of the parasite. To prove stage-specific activity of the promoter, GFP and luciferase reporter assays have been successfully established, allowing both qualitative and accurate quantitative analysis. To further characterize the promoter region, the transcription start site was mapped by rapid amplification of cDNA ends (5'-RACE. Using promoter truncation experiments and site-directed mutagenesis within potential transcription factor binding sites, we suggest that the minimal promoter contains more than one binding site for the recently identified parasite-specific ApiAP2 transcription factors. The identification of a liver stage-specific promoter in P. berghei confirms that the parasite is able to tightly regulate gene expression during its life cycle. The identified promoter region might now be used to study the biology of the Plasmodium liver stage, which has thus far proven problematic on a molecular level. Stage-specific expression of dominant-negative mutant proteins and

DESENSITIZATION PROPERTIES OF P2X3 RECEPTORS SHAPING PAIN SIGNALLING

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Rashid eGiniatullin

2013-12-01

Full Text Available ATP-gated P2X3 receptors are mostly expressed by nociceptive sensory neurons and participate in transduction of pain signals. P2X3 receptors show a combination of fast desensitization onset and slow recovery. Moreover, even low nanomolar agonist concentrations unable to evoke a response, can induce desensitization via a phenomenon called ‘high affinity desensitization’. We have also observed that recovery from desensitization is agonist-specific and can range from seconds to minutes. The recovery process displays unusually high temperature dependence. Likewise, recycling of P2X3 receptors in peri-membrane regions shows unexpectedly large temperature sensitivity. By applying kinetic modeling, we have previously shown that desensitization characteristics of P2X3 receptor are best explained with a cyclic model of receptor operation involving three agonist molecules binding a single receptor and that desensitization is primarily developing from the open receptor state. Mutagenesis experiments suggested that desensitization depends on a certain conformation of the ATP binding pocket and on the structure of the transmembrane domains forming the ion pore. Further molecular determinants of desensitization have been identified by mutating the intracellular N- and C-termini of P2X3 receptor. Unlike other P2X receptors, the P2X3 subtype is facilitated by extracellular calcium that acts via specific sites in the ectodomain neighboring the ATP binding pocket. Thus, substitution of serine275 in this region (called ‘left flipper’ converts the natural facilitation induced by extracellular calcium to receptor inhibition. Given such their strategic location in nociceptive neurons and unique desensitization properties, P2X3 receptors represent an attractive target for development of new analgesic drugs via promotion of desensitization aimed at suppressing chronic pain.

Absence of mutation at the 5'-upstream promoter region of the TPM4 gene from cardiac mutant axolotl (Ambystoma mexicanum).

Science.gov (United States)

Denz, Christopher R; Zhang, Chi; Jia, Pingping; Du, Jianfeng; Huang, Xupei; Dube, Syamalima; Thomas, Anish; Poiesz, Bernard J; Dube, Dipak K

2011-09-01

Tropomyosins are a family of actin-binding proteins that show cell-specific diversity by a combination of multiple genes and alternative RNA splicing. Of the 4 different tropomyosin genes, TPM4 plays a pivotal role in myofibrillogenesis as well as cardiac contractility in amphibians. In this study, we amplified and sequenced the upstream regulatory region of the TPM4 gene from both normal and mutant axolotl hearts. To identify the cis-elements that are essential for the expression of the TPM4, we created various deletion mutants of the TPM4 promoter DNA, inserted the deleted segments into PGL3 vector, and performed promoter-reporter assay using luciferase as the reporter gene. Comparison of sequences of the promoter region of the TPM4 gene from normal and mutant axolotl revealed no mutations in the promoter sequence of the mutant TPM4 gene. CArG box elements that are generally involved in controlling the expression of several other muscle-specific gene promoters were not found in the upstream regulatory region of the TPM4 gene. In deletion experiments, loss of activity of the reporter gene was noted upon deletion which was then restored upon further deletion suggesting the presence of both positive and negative cis-elements in the upstream regulatory region of the TPM4 gene. We believe that this is the first axolotl promoter that has ever been cloned and studied with clear evidence that it functions in mammalian cell lines. Although striated muscle-specific cis-acting elements are absent from the promoter region of TPM4 gene, our results suggest the presence of positive and negative cis-elements in the promoter region, which in conjunction with positive and negative trans-elements may be involved in regulating the expression of TPM4 gene in a tissue-specific manner.

Governmental promotion of the Information Society in the Spanish Region of Valencia

Directory of Open Access Journals (Sweden)

Emilio Feliu-García, Ph.D.

2011-01-01

Full Text Available Regional spheres are considered essential in the governmental promotion of the Information Society at the international level. The regional initiatives in Spain aim to strengthen and complement the initiatives promoted at the national level. This article analyses ICT penetration in the Valencian Community from 1996 to 2008. The objective is to identify which of the actions carried out by the Valencian Regional Government have had a positive effect on its society.The methodology employed in this study is benchmarking. The selection of indicators is based on the policies evaluation model proposed in the Plan Avanza (Spain’s national Information Society strategy. Data were collected from official statistical sources (like Spain’s National Statistics Institute, INE. Three statistical tests were applied to verify the hypotheses (Pearson’s r2, Chi-square and Student’s t.The results indicate that it is not possible to affirm that the actions implemented by the Valencian Regional Government have had a more positive effect on its society than those implemented by the Spanish Central Government. A reason for this may lie in the specific objectives of the political strategy implemented by the Valencian Government, which has focused primarily on e-Government and does not include enough projects centred on the implementation of new technologies in the private sector. Moreover, the integration of new technologies in everyday life is placed in a second level of importance despite citizens are central actors in the international agenda.

Urinary retinoic acid receptor-β2 gene promoter methylation and hyaluronidase activity as noninvasive tests for diagnosis of bladder cancer.

Science.gov (United States)

Eissa, Sanaa; Zohny, Samir F; Shehata, Hanan Hussien; Hegazy, Marwa G A; Salem, Ahmed M; Esmat, Mohamed

2012-04-01

We evaluated the significance of urinary retinoic acid receptor-β2 (RAR-β2) gene promoter methylation and hyaluronidase activity in comparison with voided urine cytology (VUC) in diagnosis of bladder cancer. This study included 100 patients diagnosed with bladder cancer, 65 patients with benign urological disorders and 51 healthy volunteers. Urine supernatant was used for determining hyaluronidase activity by zymography while urine sediment was used for cytology and detection of methylated RAR-β2 gene promoter by methylation specific nested PCR. The sensitivity and specificity were 53% and 90.5% for VUC, 65% and 89.7% for percent methylation fraction of RAR-β2 gene promoter, and 89% and 90.5% for hyaluronidase activity; combination of the three parameters increased sensitivity to 95%. A significant association was observed between investigated markers and advanced grade tumor. Combined use of RAR-β2 gene promoter methylation, hyaluronidase activity and VUC is promising non-invasive tool for bladder cancer detection. Copyright © 2012. Published by Elsevier Inc.

Orphan nuclear receptor Nur77 participates in human apolipoprotein A5 gene expression

International Nuclear Information System (INIS)

Song, Kwang-Hoon

2010-01-01

The orphan nuclear receptor Nur77 (NR4A1) has been reported to play a crucial role in the modulation of diverse metabolic processes in liver. Here, we reported the identification of human apolipoprotein A5 (ApoA5), which implicated in lowering plasma triglyceride levels, as a novel target gene of Nur77. Nur77 induced the human ApoA5 promoter activity. Using 5'-deletion and mutagenesis of human ApoA5 promoter analysis and chromatin immunoprecipitation assays, it was shown that Nur77 directly regulated human ApoA5 gene expression by binding to a Nur77 response element (AAAGGTCA) located in the proximal human ApoA5 promoter region. In addition, we demonstrated that blocking of Nur77 transcriptional activity via overexpression of dominant negative Nur77 suppressed human ApoA5 promoter activity and mRNA expression in human hepatoma cells, HepG2. Taken together, our results demonstrated that Nur77 is a novel regulator of human ApoA5 gene expression and provide a new insight into the role of this orphan nuclear receptor in lipoprotein metabolism and triglyceride homeostasis.

Orphan nuclear receptor Nur77 participates in human apolipoprotein A5 gene expression

Energy Technology Data Exchange (ETDEWEB)

Song, Kwang-Hoon, E-mail: ksong@kiom.re.kr [Korea Institute of Oriental Medicine, Daejeon 305-811 (Korea, Republic of)

2010-01-29

The orphan nuclear receptor Nur77 (NR4A1) has been reported to play a crucial role in the modulation of diverse metabolic processes in liver. Here, we reported the identification of human apolipoprotein A5 (ApoA5), which implicated in lowering plasma triglyceride levels, as a novel target gene of Nur77. Nur77 induced the human ApoA5 promoter activity. Using 5'-deletion and mutagenesis of human ApoA5 promoter analysis and chromatin immunoprecipitation assays, it was shown that Nur77 directly regulated human ApoA5 gene expression by binding to a Nur77 response element (AAAGGTCA) located in the proximal human ApoA5 promoter region. In addition, we demonstrated that blocking of Nur77 transcriptional activity via overexpression of dominant negative Nur77 suppressed human ApoA5 promoter activity and mRNA expression in human hepatoma cells, HepG2. Taken together, our results demonstrated that Nur77 is a novel regulator of human ApoA5 gene expression and provide a new insight into the role of this orphan nuclear receptor in lipoprotein metabolism and triglyceride homeostasis.

Activation of type 2 cannabinoid receptors (CB2R) promotes fatty acid oxidation through the SIRT1/PGC-1α pathway

Energy Technology Data Exchange (ETDEWEB)

Zheng, Xuqin [Department of Endocrinology, First Affiliated Hospital, Nanjing Medical University, Nanjing, Jiangsu Province 210029 (China); Sun, Tao [Department of Neurology, Jinling Hospital, Nanjing University School of Medicine, Nanjing, Jiangsu Province 210002 (China); Wang, Xiaodong, E-mail: xdwang666@hotmail.com [Department of Endocrinology, First Affiliated Hospital, Nanjing Medical University, Nanjing, Jiangsu Province 210029 (China)

2013-07-05

Highlights: •TC, a CB2R specific agonist, stimulates SIRT1 activity by PKA/CREB pathway. •TC promotes PGC-1α transcriptional activity by increasing its deacetylation. •TC increases the expression of genes linked to FAO and promotes the rate of FAO. •The effects of TC in FAO are dependent on CB2R. •Suggesting CB2R as a target to treat diseases with lipid dysregulation. -- Abstract: Abnormal fatty acid oxidation has been associated with obesity and type 2 diabetes. At the transcriptional level, peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α) has been reported to strongly increase the ability of hormone nuclear receptors PPARα and ERRα to drive transcription of fatty acid oxidation enzymes. In this study, we report that a specific agonist of the type 2 cannabinoid receptor (CB2R) can lead to fatty acid oxidation through the PGC-1α pathway. We have found that CB2R is expressed in differentiated C2C12 myotubes, and that use of the specific agonist trans-caryophyllene (TC) stimulates sirtuin 1 (SIRT1) deacetylase activity by increasing the phosphorylation of cAMP response element-binding protein (CREB), thus leading to increased levels of PGC-1α deacetylation. This use of TC treatment increases the expression of genes linked to the fatty acid oxidation pathway in a SIRT1/PGC-1α-dependent mechanism and also drastically accelerates the rate of complete fatty acid oxidation in C2C12 myotubes, neither of which occur when CB2R mRNA is knocked down using siRNA. These results reveal that activation of CB2R by a selective agonist promotes lipid oxidation through a signaling/transcriptional pathway. Our findings imply that pharmacological manipulation of CB2R may provide therapeutic possibilities to treat metabolic diseases associated with lipid dysregulation.

Selective Androgen Receptor Modulators (SARMs) as Function Promoting Therapies

Science.gov (United States)

Bhasin, Shalender; Jasuja, Ravi

2010-01-01

Purpose of review The last decade has witnessed unprecedented discovery effort to develop selective androgen receptor modulators (SARMs) that improve physical function and bone health without adversely affecting the prostate and cardiovascular outcomes. This review describes the historical evolution, the rationale for SARM development, and the mechanisms of testosterone action and SARM selectivity. Recent Findings While steroidal SARMs have been around since the 1940s, a number of nonsteroidal SARMs that do not serve as substrates for CYP19 aromatase or 5α-reductase, act as full agonists in muscle and bone and as partial agonists in prostate are in development. The differing interactions of steroidal and nonsteroidal compounds with AR contribute to their unique pharmacologic actions. Ligand binding induces specific conformational changes in the ligand binding domain, which could modulate surface topology and protein-protein interactions between AR and coregulators, resulting in tissue-specific gene regulation. Preclinical studies have demonstrated the ability of SARMs to increase muscle and bone mass in preclinical rodent models with varying degree of prostate sparing. Phase I trials of SARMs in humans have reported modest increments in fat-free mass. Summary SARMs hold promise as a new class of function promoting anabolic therapies for a number of clinical indications, including functional limitations associated with aging and chronic disease, frailty, cancer cachexia, and osteoporosis. PMID:19357508

Androgen receptor regulated microRNA miR-182-5p promotes prostate cancer progression by targeting the ARRDC3/ITGB4 pathway

Energy Technology Data Exchange (ETDEWEB)

Yao, Jingjing [Department of Urology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, 200433 (China); Xu, Chen [Research Center of Developmental Biology, Second Military Medical University, 800th Xiangyin Road, Shanghai, 200433 (China); Department of Orthopedics, Changzheng Hospital Affiliated to Second Military Medical University, 415th Feng Yang Road, Shanghai, 200003 (China); Fang, Ziyu; Li, Yaoming [Department of Urology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, 200433 (China); Liu, Houqi; Wang, Yue [Research Center of Developmental Biology, Second Military Medical University, 800th Xiangyin Road, Shanghai, 200433 (China); Translational Medicine Center, Second Military Medical University, 800th Xiangyin Road, Shanghai, 200433 (China); Xu, Chuanliang [Department of Urology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, 200433 (China); Sun, Yinghao, E-mail: sunyh@medmail.com.cn [Department of Urology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, 200433 (China)

2016-05-20

Abstracts: MicroRNAs (miRNAs) are important endogenous gene regulators that play key roles in prostate cancer development and metastasis. However, specific miRNA expression patterns in prostate cancer tissues from Chinese patients remain largely unknown. In this study, we compared miRNA expression patterns in 65 pairs of prostate cancer and para-cancer tissues by RNA sequencing and found that miR-182-5p was the most up-regulated miRNA in prostate cancer tissues. The result was validated using realtime PCR in 18 pairs of prostate cancer and para-cancer tissues. In in vitro analysis, it was confirmed that miR-182-5p promotes prostate cancer cell proliferation, invasion and migration and inhibit apoptosis. In addition, the androgen receptor directly regulated the transcription of miR-182-5p, which could target to the 3′UTR of ARRDC3 mRNA and affect the expression of ARRDC3 and its downstream gene ITGB4. For the in vivo experiment, miR-182-5p overexpression also promoted the growth and progression of prostate cancer tumors. In this regard, we suggest that miR-182-5p may be a key androgen receptor-regulated factor that contributes to the development and metastasis of Chinese prostate cancers and may be a potential target for the early diagnosis and therapeutic studies of prostate cancer. -- Highlights: •miR-182-5p is the mostly up-regulated miRNA in Chinese prostate cancer. •miR-182-5p is regulated by androgen receptor. •miR-182-5p promotes prostate cancer progression. •miR-182-5p regulates ARRDC3/ITGB4 pathway.

Androgen receptor regulated microRNA miR-182-5p promotes prostate cancer progression by targeting the ARRDC3/ITGB4 pathway

International Nuclear Information System (INIS)

Yao, Jingjing; Xu, Chen; Fang, Ziyu; Li, Yaoming; Liu, Houqi; Wang, Yue; Xu, Chuanliang; Sun, Yinghao

2016-01-01

Abstracts: MicroRNAs (miRNAs) are important endogenous gene regulators that play key roles in prostate cancer development and metastasis. However, specific miRNA expression patterns in prostate cancer tissues from Chinese patients remain largely unknown. In this study, we compared miRNA expression patterns in 65 pairs of prostate cancer and para-cancer tissues by RNA sequencing and found that miR-182-5p was the most up-regulated miRNA in prostate cancer tissues. The result was validated using realtime PCR in 18 pairs of prostate cancer and para-cancer tissues. In in vitro analysis, it was confirmed that miR-182-5p promotes prostate cancer cell proliferation, invasion and migration and inhibit apoptosis. In addition, the androgen receptor directly regulated the transcription of miR-182-5p, which could target to the 3′UTR of ARRDC3 mRNA and affect the expression of ARRDC3 and its downstream gene ITGB4. For the in vivo experiment, miR-182-5p overexpression also promoted the growth and progression of prostate cancer tumors. In this regard, we suggest that miR-182-5p may be a key androgen receptor-regulated factor that contributes to the development and metastasis of Chinese prostate cancers and may be a potential target for the early diagnosis and therapeutic studies of prostate cancer. -- Highlights: •miR-182-5p is the mostly up-regulated miRNA in Chinese prostate cancer. •miR-182-5p is regulated by androgen receptor. •miR-182-5p promotes prostate cancer progression. •miR-182-5p regulates ARRDC3/ITGB4 pathway.

Olanzapine Reverses MK-801-Induced Cognitive Deficits and Region-Specific Alterations of NMDA Receptor Subunits

Science.gov (United States)

Liu, Xiao; Li, Jitao; Guo, Chunmei; Wang, Hongli; Sun, Yaxin; Wang, Han; Su, Yun-Ai; Li, Keqing; Si, Tianmei

2018-01-01

Cognitive dysfunction constitutes an essential component in schizophrenia for its early presence in the pathophysiology of the disease and close relatedness to life quality of patients. To develop effective treatment of cognitive deficits, it is important to understand their neurobiological causes and to identify potential therapeutic targets. In this study, adopting repeated MK-801 treatment as an animal model of schizophrenia, we investigated whether antipsychotic drugs, olanzapine and haloperidol, can reverse MK-801-induced cognitive deficits and how the reversal processes recruited proteins involved in glutamate neurotransmission in rat medial prefrontal cortex (mPFC) and hippocampus. We found that low-dose chronic MK-801 treatment impaired object-in-context recognition memory and reversal learning in the Morris water maze, leaving reference memory relatively unaffected, and that these cognitive deficits can be partially reversed by olanzapine, not haloperidol, treatment. At the molecular level, chronic MK-801 treatment resulted in the reduction of multiple N-methyl-D-aspartate (NMDA) receptor subunits in rat mPFC and olanzapine, not haloperidol, treatment restored the levels of GluN1 and phosphorylated GluN2B in this region. Taken together, MK-801-induced cognitive deficits may be associated with region-specific changes in NMDA receptor subunits and the reversal of specific NMDA receptor subunits may underlie the cognition-enhancing effects of olanzapine. PMID:29375333

Death Receptor 6 Promotes Wallerian Degeneration in Peripheral Axons.

Science.gov (United States)

Gamage, Kanchana K; Cheng, Irene; Park, Rachel E; Karim, Mardeen S; Edamura, Kazusa; Hughes, Christopher; Spano, Anthony J; Erisir, Alev; Deppmann, Christopher D

2017-03-20

Axon degeneration during development is required to sculpt a functional nervous system and is also a hallmark of pathological insult, such as injury [1, 2]. Despite similar morphological characteristics, very little overlap in molecular mechanisms has been reported between pathological and developmental degeneration [3-5]. In the peripheral nervous system (PNS), developmental axon pruning relies on receptor-mediated extrinsic degeneration mechanisms to determine which axons are maintained or degenerated [5-7]. Receptors have not been implicated in Wallerian axon degeneration; instead, axon autonomous, intrinsic mechanisms are thought to be the primary driver for this type of axon disintegration [8-10]. Here we survey the role of neuronally expressed, paralogous tumor necrosis factor receptor super family (TNFRSF) members in Wallerian degeneration. We find that an orphan receptor, death receptor 6 (DR6), is required to drive axon degeneration after axotomy in sympathetic and sensory neurons cultured in microfluidic devices. We sought to validate these in vitro findings in vivo using a transected sciatic nerve model. Consistent with the in vitro findings, DR6 -/- animals displayed preserved axons up to 4 weeks after injury. In contrast to phenotypes observed in Wld s and Sarm1 -/- mice, preserved axons in DR6 -/- animals display profound myelin remodeling. This indicates that deterioration of axons and myelin after axotomy are mechanistically distinct processes. Finally, we find that JNK signaling after injury requires DR6, suggesting a link between this novel extrinsic pathway and the axon autonomous, intrinsic pathways that have become established for Wallerian degeneration. Copyright © 2017 Elsevier Ltd. All rights reserved.

Source apportionment of PM2.5 at the Lin'an regional background site in China with three receptor models

Science.gov (United States)

Deng, Junjun; Zhang, Yanru; Qiu, Yuqing; Zhang, Hongliang; Du, Wenjiao; Xu, Lingling; Hong, Youwei; Chen, Yanting; Chen, Jinsheng

2018-04-01

Source apportionment of fine particulate matter (PM2.5) were conducted at the Lin'an Regional Atmospheric Background Station (LA) in the Yangtze River Delta (YRD) region in China from July 2014 to April 2015 with three receptor models including principal component analysis combining multiple linear regression (PCA-MLR), UNMIX and Positive Matrix Factorization (PMF). The model performance, source identification and source contribution of the three models were analyzed and inter-compared. Source apportionment of PM2.5 was also conducted with the receptor models. Good correlations between the reconstructed and measured concentrations of PM2.5 and its major chemical species were obtained for all models. PMF resolved almost all masses of PM2.5, while PCA-MLR and UNMIX explained about 80%. Five, four and seven sources were identified by PCA-MLR, UNMIX and PMF, respectively. Combustion, secondary source, marine source, dust and industrial activities were identified by all the three receptor models. Combustion source and secondary source were the major sources, and totally contributed over 60% to PM2.5. The PMF model had a better performance on separating the different combustion sources. These findings improve the understanding of PM2.5 sources in background region.

Elevated chemokine CC-motif receptor-like 2 (CCRL2) promotes cell migration and invasion in glioblastoma.

Science.gov (United States)

Yin, Fengqiong; Xu, Zhenhua; Wang, Zifeng; Yao, Hong; Shen, Zan; Yu, Fang; Tang, Yiping; Fu, Dengli; Lin, Sheng; Lu, Gang; Kung, Hsiang-Fu; Poon, Wai Sang; Huang, Yunchao; Lin, Marie Chia-Mi

2012-12-14

Chemokine CC-motif receptor-like 2 (CCRL2) is a 7-transmembrane G protein-coupled receptor which plays a key role in lung dendritic cell trafficking to peripheral lymph nodes. The function and expression of CCRL2 in cancer is not understood at present. Here we report that CCRL2 expression level is elevated in human glioma patient samples and cell lines. The magnitude of increase is positively associated with increasing tumor grade, with the highest level observed in grade IV glioblastoma. By gain-of-function and loss-of-function studies, we further showed that CCRL2 did not regulate the growth of human glioblatoma U87 and U373 cells. Importantly, we demonstrated that over-expression of CCRL2 significantly enhanced the migration rate and invasiveness of the glioblastoma cells. Taken together, these results suggest for the first time that elevated CCRL2 in glioma promotes cell migration and invasion. The potential roles of CCRL2 as a novel therapeutic target and biomarker warrant further investigations. Copyright © 2012 Elsevier Inc. All rights reserved.

Mediator-dependent Nuclear Receptor Functions

Science.gov (United States)

Chen, Wei; Roeder, Robert

2011-01-01

As gene-specific transcription factors, nuclear hormone receptors are broadly involved in many important biological processes. Their function on target genes requires the stepwise assembly of different coactivator complexes that facilitate chromatin remodeling and subsequent preinitiation complex (PIC) formation and function. Mediator has proved to be a crucial, and general, nuclear receptor-interacting coactivator, with demonstrated functions in transcription steps ranging from chromatin remodeling to subsequent PIC formation and function. Here we discuss (i) our current understanding of pathways that nuclear receptors and other interacting cofactors employ to recruit Mediator to target gene enhancers and promoters, including conditional requirements for the strong NR-Mediator interactions mediated by the NR AF2 domain and the MED1 LXXLLL motifs and (ii) mechanisms by which Mediator acts to transmit signals from enhancer-bound nuclear receptors to the general transcription machinery at core promoters to effect PIC formation and function. PMID:21854863

Systematic screening for mutations in the 5{prime}-regulatory region of the human dopamine D{sub 1} receptor (DRD1) gene in patients with schizophrenia and bipolar affective disorder

Energy Technology Data Exchange (ETDEWEB)

Cichon, S.; Noethen, M.M.; Stoeber, G. [Univ. of Bonn (Germany)] [and others

1996-07-26

A possible dysregulation of dopaminergic neurotransmission has been implicated in a variety of neuropsychiatric diseases. In the present study we systematically searched for the presence of mutations in the 5{prime}-flanking region of the dopamine D{sub 1} receptor (DRD1) gene. This region has previously been shown to contain a functional promoter. We investigated 119 unrelated individuals (including 36 schizophrenic patients, 38 bipolar affective patients, and 45 healthy controls) using single-strand conformation analysis (SSCA). Eleven overlapping PCR fragments covered 2,189 bp of DNA sequence. We identified six single base substitutions: -2218T/C, -2102C/A, -2030T/C, -1992G/A, -1251G/C, and -800T/C. None of the mutations was found to be located in regions which have important influence on the level of transcriptional activity. Allele frequencies were similar in patients and controls, indicating that genetic variation in the 5{prime}-regulatory region of the DRD1 gene is unlikely to play a frequent, major role in the genetic predisposition to either schizophrenia or bipolar affective disorder. 31 refs., 3 tabs.

Tumour MLH1 promoter region methylation testing is an effective prescreen for Lynch Syndrome (HNPCC).

Science.gov (United States)

Newton, K; Jorgensen, N M; Wallace, A J; Buchanan, D D; Lalloo, F; McMahon, R F T; Hill, J; Evans, D G

2014-12-01

Lynch syndrome (LS) patients have DNA mismatch repair deficiency and up to 80% lifetime risk of colorectal cancer (CRC). Screening of mutation carriers reduces CRC incidence and mortality. Selection for constitutional mutation testing relies on family history (Amsterdam and Bethesda Guidelines) and tumour-derived biomarkers. Initial biomarker analysis uses mismatch repair protein immunohistochemistry and microsatellite instability. Abnormalities in either identify mismatch repair deficiency but do not differentiate sporadic epigenetic defects, due to MLH1 promoter region methylation (13% of CRCs) from LS (4% of CRCs). A diagnostic biomarker capable of making this distinction would be valuable. This study compared two biomarkers in tumours with mismatch repair deficiency; quantification of methylation of the MLH1 promoter region using a novel assay and BRAF c.1799T>A, p.(Val600Glu) mutation status in the identification of constitutional mutations. Tumour DNA was extracted (formalin fixed, paraffin embedded, FFPE tissue) and pyrosequencing used to test for MLH1 promoter methylation and presence of the BRAF c.1799T>A, p.(Val600Glu) mutation 71 CRCs from individuals with pathogenic MLH1 mutations and 73 CRCs with sporadic MLH1 loss. Specificity and sensitivity was compared. Unmethylated MLH1 promoter: sensitivity 94.4% (95% CI 86.2% to 98.4%), specificity 87.7% (95% CI 77.9% to 94.2%), Wild-type BRAF (codon 600): sensitivity 65.8% (95% CI 53.7% to 76.5%), specificity 98.6% (95% CI 92.4% to 100.0%) for the identification of those with pathogenic MLH1 mutations. Quantitative MLH1 promoter region methylation using pyrosequencing is superior to BRAF codon 600 mutation status in identifying constitutional mutations in mismatch repair deficient tumours. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

Phenobarbital indirectly activates the constitutive active androstane receptor (CAR) by inhibition of epidermal growth factor receptor signaling.

Science.gov (United States)

Mutoh, Shingo; Sobhany, Mack; Moore, Rick; Perera, Lalith; Pedersen, Lee; Sueyoshi, Tatsuya; Negishi, Masahiko

2013-05-07

Phenobarbital is a central nervous system depressant that also indirectly activates nuclear receptor constitutive active androstane receptor (CAR), which promotes drug and energy metabolism, as well as cell growth (and death), in the liver. We found that phenobarbital activated CAR by inhibiting epidermal growth factor receptor (EGFR) signaling. Phenobarbital bound to EGFR and potently inhibited the binding of EGF, which prevented the activation of EGFR. This abrogation of EGFR signaling induced the dephosphorylation of receptor for activated C kinase 1 (RACK1) at Tyr(52), which then promoted the dephosphorylation of CAR at Thr(38) by the catalytic core subunit of protein phosphatase 2A. The findings demonstrated that the phenobarbital-induced mechanism of CAR dephosphorylation and activation is mediated through its direct interaction with and inhibition of EGFR.

The lactate receptor, G-protein-coupled receptor 81/hydroxycarboxylic acid receptor 1

DEFF Research Database (Denmark)

Morland, Cecilie; Lauritzen, Knut Huso; Puchades, Maja

2015-01-01

We have proposed that lactate is a “volume transmitter” in the brain and underpinned this by showing that the lactate receptor, G-protein-coupled receptor 81 (GPR81, also known as HCA1 or HCAR1), which promotes lipid storage in adipocytes, is also active in the mammalian brain. This includes......, energy metabolism, and energy substrate availability, including a glucose- and glycogen-saving response. HCAR1 may contribute to optimizing the cAMP concentration. For instance, in the prefrontal cortex, excessively high cAMP levels are implicated in impaired cognition in old age, fatigue, stress...

Genomic organization and identification of promoter regions for the BDNF gene in the pond turtle Trachemys scripta elegans.

Science.gov (United States)

Ambigapathy, Ganesh; Zheng, Zhaoqing; Keifer, Joyce

2014-08-01

Brain-derived neurotrophic factor (BDNF) is an important regulator of neuronal development and synaptic function. The BDNF gene undergoes significant activity-dependent regulation during learning. Here, we identified the BDNF promoter regions, transcription start sites, and potential regulatory sequences for BDNF exons I-III that may contribute to activity-dependent gene and protein expression in the pond turtle Trachemys scripta elegans (tBDNF). By using transfection of BDNF promoter/luciferase plasmid constructs into human neuroblastoma SHSY5Y cells and mouse embryonic fibroblast NIH3T3 cells, we identified the basal regulatory activity of promoter sequences located upstream of each tBDNF exon, designated as pBDNFI-III. Further, through chromatin immunoprecipitation (ChIP) assays, we detected CREB binding directly to exon I and exon III promoters, while BHLHB2, but not CREB, binds within the exon II promoter. Elucidation of the promoter regions and regulatory protein binding sites in the tBDNF gene is essential for understanding the regulatory mechanisms that control tBDNF gene expression.

A Novel Tenebrio molitor Cadherin Is a Functional Receptor for Bacillus thuringiensis Cry3Aa Toxin*

Science.gov (United States)

Fabrick, Jeff; Oppert, Cris; Lorenzen, Marcé D.; Morris, Kaley; Oppert, Brenda; Jurat-Fuentes, Juan Luis

2009-01-01

Cry toxins produced by the bacterium Bacillus thuringiensis are effective biological insecticides. Cadherin-like proteins have been reported as functional Cry1A toxin receptors in Lepidoptera. Here we present data that demonstrate that a coleopteran cadherin is a functional Cry3Aa toxin receptor. The Cry3Aa receptor cadherin was cloned from Tenebrio molitor larval midgut mRNA, and the predicted protein, TmCad1, has domain structure and a putative toxin binding region similar to those in lepidopteran cadherin B. thuringiensis receptors. A peptide containing the putative toxin binding region from TmCad1 bound specifically to Cry3Aa and promoted the formation of Cry3Aa toxin oligomers, proposed to be mediators of toxicity in lepidopterans. Injection of TmCad1-specific double-stranded RNA into T. molitor larvae resulted in knockdown of the TmCad1 transcript and conferred resistance to Cry3Aa toxicity. These data demonstrate the functional role of TmCad1 as a Cry3Aa receptor in T. molitor and reveal similarities between the mode of action of Cry toxins in Lepidoptera and Coleoptera. PMID:19416969

Nogo-66 receptor antagonist peptide (NEP1-40) administration promotes functional recovery and axonal growth after lateral funiculus injury in the adult rat

NARCIS (Netherlands)

Cao, Y.; Shumsky, J. S.; Sabol, M. A.; Kushner, R. A.; Strittmatter, S.; Hamers, F. P. T.; Lee, D. H. S.; Rabacchi, S. A.; Murray, M.

2008-01-01

Objective. The myelin protein Nogo inhibits axon regeneration by binding to its receptor (NgR) on axons. Intrathecal delivery of an NgR antagonist (NEP1-40) promotes growth of injured corticospinal axons and recovery of motor function following a dorsal hemisection. The authors used a similar design

Early vertebrate chromosome duplications and the evolution of the neuropeptide Y receptor gene regions

Directory of Open Access Journals (Sweden)

Brenner Sydney

2008-06-01

Full Text Available Abstract Background One of the many gene families that expanded in early vertebrate evolution is the neuropeptide (NPY receptor family of G-protein coupled receptors. Earlier work by our lab suggested that several of the NPY receptor genes found in extant vertebrates resulted from two genome duplications before the origin of jawed vertebrates (gnathostomes and one additional genome duplication in the actinopterygian lineage, based on their location on chromosomes sharing several gene families. In this study we have investigated, in five vertebrate genomes, 45 gene families with members close to the NPY receptor genes in the compact genomes of the teleost fishes Tetraodon nigroviridis and Takifugu rubripes. These correspond to Homo sapiens chromosomes 4, 5, 8 and 10. Results Chromosome regions with conserved synteny were identified and confirmed by phylogenetic analyses in H. sapiens, M. musculus, D. rerio, T. rubripes and T. nigroviridis. 26 gene families, including the NPY receptor genes, (plus 3 described recently by other labs showed a tree topology consistent with duplications in early vertebrate evolution and in the actinopterygian lineage, thereby supporting expansion through block duplications. Eight gene families had complications that precluded analysis (such as short sequence length or variable number of repeated domains and another eight families did not support block duplications (because the paralogs in these families seem to have originated in another time window than the proposed genome duplication events. RT-PCR carried out with several tissues in T. rubripes revealed that all five NPY receptors were expressed in the brain and subtypes Y2, Y4 and Y8 were also expressed in peripheral organs. Conclusion We conclude that the phylogenetic analyses and chromosomal locations of these gene families support duplications of large blocks of genes or even entire chromosomes. Thus, these results are consistent with two early vertebrate

The Coordinated P53 and Estrogen Receptor Cis-Regulation at an FLT1 Promoter SNP Is Specific to Genotoxic Stress and Estrogenic Compound

Science.gov (United States)

Langen, Jan-Stephan; Schoenfelder, Gilbert; Resnick, Michael A.; Inga, Alberto

2010-01-01

Background Recently, we established that a C>T single nucleotide polymorphism (SNP) in the promoter of the VEGF receptor FLT1 gene generates a ½ site p53 response element (RE-T) that results in p53 responsiveness of the promoter. The transcriptional control required an estrogen receptor (ER) ½ site response element (ERE1) 225 nt upstream to the RE-T. Methodology/Principal Findings Here we report the identification of a second ER ½ site (ERE2) located 145 bp downstream of the RE-T and establish that both EREs can impact p53-mediated transactivation of FLT1-T in a manner that is cell type and ER level dependent. Gene reporter assays and ChIP experiments conducted in the breast cancer-derived MCF7 cells revealed that the ERE2 site was sufficient for p53-mediated ERα recruitment and transactivation of the FLT1-T promoter/reporter construct. Surprisingly, unlike the case for other p53 target promoters, p53-mediated transactivation of FLT1-T constructs or expression of the endogenous FLT1 gene, as well as binding of p53 and ER at the promoter constructs, was inducible by doxorubicin but not by 5-fluorouracil. Furthermore, ER activity at FLT1-T was differentially affected by ER ligands, compared to a control TFF1/pS2 ER target promoter. The p53-related transcription factors (TFs) p73 and p63 had no effect on FLT1 transactivation. Conclusions/Significance We establish a new dimension to the p53 master regulatory network where p53-mediated transcription from a ½ site RE can be determined by ER binding at one or more cis-acting EREs in manner that is dependent on level of ER protein, the type of ER ligand and the specific p53-inducing agent. PMID:20422012

Mediator and p300/CBP-Steroid Receptor Coactivator Complexes Have Distinct Roles, but Function Synergistically, during Estrogen Receptor α-Dependent Transcription with Chromatin Templates

OpenAIRE

Acevedo, Mari Luz; Kraus, W. Lee

2003-01-01

Ligand-dependent transcriptional activation by nuclear receptors involves the recruitment of various coactivators to the promoters of hormone-regulated genes assembled into chromatin. Nuclear receptor coactivators include histone acetyltransferase complexes, such as p300/CBP-steroid receptor coactivator (SRC), as well as the multisubunit mediator complexes (“Mediator”), which may help recruit RNA polymerase II to the promoter. We have used a biochemical approach, including an in vitro chromat...

Hypermethylation of E-Cadherin and Estrogen Receptor-a Gene Promoter and Its Association with Clinicopathological Features of Breast Cancer in Iranian Patients

Directory of Open Access Journals (Sweden)

Mozhgan Rasti

2009-06-01

Full Text Available Background: Aberrant methylation of cytosine-guanine dinucleotideislands leads to inactivation of tumor suppressorgenes in breast cancer. Tumor suppressor genes are unmethylatedin normal tissue and often become hypermethylatedduring tumor formation, leading to gene silencing. We investigatedthe association between E-cadherin (CDH1 and estrogenreceptor-α (ESRα gene promoter methylation andmajor clinical and pathological features of breast cancer inIranian women.Methods: DNA was extracted from 67 primary breast tumorsand gene promoter methylation was analyzed by methylationspecificpolymerase chain reaction method.Results: Fifty percent of the samples showed aberrant methylationin at least one of the two tested loci. We detectedCDH1 hypermethylation in 41% of invasive tumors and receptor-α gene hypermethylation in 18% of invasive tumorsamples. We found no association between CDH1 and receptor-α gene hypermethylation (P=0.45. There was a correlationbetween hypermethylation of CDH1 locus and tumorsize ≥5 cm (P=0.019.Conclusion: Our data suggest that the malignant progressionof human ductal and lobular breast carcinoma in Iranianwomen involves a heterogeneous pattern of cytosine-guaninedinucleotide island hypermethylation of the CDH1 gene.

SPARC and GluA1-Containing AMPA Receptors Promote Neuronal Health Following CNS Injury

Directory of Open Access Journals (Sweden)

Emma V. Jones

2018-02-01

Full Text Available The proper formation and maintenance of functional synapses in the central nervous system (CNS requires communication between neurons and astrocytes and the ability of astrocytes to release neuromodulatory molecules. Previously, we described a novel role for the astrocyte-secreted matricellular protein SPARC (Secreted Protein, Acidic and Rich in Cysteine in regulating α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs and plasticity at developing synapses. SPARC is highly expressed by astrocytes and microglia during CNS development but its level is reduced in adulthood. Interestingly, SPARC has been shown to be upregulated in CNS injury and disease. However, the role of SPARC upregulation in these contexts is not fully understood. In this study, we investigated the effect of chronic SPARC administration on glutamate receptors on mature hippocampal neuron cultures and following CNS injury. We found that SPARC treatment increased the number of GluA1-containing AMPARs at synapses and enhanced synaptic function. Furthermore, we determined that the increase in synaptic strength induced by SPARC could be inhibited by Philanthotoxin-433, a blocker of homomeric GluA1-containing AMPARs. We then investigated the effect of SPARC treatment on neuronal health in an injury context where SPARC expression is upregulated. We found that SPARC levels are increased in astrocytes and microglia following middle cerebral artery occlusion (MCAO in vivo and oxygen-glucose deprivation (OGD in vitro. Remarkably, chronic pre-treatment with SPARC prevented OGD-induced loss of synaptic GluA1. Furthermore, SPARC treatment reduced neuronal death through Philanthotoxin-433 sensitive GluA1 receptors. Taken together, this study suggests a novel role for SPARC and GluA1 in promoting neuronal health and recovery following CNS damage.

Prostaglandin D2 effects and DP1 /DP2 receptor distribution in guinea pig urinary bladder out-flow region.

Science.gov (United States)

Guan, Na N; Svennersten, Karl; de Verdier, Petra J; Wiklund, N Peter; Gustafsson, Lars E

2017-02-01

The proximal urethra and urinary bladder trigone play important roles in continence. We have previously shown that PGD 2 is released from guinea pig bladder urothelium/suburothelium and can inhibit detrusor contractile responses. We presently wished to investigate PGD 2 actions in guinea pig out-flow region and the distribution of DP 1 /DP 2 receptors. The effects of PGD 2 on urothelium-intact trigone and proximal urethra contractility were studied in organ bath experiments. Expression of DP 1 /DP 2 receptor proteins was analysed by western blot. Immunohistochemistry was used to identify distribution of DP 1 /DP 2 receptors. PGD 2 in a dose-dependent manner inhibited trigone contractions induced by electrical field stimulation (EFS) and inhibited spontaneous contractions of the proximal urethra. PGD 2 was equally (trigone) or slightly less potent (urethra) compared with PGE 2 . Expression of DP 1 and DP 2 receptors was found in male guinea pig bladder trigone, neck and proximal urethra. In the trigone and proximal urethra, DP 1 receptors were found on the membrane of smooth muscle cells and weak immunoreactivty was observed in the urothelium. DP 2 receptors were distributed more widespread, weakly and evenly in the urothelium and smooth muscles. Inhibitory effects by PGD 2 on motor activity of guinea pig trigone and proximal urethra are consistent with finding DP 1 and DP 2 receptors located in the urothelium and smooth muscle cells of the trigone and proximal urethra, and PGD 2 may therefore be a modulator of the bladder out-flow region, possibly having a function in regulation of micturition and a role in overactive bladder syndrome. © 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

New PAH gene promoter KLF1 and 3'-region C/EBPalpha motifs influence transcription in vitro.

Science.gov (United States)

Klaassen, Kristel; Stankovic, Biljana; Kotur, Nikola; Djordjevic, Maja; Zukic, Branka; Nikcevic, Gordana; Ugrin, Milena; Spasovski, Vesna; Srzentic, Sanja; Pavlovic, Sonja; Stojiljkovic, Maja

2017-02-01

Phenylketonuria (PKU) is a metabolic disease caused by mutations in the phenylalanine hydroxylase (PAH) gene. Although the PAH genotype remains the main determinant of PKU phenotype severity, genotype-phenotype inconsistencies have been reported. In this study, we focused on unanalysed sequences in non-coding PAH gene regions to assess their possible influence on the PKU phenotype. We transiently transfected HepG2 cells with various chloramphenicol acetyl transferase (CAT) reporter constructs which included PAH gene non-coding regions. Selected non-coding regions were indicated by in silico prediction to contain transcription factor binding sites. Furthermore, electrophoretic mobility shift assay (EMSA) and supershift assays were performed to identify which transcriptional factors were engaged in the interaction. We found novel KLF1 motif in the PAH promoter, which decreases CAT activity by 50 % in comparison to basal transcription in vitro. The cytosine at the c.-170 promoter position creates an additional binding site for the protein complex involving KLF1 transcription factor. Moreover, we assessed for the first time the role of a multivariant variable number tandem repeat (VNTR) region located in the 3'-region of the PAH gene. We found that the VNTR3, VNTR7 and VNTR8 constructs had approximately 60 % of CAT activity. The regulation is mediated by the C/EBPalpha transcription factor, present in protein complex binding to VNTR3. Our study highlighted two novel promoter KLF1 and 3'-region C/EBPalpha motifs in the PAH gene which decrease transcription in vitro and, thus, could be considered as PAH expression modifiers. New transcription motifs in non-coding regions will contribute to better understanding of the PKU phenotype complexity and may become important for the optimisation of PKU treatment.

Cytosine deletion at AP2-box region of HSP70 promoter and its ...

Indian Academy of Sciences (India)

Cytosine deletion at AP2-box region of HSP70 promoter and its influence on semen quality traits in crossbred bulls ... Laboratory, ICAR-Central Institute for Research on Cattle, Meerut 250 001, India; School of Atmospheric Stress Management, ICAR-National Institute of Abiotic Stress Management, Baramati 413 115, India ...

“Fear or Love Thy Neighbour”? The EU Framework for Promoting Regional Cooperation in the South Caucasus

Directory of Open Access Journals (Sweden)

Nelli Babayan

2011-12-01

Full Text Available Building on the model of the enlargement policy, the European Union (EU designed the European Neighbourhood Policy and the Eastern Partnership to further promote its norms and principles. One of the goals of its new policies has been to foster regional cooperation among partner countries and their neighbours. This article specifies the EU’s framework for promoting regional cooperation through the aforementioned policies and discusses its potential impact on the example of the South Caucasus republics of Armenia, Azerbaijan, and Georgia. The South Caucasus has not only been an arena of intraregional conflicts, but has also often been troubled by disputes between its neighbours. This article argues that, due to a lack of proactive and consistent engagement, the EU’s framework risks leaving regional conflicts in the current state of stagnation and without advancement in regional cooperation.

Functional importance of the Ala(116)-Pro(136) region in the calcium-sensing receptor. Constitutive activity and inverse agonism in a family C G-protein-coupled receptor

DEFF Research Database (Denmark)

Jensen, Anders A.; Spalding, T A; Burstein, E S

2000-01-01

The calcium-sensing receptor (CaR) belongs to family C of the G-protein-coupled receptor superfamily. To date 14 activating mutations in CaR showing increased sensitivity to Ca(2+) have been identified in humans with autosomal dominant hypocalcemia. Four of these activating mutations are found......, suppressed the elevated basal response of the constitutively activated Ca/1a mutants demonstrating inverse agonist activity of CPCCOEt. Taken together, our results demonstrate that the Ala(116)-Pro(136) region is of key importance for the maintenance of the inactive conformation of CaR....

Autocrine CSF-1 and CSF-1 Receptor Co-expression Promotes Renal Cell Carcinoma Growth

Science.gov (United States)

Menke, Julia; Kriegsmann, Jörg; Schimanski, Carl Christoph; Schwartz, Melvin M.; Schwarting, Andreas; Kelley, Vicki R.

2011-01-01

Renal cell carcinoma is increasing in incidence but the molecular mechanisms regulating its growth remain elusive. Co-expression of the monocytic growth factor CSF-1 and its receptor CSF-1R on renal tubular epithelial cells (TEC) will promote proliferation and anti-apoptosis during regeneration of renal tubules. Here we show that a CSF-1-dependent autocrine pathway is also responsible for the growth of renal cell carcinoma (RCC). CSF-1 and CSF-1R were co-expressed in RCC and TEC proximally adjacent to RCC. CSF-1 engagement of CSF-1R promoted RCC survival and proliferation and reduced apoptosis, in support of the likelihood that CSF-1R effector signals mediate RCC growth. In vivo CSF-1R blockade using a CSF-1R tyrosine kinase inhibitor decreased RCC proliferation and macrophage infiltration in a manner associated with a dramatic reduction in tumor mass. Further mechanistic investigations linked CSF-1 and EGF signaling in RCC. Taken together, our results suggest that budding RCC stimulates the proximal adjacent microenvironment in the kidney to release mediators of CSF-1, CSF-1R and EGF expression in RCC. Further, our findings imply that targeting CSF-1/CSF-1R signaling may be therapeutically effective in RCC. PMID:22052465

Mutation of the SHP-2 binding site in growth hormone (GH) receptor prolongs GH-promoted tyrosyl phosphorylation of GH receptor, JAK2, and STAT5B

DEFF Research Database (Denmark)

Stofega, M R; Herrington, J; Billestrup, Nils

2000-01-01

phosphorylation. Consistent with the effects on STAT5B phosphorylation, tyrosine-to-phenylalanine mutation of tyrosine 595 prolongs the duration of tyrosyl phosphorylation of GHR and JAK2. These data suggest that tyrosine 595 is a major site of interaction of GHR with SHP-2, and that GHR-bound SHP-2 negatively......Binding of GH to GH receptor (GHR) rapidly and transiently activates multiple signal transduction pathways that contribute to the growth-promoting and metabolic effects of GH. While the events that initiate GH signal transduction, such as activation of the Janus tyrosine kinase JAK2, are beginning...

Data describing the effect of DRD4 promoter polymorphisms on promoter activity

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Shoin Tei

2016-06-01

Full Text Available This data article tested whether polymorphisms within the dopamine D4 receptor (DRD4 gene promoter can lead to differences in the promoter activity. The variants, a 120-bp variable number tandem repeat (VNTR, −906 T/C, −809 G/A, −616G/C, and −521C/T, were introduced into the DRD4 promoter and the promoter activity was measured in a neural cell line using the luciferase assay. However, no differences were detected among the haplotypes investigated, and the in vitro data obtained from our protocol could not support the involvement of DRD4 promoter polymorphisms in heritable human traits.

NK-1 receptor antagonists as anti-cancer drugs

Indian Academy of Sciences (India)

The substance P (SP)/neurokinin (NK)-1 receptor system plays an important role in cancer. SP promotes the proliferation of tumour cells, angiogenesis and the migration of tumour cells. We review the involvement of SP, the NK-1 receptor and NK-1 receptor antagonists in cancer. Tumour cells overexpress NK-1 receptors, ...

Deletion of G-protein-coupled receptor 55 promotes obesity by reducing physical activity.

Science.gov (United States)

Meadows, A; Lee, J H; Wu, C-S; Wei, Q; Pradhan, G; Yafi, M; Lu, H-C; Sun, Y

2016-03-01

Cannabinoid receptor 1 (CB1) is the best-characterized cannabinoid receptor, and CB1 antagonists are used in clinical trials to treat obesity. Because of the wide range of CB1 functions, the side effects of CB1 antagonists pose serious concerns. G-protein-coupled receptor 55 (GPR55) is an atypical cannabinoid receptor, and its pharmacology and functions are distinct from CB1. GPR55 regulates neuropathic pain, gut, bone, immune functions and motor coordination. GPR55 is expressed in various brain regions and peripheral tissues. However, the roles of GPR55 in energy and glucose homeostasis are unknown. Here we have investigated the roles of GPR55 in energy balance and insulin sensitivity using GPR55-null mice (GPR55(-/-)). Body composition of the mice was measured by EchoMRI. Food intake, feeding behavior, energy expenditure and physical activity of GPR55(-/-) mice were determined by indirect calorimetry. Muscle function was assessed by forced treadmill running test. Insulin sensitivity was evaluated by glucose and insulin tolerance tests. Adipose inflammation was assessed by flow cytometry analysis of adipose tissue macrophages. The expression of inflammatory markers in adipose tissues and orexigenic/anorexigenic peptides in the hypothalamus was also analyzed by real-time PCR. GPR55(-/-) mice had normal total energy intake and feeding pattern (i.e., no changes in meal size, meal number or feeding frequency). Intriguingly, whereas adult GPR55(-/-) mice only showed a modest increase in overall body weight, they exhibited significantly increased fat mass and insulin resistance. The spontaneous locomotor activity of GPR55(-/-) mice was dramatically decreased, whereas resting metabolic rate and non-shivering thermogenesis were unchanged. Moreover, GPR55(-/-) mice exhibited significantly decreased voluntary physical activity, showing reduced running distance on the running wheels, whereas muscle function appeared to be normal. GPR55 has an important role in energy

Testosterone-Dependent Interaction between Androgen Receptor and Aryl Hydrocarbon Receptor Induces Liver Receptor Homolog 1 Expression in Rat Granulosa Cells

Science.gov (United States)

Wu, Yanguang; Baumgarten, Sarah C.; Zhou, Ping

2013-01-01

Androgens play a major role in the regulation of normal ovarian function; however, they are also involved in the development of ovarian pathologies. These contrasting effects may involve a differential response of granulosa cells to the androgens testosterone (T) and dihydrotestosterone (DHT). To determine the molecular pathways that mediate the distinct effects of T and DHT, we studied the expression of the liver receptor homolog 1 (LRH-1) gene, which is differentially regulated by these steroids. We found that although both T and DHT stimulate androgen receptor (AR) binding to the LRH-1 promoter, DHT prevents T-mediated stimulation of LRH-1 expression. T stimulated the expression of aryl hydrocarbon receptor (AHR) and its interaction with the AR. T also promoted the recruitment of the AR/AHR complex to the LRH-1 promoter. These effects were not mimicked by DHT. We also observed that the activation of extracellular regulated kinases by T is required for AR and AHR interaction. In summary, T, but not DHT, stimulates AHR expression and the interaction between AHR and AR, leading to the stimulation of LRH-1 expression. These findings could explain the distinct response of granulosa cells to T and DHT and provide a molecular mechanism by which DHT negatively affects ovarian function. PMID:23689136

Generation and reactivation of T-cell receptor A joining region pseudogenes in primates

Energy Technology Data Exchange (ETDEWEB)

Thiel, C.; Lanchbury, J.S. [Guy`s Hospital, London (United Kingdom); Otting, N. [Biomedical Primate Research Centre, Rijswijk (Netherlands)] [and others

1996-06-01

Tandemly duplicated T-cell receptor (Tcr) AJ (J{alpha}) segments contribute significantly to TCRA chain junctional region diversity in mammals. Since only limited data exists on TCRA diversity in nonhuman primates, we examined the TCRAJ regions of 37 chimpanzee and 71 rhesus macaque TCRA cDNA clones derived from inverse polymerase chain reaction on peripheral blood mononuclear cell cDNA of healthy animals. Twenty-five different TCRAJ regions were characterized in the chimpanzee and 36 in the rhesus macaque. Each bears a close structural relationship to an equivalent human TCRAJ region. Conserved amino acid motifs are shared between all three species. There are indications that differences between nonhuman primates and humans exist in the generation of TCRAJ pseudogenes. The nucleotide and amino acid sequences of the various characterized TCRAJ of each species are reported and we compare our results to the available information on human genomic sequences. Although we provide evidence of dynamic processes modifying TCRAJ segments during primate evolution, their repertoire and primary structure appears to be relatively conserved. 21 refs., 2 figs.

Multiple 5' ends of human cytomegalovirus UL57 transcripts identify a complex, cycloheximide-resistant promoter region that activates oriLyt

International Nuclear Information System (INIS)

Kiehl, Anita; Huang, Lili; Franchi, David; Anders, David G.

2003-01-01

The human cytomegalovirus (HCMV) UL57 gene lies adjacent to HCMV oriLyt, from which it is separated by an organizationally conserved, mostly noncoding region that is thought to both regulate UL57 expression and activate oriLyt function. However, the UL57 promoter has not been studied. We determined the 5' ends of UL57 transcripts toward an understanding of the potential relationship between UL57 expression and oriLyt activation. The results presented here identified three distinct 5' ends spread over 800 bp, at nt 90302, 90530, and 91138; use of these sites exhibited differential sensitivity to phosphonoformic acid treatment. Interestingly, a 10-kb UL57 transcript accumulated in cycloheximide-treated infected cells, even though other early transcripts were not detectable. However, the 10-kb transcript did not accumulate in cells treated with the more stringent translation inhibitor anisomycin. Consistent with the notion that the identified 5' ends arise from distinct transcription start sites, the sequences upstream of sites I and II functioned as promoters responsive to HCMV infection in transient assays. However, the origin-proximal promoter region III required downstream sequences for transcriptional activity. Mutation of candidate core promoter elements suggested that promoter III is regulated by an initiator region (Inr) and a downstream promoter element. Finally, a 42-bp sequence containing the candidate Inr activated a minimal oriLyt core construct in transient replication assays. Thus, these studies showed that a large, complex promoter region with novel features controls UL57 expression, and identified a sequence that regulates both UL57 transcription and oriLyt activation

Identification of the subunit of cAMP receptor protein (CRP) that functionally interacts with CytR in CRP-CytR-mediated transcriptional repression

DEFF Research Database (Denmark)

Meibom, K L; Kallipolitis, B H; Ebright, R H

2000-01-01

At promoters of the Escherichia coli CytR regulon, the cAMP receptor protein (CRP) interacts with the repressor CytR to form transcriptionally inactive CRP-CytR-promoter or (CRP)(2)-CytR-promoter complexes. Here, using "oriented heterodimer" analysis, we show that only one subunit of the CRP dimer......, the subunit proximal to CytR, functionally interacts with CytR in CRP-CytR-promoter and (CRP)(2)-CytR-promoter complexes. Our results provide information about the architecture of CRP-CytR-promoter and (CRP)(2)-CytR-promoter complexes and rule out the proposal that masking of activating region 2 of CRP...

Genetic variants in promoters and coding regions of the muscle glycogen synthase and the insulin-responsive GLUT4 genes in NIDDM

DEFF Research Database (Denmark)

Bjørbaek, C; Echwald, Søren Morgenthaler; Hubricht, P

1994-01-01

To examine the hypothesis that variants in the regulatory or coding regions of the glycogen synthase (GS) and insulin-responsive glucose transporter (GLUT4) genes contribute to insulin-resistant glucose processing of muscle from non-insulin-dependent diabetes mellitus (NIDDM) patients, promoter...... volunteers. By applying inverse polymerase chain reaction and direct DNA sequencing, 532 base pairs (bp) of the GS promoter were identified and the transcriptional start site determined by primer extension. SSCP scanning of the promoter region detected five single nucleotide substitutions, positioned at 42......'-untranslated region, and the coding region of the GLUT4 gene showed four polymorphisms, all single nucleotide substitutions, positioned at -581, 1, 30, and 582. None of the three changes in the regulatory region of the gene had any major influence on expression of the GLUT4 gene in muscle. The variant at 582...

Adenosine Receptors and Wound Healing

Directory of Open Access Journals (Sweden)

Bruce N. Cronstein

2004-01-01

Full Text Available Recent studies have demonstrated that application of topical adenosine A2A receptor agonists promotes more rapid wound closure and clinical studies are currently underway to determine the utility of topical A2A adenosine receptor agonists in the therapy of diabetic foot ulcers. The effects of adenosine A2A receptors on the cells and tissues of healing wounds have only recently been explored. We review here the known effects of adenosine A2A receptor occupancy on the cells involved in wound healing.

MECP2 promoter methylation and X chromosome inactivation in autism.

Science.gov (United States)

Nagarajan, Raman P; Patzel, Katherine A; Martin, Michelle; Yasui, Dag H; Swanberg, Susan E; Hertz-Picciotto, Irva; Hansen, Robin L; Van de Water, Judy; Pessah, Isaac N; Jiang, Ruby; Robinson, Wendy P; LaSalle, Janine M

2008-06-01

Epigenetic mechanisms have been proposed to play a role in the etiology of autism. This hypothesis is supported by the discovery of increased MECP2 promoter methylation associated with decreased MeCP2 protein expression in autism male brain. To further understand the influence of female X chromosome inactivation (XCI) and neighboring methylation patterns on aberrant MECP2 promoter methylation in autism, multiple methylation analyses were peformed on brain and blood samples from individuals with autism. Bisulfite sequencing analyses of a region 0.6 kb upstream of MECP2 in brain DNA samples revealed an abrupt transition from a highly methylated region in both sexes to a region unmethylated in males and subject to XCI in females. Chromatin immunoprecipitation analysis demonstrated that the CCTC-binding factor (CTCF) bound to this transition region in neuronal cells, consistent with a chromatin boundary at the methylation transition. Male autism brain DNA samples displayed a slight increase in methylation in this transition region, suggesting a possible aberrant spreading of methylation into the MECP2 promoter in autism males across this boundary element. In addition, autistic female brain DNA samples showed evidence for aberrant MECP2 promoter methylation as an increase in the number of bisulfite sequenced clones with undefined XCI status for MECP2 but not androgen receptor (AR). To further investigate the specificity of MECP2 methylation alterations in autism, blood DNA samples from females and mothers of males with autism were also examined for XCI skewing at AR, but no significant increase in XCI skewing was observed compared to controls. These results suggest that the aberrant MECP2 methylation in autism brain DNA samples is due to locus-specific rather than global X chromosome methylation changes.

Salicylic acid receptors activate jasmonic acid signalling through a non-canonical pathway to promote effector-triggered immunity.

Science.gov (United States)

Liu, Lijing; Sonbol, Fathi-Mohamed; Huot, Bethany; Gu, Yangnan; Withers, John; Mwimba, Musoki; Yao, Jian; He, Sheng Yang; Dong, Xinnian

2016-10-11

It is an apparent conundrum how plants evolved effector-triggered immunity (ETI), involving programmed cell death (PCD), as a major defence mechanism against biotrophic pathogens, because ETI-associated PCD could leave them vulnerable to necrotrophic pathogens that thrive on dead host cells. Interestingly, during ETI, the normally antagonistic defence hormones, salicylic acid (SA) and jasmonic acid (JA) associated with defence against biotrophs and necrotrophs respectively, both accumulate to high levels. In this study, we made the surprising finding that JA is a positive regulator of RPS2-mediated ETI. Early induction of JA-responsive genes and de novo JA synthesis following SA accumulation is activated through the SA receptors NPR3 and NPR4, instead of the JA receptor COI1. We provide evidence that NPR3 and NPR4 may mediate this effect by promoting degradation of the JA transcriptional repressor JAZs. This unique interplay between SA and JA offers a possible explanation of how plants can mount defence against a biotrophic pathogen without becoming vulnerable to necrotrophic pathogens.

The insulin and IGF1 receptor kinase domains are functional dimers in the activated state

Science.gov (United States)

Cabail, M. Zulema; Li, Shiqing; Lemmon, Eric; Bowen, Mark E.; Hubbard, Stevan R.; Miller, W. Todd

2015-03-01

The insulin receptor (IR) and insulin-like growth factor-1 receptor (IGF1R) are highly related receptor tyrosine kinases with a disulfide-linked homodimeric architecture. Ligand binding to the receptor ectodomain triggers tyrosine autophosphorylation of the cytoplasmic domains, which stimulates catalytic activity and creates recruitment sites for downstream signalling proteins. Whether the two phosphorylated tyrosine kinase domains within the receptor dimer function independently or cooperatively to phosphorylate protein substrates is not known. Here we provide crystallographic, biophysical and biochemical evidence demonstrating that the phosphorylated kinase domains of IR and IGF1R form a specific dimeric arrangement involving an exchange of the juxtamembrane region proximal to the kinase domain. In this dimer, the active position of α-helix C in the kinase N lobe is stabilized, which promotes downstream substrate phosphorylation. These studies afford a novel strategy for the design of small-molecule IR agonists as potential therapeutic agents for type 2 diabetes.

Tumour MLH1 promoter region methylation testing is an effective pre-screen for Lynch Syndrome (HNPCC)

Science.gov (United States)

Newton, K; Jorgensen, NM; Wallace, AJ; Buchanan, DD; Lalloo, F; McMahon, RFT; Hill, J; Evans, DG

2016-01-01

Background & Aims Lynch syndrome patients have DNA mismatch repair deficiency and up to 80% life-time risk of colorectal cancer. Screening of mutation carriers reduces colorectal cancer incidence and mortality. Selection for constitutional mutation testing relies on family history (Amsterdam and Bethesda Guidelines) and tumour derived biomarkers. Initial biomarker analysis uses mismatch repair protein immunohistochemistry and microsatellite instability. Abnormalities in either identify mismatch repair deficiency but do not differentiate sporadic epigenetic defects, due to MLH1 promoter region methylation (13% of CRCs) from Lynch Syndrome (4% of CRCs). A diagnostic biomarker capable of making this distinction would be valuable. This study compared two biomarkers in tumours with mismatch repair deficiency; quantification of methylation of the MLH1 promoter region using a novel assay and BRAF c.1799T>A, p.(Val600Glu) mutation status in the identification of constitutional mutations. Methods Tumour DNA was extracted (FFPE tissue) and pyrosequencing used to test for MLH1 promoter methylation and presence of the BRAF c.1799T>A, p.(Val600Glu) mutation 71 CRCs from individuals with pathogenic MLH1 mutations and 73 CRCs with sporadic MLH1 loss. Specificity and sensitivity was compared. Findings Unmethylated MLH1 promoter: sensitivity 94.4% (95% CI 86.2–98.4%), specificity 87.7% (95% CI 77.9–94.2%), Wild-type BRAF (codon 600): sensitivity 65.8% (95% CI 53.7–76.5%), specificity 98.6% (95% CI 92.4–100.0%) for the identification of those with pathogenic MLH1 mutations. Conclusions Quantitative MLH1 promoter region methylation using pyrosequencing is superior to BRAF codon 600 mutation status in identifying constitutional mutations in mismatch repair deficient tumours. PMID:25280751

Estradiol represses Insulin-like 3 expression and promoter activity in MA-10 Leydig cells

International Nuclear Information System (INIS)

Lague, Eric; Tremblay, Jacques J.

2009-01-01

There are increasing evidence in the literature reporting the detrimental effects of endocrine disruptors on the development and function of the male reproductive system. One example is cryptorchidism, or undescended testis, caused by exposure to excessive estrogens. Estrogens, acting through the estrogen receptor α (ERα), have been shown to repress expression of the gene encoding insulin-like 3 (INSL3), a small peptide produced by testicular Leydig cells that is essential for normal testis descent. The molecular mechanism of estrogen/ER action on Insl3 expression, however, remains poorly understood. Here we report estradiol (E 2 ) represses Insl3 mRNA levels in MA-10 cells, a Leydig cell line model. We also found that E 2 represses the activity of the human and mouse Insl3 promoter in these cells. The E 2 -responsive region of the human INSL3 promoter was located to the proximal INSL3 promoter. This region does not contain a consensus estrogen response element indicating an indirect mechanism of action. In agreement with this, we found that E 2 -responsiveness was lost when two previously characterized binding sites for the nuclear receptors NUR77 and SF1 were mutated. Finally we show that the E 2 repressive effect could be overcome by cotreatment with testosterone, a positive regulator of Insl3 transcription. Collectively our data provide important new insights into the molecular mechanism of estrogen action in Insl3 transcription in Leydig cells

The system of Regional Contact Offices for promoting GMES services and the use of Space Technologies in European Regions.

Science.gov (United States)

Carrara, Paola; Antoninetti, Massimo; Bacai, Hina; Basoni, Anna; Bosc, Christelle; Clave, Magali; Cornacchia, Carmela; L'Astorina, Alba; Monbet, Philippe; Mueller, Bastian; Nicolau, Sonia; Pergola, Nicola; Rampini, Anna; Tramutoli, Valerio; Schumacher, Volker; Wells, Alan; Zepeda Juarez, Jesus; Zolotikova, Svetlana

2013-04-01

which have significant impact on the economy, environment and the quality of life of the citizens To this aim since 2011 the system of Regional Contact Offices (RCOs) was promoted by the EU FP7 DORIS_Net (Downsteam Observatory organized by Regions Active in Space - Network, http://www.doris-net.eu/) project as the regional link to the services provided by the European GMES programme. Since then a first nucleus of 12 pilot European Regions were working together establishing 6 first RCOs around Europe. This paper will present RCOs network goals, achievements and perspectives as well as its planned actions devoted to improve quality of Space Technology products from one side, to promote awareness and use of them by potential end-users (and particularly LRAs), from the other side.

Cell- and stage-specific chromatin structure across the Complement receptor 2 (CR2/CD21) promoter coincide with CBF1 and C/EBP-beta binding in B cells.

Science.gov (United States)

Cruickshank, Mark N; Fenwick, Emily; Karimi, Mahdad; Abraham, Lawrence J; Ulgiati, Daniela

2009-08-01

Stringent developmental transcription requires multiple transcription factor (TF) binding sites, cell-specific expression of signaling molecules, TFs and co-regulators and appropriate chromatin structure. During B-lymphopoiesis, human Complement receptor 2 (CR2/CD21) is detected on immature and mature B cells but not on B cell precursors and plasma cells. We examined cell- and stage-specific human CR2 gene regulation using cell lines modeling B-lymphopoiesis. Chromatin accessibility assays revealed a region between -409 and -262 with enhanced accessibility in mature B cells and pre-B cells, compared to either non-lymphoid or plasma cell-types, however, accessibility near the transcription start site (TSS) was elevated only in CR2-expressing B cells. A correlation between histone acetylation and CR2 expression was observed, while histone H3K4 dimethylation was enriched near the TSS in both CR2-expressing B cells and non-expressing pre-B cells. Candidate sites within the CR2 promoter were identified which could regulate chromatin, including a matrix attachment region associated with CDP, SATB1/BRIGHT and CEBP-beta sites as well as two CBF1 sites. ChIP assays verified that both CBF1 and C/EBP-beta bind the CR2 promoter in B cells raising the possibility that these factors facilitate or respond to alterations in chromatin structure to control the timing and/or level of CR2 transcription.

Regulation of CYP3A4 by pregnane X receptor: The role of nuclear receptors competing for response element binding

Energy Technology Data Exchange (ETDEWEB)

Istrate, Monica A., E-mail: monicai@scripps.edu [Dr. Margarete Fischer-Bosch-Institute of Clinical Pharmacology, Stuttgart, Germany, and University of Tuebingen, Auerbachstr. 112, D-70376 Stuttgart (Germany); Nussler, Andreas K., E-mail: nuessler@uchir.me.tum.de [Department of Traumatology, Technical University Munich, Ismaningerstr. 22, 81675 Munich (Germany); Eichelbaum, Michel, E-mail: michel.eichelbaum@ikp-stuttgart.de [Dr. Margarete Fischer-Bosch-Institute of Clinical Pharmacology, Stuttgart, Germany, and University of Tuebingen, Auerbachstr. 112, D-70376 Stuttgart (Germany); Burk, Oliver, E-mail: oliver.burk@ikp-stuttgart.de [Dr. Margarete Fischer-Bosch-Institute of Clinical Pharmacology, Stuttgart, Germany, and University of Tuebingen, Auerbachstr. 112, D-70376 Stuttgart (Germany)

2010-03-19

Induction of the major drug metabolizing enzyme CYP3A4 by xenobiotics contributes to the pronounced interindividual variability of its expression and often results in clinically relevant drug-drug interactions. It is mainly mediated by PXR, which regulates CYP3A4 expression by binding to several specific elements in the 5' upstream regulatory region of the gene. Induction itself shows a marked interindividual variability, whose underlying determinants are only partly understood. In this study, we investigated the role of nuclear receptor binding to PXR response elements in CYP3A4, as a potential non-genetic mechanism contributing to interindividual variability of induction. By in vitro DNA binding experiments, we showed that several nuclear receptors bind efficiently to the proximal promoter ER6 and distal xenobiotic-responsive enhancer module DR3 motifs. TR{alpha}1, TR{beta}1, COUP-TFI, and COUP-TFII further demonstrated dose-dependent repression of PXR-mediated CYP3A4 enhancer/promoter reporter activity in transient transfection in the presence and absence of the PXR inducer rifampin, while VDR showed this effect only in the absence of treatment. By combining functional in vitro characterization with hepatic expression analysis, we predict that TR{alpha}1, TR{beta}1, COUP-TFI, and COUP-TFII show a strong potential for the repression of PXR-mediated activation of CYP3A4 in vivo. In summary, our results demonstrate that nuclear receptor binding to PXR response elements interferes with PXR-mediated expression and induction of CYP3A4 and thereby contributes to the interindividual variability of induction.

Increased Eps15 homology domain 1 and RAB11FIP3 expression regulate breast cancer progression via promoting epithelial growth factor receptor recycling.

Science.gov (United States)

Tong, Dandan; Liang, Ya-Nan; Stepanova, A A; Liu, Yu; Li, Xiaobo; Wang, Letian; Zhang, Fengmin; Vasilyeva, N V

2017-02-01

. Furthermore, RAB11FIP3 combines with Eps15 homology domain 1 to promote the endocytosis recycling of phosphorylation of epithelial growth factor receptor.

Assessing the effects of data selection and representation on the development of reliable E. coli sigma 70 promoter region predictors.

Directory of Open Access Journals (Sweden)

Mostafa M Abbas

Full Text Available As the number of sequenced bacterial genomes increases, the need for rapid and reliable tools for the annotation of functional elements (e.g., transcriptional regulatory elements becomes more desirable. Promoters are the key regulatory elements, which recruit the transcriptional machinery through binding to a variety of regulatory proteins (known as sigma factors. The identification of the promoter regions is very challenging because these regions do not adhere to specific sequence patterns or motifs and are difficult to determine experimentally. Machine learning represents a promising and cost-effective approach for computational identification of prokaryotic promoter regions. However, the quality of the predictors depends on several factors including: i training data; ii data representation; iii classification algorithms; iv evaluation procedures. In this work, we create several variants of E. coli promoter data sets and utilize them to experimentally examine the effect of these factors on the predictive performance of E. coli σ70 promoter models. Our results suggest that under some combinations of the first three criteria, a prediction model might perform very well on cross-validation experiments while its performance on independent test data is drastically very poor. This emphasizes the importance of evaluating promoter region predictors using independent test data, which corrects for the over-optimistic performance that might be estimated using the cross-validation procedure. Our analysis of the tested models shows that good prediction models often perform well despite how the non-promoter data was obtained. On the other hand, poor prediction models seems to be more sensitive to the choice of non-promoter sequences. Interestingly, the best performing sequence-based classifiers outperform the best performing structure-based classifiers on both cross-validation and independent test performance evaluation experiments. Finally, we propose a

Hypoxia regulates the expression of the neuromedin B receptor through a mechanism dependent on hypoxia-inducible factor-1α.

Directory of Open Access Journals (Sweden)

Hyun-Joo Park

Full Text Available The neuromedin B receptor (NMB-R, a member of the mammalian bombesin receptor family, is frequently overexpressed in various tumors. In the present study, we found that exposure to hypoxic conditions increases the levels of NMBR mRNA and protein in breast cancer cells, which are tightly regulated by hypoxia-inducible factor-1α (HIF-1α. We confirmed the effect of HIF-1α on NMBR transcription by performing an NMBR promoter-driven reporter assay and then identified a functional hypoxia-responsive element (HRE in the human NMBR promoter region. Further, the binding of HIF-1α to the NMBR promoter was corroborated by electrophoretic mobility shift and chromatin immunoprecipitation assays, which showed that HIF-1α specifically and directly bound to the NMBR promoter in response to hypoxia. Immunohistochemical analysis of a xenograft and a human breast cancer tissue array revealed a significant correlation between NMB-R and HIF-1α expression. Taken together, our findings indicate that hypoxia induces NMB-R expression through a novel mechanism to regulate HIF-1α expression in breast cancer cells.

Regional Cooperation Efforts in the Mekong River Basin: Mitigating river-related security threats and promoting regional development

Directory of Open Access Journals (Sweden)

Susanne Schmeier

2009-01-01

Full Text Available The development of international rivers is often perceived as leading to conflicts or even water wars. However, as the development of the Mekong River shows, cooperation has not only prevailed in the last decades, but River Basin Organizations (RBOs, established to mitigate river-related conflicts and/or develop the river basin, have also contributed to the emergence of more general cooperation structures, mainly by creating spill-over effects in other issue-areas, bringing cooperation to policy fields beyond the river itself. This article assesses the contribution of the Mekong River Commission (MRC and the Greater Mekong Sub-Region (GMS to the sustainable development of the Mekong Region as well as to the promotion of regional cooperation in mainland South-East Asia in general. --- Die Entwicklung grenzüberschreitender Flüsse wird oft mit Konflikten oder gar Kriegen um Wasser assoziiert. Wie jedoch die Entwicklung im Mekong-Becken zeigt, waren die vergangenen Jahrzehnte nicht nur von Kooperation gezeichnet, sondern Flussbeckenorganisationen konnten außerdem dazu beitragen, weitreichendere Kooperationsstrukturen zu entwickeln, die sich auf andere Politikfelder ausdehnen. Dieser Artikel beschäftigt sich mit dem Beitrag der Mekong River Commission (MRC und der Greater Mekong Sub-Region (GMS zur nachhaltigen Entwicklung in der Mekong Region sowie zur Förderung allgemeiner regionaler Kooperation im Festländischen Südostasien.

Mesotocin and nonapeptide receptors promote estrildid flocking behavior.

Science.gov (United States)

Goodson, James L; Schrock, Sara E; Klatt, James D; Kabelik, David; Kingsbury, Marcy A

2009-08-14

Proximate neural mechanisms that influence preferences for groups of a given size are almost wholly unknown. In the highly gregarious zebra finch (Estrildidae: Taeniopygia guttata), blockade of nonapeptide receptors by an oxytocin (OT) antagonist significantly reduced time spent with large groups and familiar social partners independent of time spent in social contact. Opposing effects were produced by central infusions of mesotocin (MT, avian homolog of OT). Most drug effects appeared to be female-specific. Across five estrildid finch species, species-typical group size correlates with nonapeptide receptor distributions in the lateral septum, and sociality in female zebra finches was reduced by OT antagonist infusions into the septum but not a control area. We propose that titration of sociality by MT represents a phylogenetically deep framework for the evolution of OT's female-specific roles in pair bonding and maternal functions.

Overexpression of the E2F target gene CENPI promotes chromosome instability and predicts poor prognosis in estrogen receptor-positive breast cancer.

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Thangavelu, Pulari U; Lin, Cheng-Yu; Vaidyanathan, Srividya; Nguyen, Thu H M; Dray, Eloise; Duijf, Pascal H G

2017-09-22

During cell division, chromosome segregation is facilitated by the mitotic checkpoint, or spindle assembly checkpoint (SAC), which ensures correct kinetochore-microtubule attachments and prevents premature sister-chromatid separation. It is well established that misexpression of SAC components on the outer kinetochores promotes chromosome instability (CIN) and tumorigenesis. Here, we study the expression of CENP-I, a key component of the HIKM complex at the inner kinetochores, in breast cancer, including ductal, lobular, medullary and male breast carcinomas. CENPI mRNA and protein levels are significantly elevated in estrogen receptor-positive (ER+) but not in estrogen receptor-negative (ER-) breast carcinoma. Well-established prognostic tests indicate that CENPI overexpression constitutes a powerful independent marker for poor patient prognosis and survival in ER+ breast cancer. We further demonstrate that CENPI is an E2F target gene. Consistently, it is overexpressed in RB1 -deficient breast cancers. However, CENP-I overexpression is not purely due to cell cycle-associated expression. In ER+ breast cancer cells, CENP-I overexpression promotes CIN, especially chromosome gains. In addition, in ER+ breast carcinomas the degree of CENPI overexpression is proportional to the level of aneuploidy and CENPI overexpression is one of the strongest markers for CIN identified to date. Our results indicate that overexpression of the inner kinetochore protein CENP-I promotes CIN and forecasts poor prognosis for ER+ breast cancer patients. These observations provide novel mechanistic insights and have important implications for breast cancer diagnostics and potentially therapeutic targeting.

Peroxisome proliferator-activated receptor gamma recruits the positive transcription elongation factor b complex to activate transcription and promote adipogenesis

DEFF Research Database (Denmark)

Iankova, Irena; Petersen, Rasmus K; Annicotte, Jean-Sébastien

2006-01-01

Positive transcription elongation factor b (P-TEFb) phosphorylates the C-terminal domain of RNA polymerase II, facilitating transcriptional elongation. In addition to its participation in general transcription, P-TEFb is recruited to specific promoters by some transcription factors such as c......-Myc or MyoD. The P-TEFb complex is composed of a cyclin-dependent kinase (cdk9) subunit and a regulatory partner (cyclin T1, cyclin T2, or cyclin K). Because cdk9 has been shown to participate in differentiation processes, such as muscle cell differentiation, we studied a possible role of cdk9...... with and phosphorylation of peroxisome proliferator-activated receptor gamma (PPARgamma), which is the master regulator of this process, on the promoter of PPARgamma target genes. PPARgamma-cdk9 interaction results in increased transcriptional activity of PPARgamma and therefore increased adipogenesis....

Modulating Estrogen Receptor-related Receptor-α Activity Inhibits Cell Proliferation*

Science.gov (United States)

Bianco, Stéphanie; Lanvin, Olivia; Tribollet, Violaine; Macari, Claire; North, Sophie; Vanacker, Jean-Marc

2009-01-01

High expression of the estrogen receptor-related receptor (ERR)-α in human tumors is correlated to a poor prognosis, suggesting an involvement of the receptor in cell proliferation. In this study, we show that a synthetic compound (XCT790) that modulates the activity of ERRα reduces the proliferation of various cell lines and blocks the G1/S transition of the cell cycle in an ERRα-dependent manner. XCT790 induces, in a p53-independent manner, the expression of the cell cycle inhibitor p21waf/cip1 at the protein, mRNA, and promoter level, leading to an accumulation of hypophosphorylated Rb. Finally, XCT790 reduces cell tumorigenicity in Nude mice. PMID:19546226

Leukemia inhibitory factor promote trophoblast invasion via urokinase-type plasminogen activator receptor in preeclampsia.

Science.gov (United States)

Zheng, Qin; Dai, Kuixing; Cui, Xinyuan; Yu, Ming; Yang, Xuesong; Yan, Bin; Liu, Shuai; Yan, Qiu

2016-05-01

Preeclampsia is a pregnancy-related syndrome which can cause perinatal mortality and morbidity. Inadequate invasion by trophoblast cells may lead to poor perfusion of the placenta, even result in preeclampsia. Understanding the molecular mechanisms underlying placentation facilitates the better intervention of preeclampsia. Urokinase-type plasminogen activator receptor (uPAR) is involved in the physiological and pathological processes. Leukemia inhibitory factor (LIF) is an important regulator in the establishment of pregnancy. However, the expression of uPAR in preeclamptic patients and its relationship with LIF remains unclear. In the current study, we found that the level of uPAR was relatively lower in the placentas from preeclamptic patients as compared with normal pregnant women. LIF promoted trophoblast cell outgrowth by upregulating uPAR in an explants culture, and LIF also enhanced migration and invasion potential through uPAR in trophoblast JAR and JEG-3 cell lines, and with increased gelatinolytic activities of matrix metalloproteinase 2 (MMP-2). The effect of LIF and uPAR on trophoblast migration and invasion was mediated by PI3K/AKT signaling pathway. Our data indicates the roles of LIF in promoting trophoblast migration and invasion through uPAR and suggest that abnormal expression of uPAR might be associated with the etiology of preeclampsia. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Regional specific binding of [11C]RO 15 1788 to central type benzodiazepine receptors in human brain: quantitative evaluation by PET

International Nuclear Information System (INIS)

Pappata, S.; Samson, Y.; Chavoix, C.; Prenant, C.; Maziere, M.; Baron, J.C.

1988-01-01

The central type benzodiazepine receptors were studied in 17 healthy human subjects with 11 C-RO 15 1788 and positron emission tomography (PET). The brain regional distribution of the tracer in eight control studies performed after injection of trace doses of 11 C-RO 15 1788 was consistent with that of benzodiazepine receptors. Saturation studies with co-injected cold RO 15 1788 in the remaining subjects showed a dose-dependent decrease of brain radiotracer until full inhibition of specific binding was achieved with doses above 0.1 mg/kg (four studies). Based on the results, a simple method to estimate the specifically bound 11 C-RO 15 1788 regionally in a single PET study is proposed, using the data from the full-saturation studies as a stable estimate of the nondisplaceable radioligand concentration. Using this method, it was found that quasiequilibrium between the estimated specifically bound and nondisplaceable components was achieved at times equal to or longer than 20 min after tracer administration. The validity of this method was partly supported by further results, showing a good agreement between the regional specific binding so calculated and postmortem data of receptor density

Androgen Receptor Signaling in Bladder Cancer

OpenAIRE

Li, Peng; Chen, Jinbo; Miyamoto, Hiroshi

2017-01-01

Emerging preclinical findings have indicated that steroid hormone receptor signaling plays an important role in bladder cancer outgrowth. In particular, androgen-mediated androgen receptor signals have been shown to correlate with the promotion of tumor development and progression, which may clearly explain some sex-specific differences in bladder cancer. This review summarizes and discusses the available data, suggesting the involvement of androgens and/or the androgen receptor pathways in u...

Identification of distal regulatory regions in the human alpha IIb gene locus necessary for consistent, high-level megakaryocyte expression.

Science.gov (United States)

Thornton, Michael A; Zhang, Chunyan; Kowalska, Maria A; Poncz, Mortimer

2002-11-15

The alphaIIb/beta3-integrin receptor is present at high levels only in megakaryocytes and platelets. Its presence on platelets is critical for hemostasis. The tissue-specific nature of this receptor's expression is secondary to the restricted expression of alphaIIb, and studies of the alphaIIb proximal promoter have served as a model of a megakaryocyte-specific promoter. We have examined the alphaIIb gene locus for distal regulatory elements. Sequence comparison between the human (h) and murine (m) alphaIIb loci revealed high levels of conservation at intergenic regions both 5' and 3' to the alphaIIb gene. Additionally, deoxyribonuclease (DNase) I sensitivity mapping defined tissue-specific hypersensitive (HS) sites that coincide, in part, with these conserved regions. Transgenic mice containing various lengths of the h(alpha)IIb gene locus, which included or excluded the various conserved/HS regions, demonstrated that the proximal promoter was sufficient for tissue specificity, but that a region 2.5 to 7.1 kb upstream of the h(alpha)IIb gene was necessary for consistent expression. Another region 2.2 to 7.4 kb downstream of the gene enhanced expression 1000-fold and led to levels of h(alpha)IIb mRNA that were about 30% of the native m(alpha)IIb mRNA level. These constructs also resulted in detectable h(alpha)IIb/m(beta)3 on the platelet surface. This work not only confirms the importance of the proximal promoter of the alphaIIb gene for tissue specificity, but also characterizes the distal organization of the alphaIIb gene locus and provides an initial localization of 2 important regulatory regions needed for the expression of the alphaIIb gene at high levels during megakaryopoiesis.

Methylation of the claudin 1 promoter is associated with loss of expression in estrogen receptor positive breast cancer.

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Francescopaolo Di Cello

Full Text Available Downregulation of the tight junction protein claudin 1 is a frequent event in breast cancer and is associated with recurrence, metastasis, and reduced survival, suggesting a tumor suppressor role for this protein. Tumor suppressor genes are often epigenetically silenced in cancer. Downregulation of claudin 1 via DNA promoter methylation may thus be an important determinant in breast cancer development and progression. To investigate if silencing of claudin 1 has an epigenetic etiology in breast cancer we compared gene expression and methylation data from 217 breast cancer samples and 40 matched normal samples available through the Cancer Genome Atlas (TCGA. Moreover, we analyzed claudin 1 expression and methylation in 26 breast cancer cell lines. We found that methylation of the claudin 1 promoter CpG island is relatively frequent in estrogen receptor positive (ER+ breast cancer and is associated with low claudin 1 expression. In contrast, the claudin 1 promoter was not methylated in most of the ER-breast cancers samples and some of these tumors overexpress claudin 1. In addition, we observed that the demethylating agents, azacitidine and decitabine can upregulate claudin 1 expression in breast cancer cell lines that have a methylated claudin 1 promoter. Taken together, our results indicate that DNA promoter methylation is causally associated with downregulation of claudin 1 in a subgroup of breast cancer that includes mostly ER+ tumors, and suggest that epigenetic therapy to restore claudin 1 expression might represent a viable therapeutic strategy in this subtype of breast cancer.

Interaction between retinoid acid receptor-related orphan receptor alpha (RORA and neuropeptide S receptor 1 (NPSR1 in asthma.

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Nathalie Acevedo

Full Text Available Retinoid acid receptor-related Orphan Receptor Alpha (RORA was recently identified as a susceptibility gene for asthma in a genome-wide association study. To investigate the impact of RORA on asthma susceptibility, we performed a genetic association study between RORA single nucleotide polymorphisms (SNPs in the vicinity of the asthma-associated SNP (rs11071559 and asthma-related traits. Because the regulatory region of a previously implicated asthma susceptibility gene, Neuropeptide S receptor 1 (NPSR1, has predicted elements for RORA binding, we hypothesized that RORA may interact biologically and genetically with NPSR1. 37 RORA SNPs and eight NPSR1 SNPs were genotyped in the Swedish birth cohort BAMSE (2033 children and the European cross-sectional PARSIFAL study (1120 children. Seven RORA SNPs confined into a 49 kb region were significantly associated with physician-diagnosed childhood asthma. The most significant association with rs7164773 (T/C was driven by the CC genotype in asthma cases (OR = 2.0, 95%CI 1.36-2.93, p = 0.0003 in BAMSE; and 1.61, 1.18-2.19, p = 0.002 in the combined BAMSE-PARSIFAL datasets, respectively, and strikingly, the risk effect was dependent on the Gln344Arg mutation in NPSR1. In cell models, stimulation of NPSR1 activated a pathway including RORA and other circadian clock genes. Over-expression of RORA decreased NPSR1 promoter activity further suggesting a regulatory loop between these genes. In addition, Rora mRNA expression was lower in the lung tissue of Npsr1 deficient mice compared to wildtype littermates during the early hours of the light period. We conclude that RORA SNPs are associated with childhood asthma and show epistasis with NPSR1, and the interaction between RORA and NPSR1 may be of biological relevance. Combinations of common susceptibility alleles and less common functional polymorphisms may modify the joint risk effects on asthma susceptibility.

Effects of swim stress and fluoxetine on 5-HT1A receptor gene expression and monoamine metabolism in the rat brain regions.

Science.gov (United States)

Shishkina, G T; Kalinina, T S; Dygalo, N N

2012-07-01

Changes in gene expression of the brain serotonin (5-HT) 1A receptors may be important for the development and ameliorating depression, however identification of specific stimuli that activate or reduce the receptor transcriptional activity is far from complete. In the present study, the forced swim test (FST) exposure, the first stress session of which is already sufficient to induce behavioral despair in rats, significantly increased 5-HT1A receptor mRNA levels in the brainstem, frontal cortex, and hippocampus at 24 h. In the brainstem and frontal cortex, the elevation in the receptor gene expression after the second forced swim session was not affected following chronic administration of fluoxetine, while in the cortex, both control and FST values were significantly reduced in fluoxetine-treated rats. In contrast to untreated rats, no increase in hippocampal 5-HT1A receptor mRNA was observed in response to FST in rats chronically treated with fluoxetine. Metabolism of 5-HT (5-HIAA/5-HT) in the brainstem was significantly decreased by fluoxetine and further reduced by swim stress, showing a certain degree of independence of these changes on 5-HT1A receptor gene expression that was increased in this brain region only after the FST, but not after fluoxetine. FST exposure also decreased the brainstem dopamine metabolism, which was unexpectedly positively correlated with 5-HT1A receptor mRNA levels in the frontal cortex. Together, these data suggest that the effects of the forced swim stress as well as fluoxetine involve brain region-dependent alterations in 5-HT1A receptor gene transcription, some of which may be interrelated with concomitant changes in catecholamine metabolism.

Graduate Management Project (GMP) Retrospective Analysis of Promotional Mediums for Tricare Prime in Tricare Region 11

National Research Council Canada - National Science Library

Carpenter, Steven

1997-01-01

This study provides retrospective market research information about the population who enrolled in TRICARE Prime in TRICARE Region 11 and the advertising mediums used to promote enrollment in the TRICARE Prime program...

Thermogenic characterization of ghrelin receptor null mice

Science.gov (United States)

Ghrelin is the only known circulating orexigenic hormone that increases food intake and promotes adiposity, and these physiological functions of ghrelin are mediated through its receptor growth hormone secretagogue receptor (GHS-R). Ghrelin/GHS-R signaling plays a crucial role in energy homeostasis....

Muscle Plasticity and β2-Adrenergic Receptors: Adaptive Responses of β2-Adrenergic Receptor Expression to Muscle Hypertrophy and Atrophy

OpenAIRE

Shogo Sato; Ken Shirato; Kaoru Tachiyashiki; Kazuhiko Imaizumi

2011-01-01

We discuss the functional roles of β2-adrenergic receptors in skeletal muscle hypertrophy and atrophy as well as the adaptive responses of β2-adrenergic receptor expression to anabolic and catabolic conditions. β2-Adrenergic receptor stimulation using anabolic drugs increases muscle mass by promoting muscle protein synthesis and/or attenuating protein degradation. These effects are prevented ...

β2-Adrenergic receptor promoter haplotype influences the severity of acute viral respiratory tract infection during infancy: a prospective cohort study.

Science.gov (United States)

Wu, Pingsheng; Larkin, Emma K; Reiss, Sara S; Carroll, Kecia N; Summar, Marshall L; Minton, Patricia A; Woodward, Kimberly B; Liu, Zhouwen; Islam, Jessica Y; Hartert, Tina V; Moore, Paul E

2015-09-14

Despite the significant interest in β2-Adrenergic receptor (ADRB2) polymorphisms related to asthma, whether ADRB2 genetic variants are similarly associated with acute respiratory tract infections have not been studied. We hypothesized that genetic variants in ADRB2 associated with a response to asthma therapy during an asthma exacerbation were also associated with severity of acute respiratory tract infections. To test this hypothesis, we genotyped 5 common polymorphisms in the promoter region and coding block of the ADRB2 gene (loci -2387, -2274, -1343, +46, and +79) from 374 Caucasian and African American term infants who were enrolled at the time of acute respiratory illness over four respiratory viral seasons. Severity of respiratory tract infections was measured using a bronchiolitis severity score (BSS; range = 0-12, clinically significant difference = 0.5) with a higher score indicating more severe disease. We assigned the promoter, coding and combined promoter and coding haplotypes to the unphased genotype data. The associations between each of these five single-nucleotide polymorphisms (SNPs) as well as the haplotypes and infant BSS were analyzed using nonparametric univariate analysis and multivariable proportional odds model separately in Caucasians and African Americans. There was no significant association between infant BSS and each of the SNPs in both Caucasians and African Americans. However, promoter haplotype CCA was associated with a decreased BSS in African Americans in a dose dependent manner. The median (interquartile range) BSS of infants with no copies of the CCA haplotype, one copy, and two copies of the CCA haplotype were 5.5 (2.0, 8.0), 4.0 (1.0, 7.5), and 3.0 (1.0, 4.0), respectively. This dose dependent relationship persisted after adjusting for infant age, gender, daycare exposure, secondhand smoke exposure, prior history of breastfeeding, siblings at home, and enrollment season (adjusted odds ratio: 0.59, 95% confidence

Extended hormone binding site of the human thyroid stimulating hormone receptor: distinctive acidic residues in the hinge region are involved in bovine thyroid stimulating hormone binding and receptor activation.

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Mueller, Sandra; Kleinau, Gunnar; Jaeschke, Holger; Paschke, Ralf; Krause, Gerd

2008-06-27

The human thyroid stimulating hormone receptor (hTSHR) belongs to the glycoprotein hormone receptors that bind the hormones at their large extracellular domain. The extracellular hinge region of the TSHR connects the N-terminal leucine-rich repeat domain with the membrane-spanning serpentine domain. From previous studies we reasoned that apart from hormone binding at the leucine-rich repeat domain, additional multiple hormone contacts might exist at the hinge region of the TSHR by complementary charge-charge recognition. Here we investigated highly conserved charged residues in the hinge region of the TSHR by site-directed mutagenesis to identify amino acids interacting with bovine TSH (bTSH). Indeed, the residues Glu-297, Glu-303, and Asp-382 in the TSHR hinge region are essential for bTSH binding and partially for signal transduction. Side chain substitutions showed that the negative charge of Glu-297 and Asp-382 is necessary for recognition of bTSH by the hTSHR. Multiple combinations of alanine mutants of the identified positions revealed an increased negative effect on hormone binding. An assembled model suggests that the deciphered acidic residues form negatively charged patches at the hinge region resulting in an extended binding mode for bTSH on the hTSHR. Our data indicate that certain positively charged residues of bTSH might be involved in interaction with the identified negatively charged amino acids of the hTSHR hinge region. We demonstrate that the hinge region represents an extracellular intermediate connector for both hormone binding and signal transduction of the hTSHR.

Neurotrophin receptors expression and JNK pathway activation in human astrocytomas

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Maraziotis Theodore

2007-10-01

significantly co-expressed in a tumor grade-dependent manner (p Conclusion In the context of astrocytomas, Trk receptors (TrkA, TrkB, TrkC expression may promote tumor growth independently of grade. Furthermore, activation of JNK pathway may contribute to progression towards malignancy. Considering the fact that regional tumor heterogeneity may be a limiting factor for immunohistochemical studies, the significance of the reverse relationship between Trk receptors and pc-Jun/pJNK LIs with respect to biological behavior of human astrocytomas requires further evaluation.

Orphan Nuclear Receptor ERRα Controls Macrophage Metabolic Signaling and A20 Expression to Negatively Regulate TLR-Induced Inflammation.

Science.gov (United States)

Yuk, Jae-Min; Kim, Tae Sung; Kim, Soo Yeon; Lee, Hye-Mi; Han, Jeongsu; Dufour, Catherine Rosa; Kim, Jin Kyung; Jin, Hyo Sun; Yang, Chul-Su; Park, Ki-Sun; Lee, Chul-Ho; Kim, Jin-Man; Kweon, Gi Ryang; Choi, Hueng-Sik; Vanacker, Jean-Marc; Moore, David D; Giguère, Vincent; Jo, Eun-Kyeong

2015-07-21

The orphan nuclear receptor estrogen-related receptor α (ERRα; NR3B1) is a key metabolic regulator, but its function in regulating inflammation remains largely unknown. Here, we demonstrate that ERRα negatively regulates Toll-like receptor (TLR)-induced inflammation by promoting Tnfaip3 transcription and fine-tuning of metabolic reprogramming in macrophages. ERRα-deficient (Esrra(-/-)) mice showed increased susceptibility to endotoxin-induced septic shock, leading to more severe pro-inflammatory responses than control mice. ERRα regulated macrophage inflammatory responses by directly binding the promoter region of Tnfaip3, a deubiquitinating enzyme in TLR signaling. In addition, Esrra(-/-) macrophages showed an increased glycolysis, but impaired mitochondrial respiratory function and biogenesis. Further, ERRα was required for the regulation of NF-κB signaling by controlling p65 acetylation via maintenance of NAD(+) levels and sirtuin 1 activation. These findings unravel a previously unappreciated role for ERRα as a negative regulator of TLR-induced inflammatory responses through inducing Tnfaip3 transcription and controlling the metabolic reprogramming. Copyright © 2015 Elsevier Inc. All rights reserved.

TARM1 Is a Novel Leukocyte Receptor Complex-Encoded ITAM Receptor That Costimulates Proinflammatory Cytokine Secretion by Macrophages and Neutrophils

DEFF Research Database (Denmark)

Radjabova, Valeria; Mastroeni, Piero; Skjødt, Karsten

2015-01-01

We identified a novel, evolutionarily conserved receptor encoded within the human leukocyte receptor complex and syntenic region of mouse chromosome 7, named T cell-interacting, activating receptor on myeloid cells-1 (TARM1). The transmembrane region of TARM1 contained a conserved arginine residu...

A developmental timing switch promotes axon outgrowth independent of known guidance receptors.

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Katherine Olsson-Carter

2010-08-01

Full Text Available To form functional neuronal connections, axon outgrowth and guidance must be tightly regulated across space as well as time. While a number of genes and pathways have been shown to control spatial features of axon development, very little is known about the in vivo mechanisms that direct the timing of axon initiation and elongation. The Caenorhabditis elegans hermaphrodite specific motor neurons (HSNs extend a single axon ventrally and then anteriorly during the L4 larval stage. Here we show the lin-4 microRNA promotes HSN axon initiation after cell cycle withdrawal. Axons fail to form in lin-4 mutants, while they grow prematurely in lin-4-overexpressing animals. lin-4 is required to down-regulate two inhibitors of HSN differentiation--the transcriptional regulator LIN-14 and the "stemness" factor LIN-28--and it likely does so through a cell-autonomous mechanism. This developmental switch depends neither on the UNC-40/DCC and SAX-3/Robo receptors nor on the direction of axon growth, demonstrating that it acts independently of ventral guidance signals to control the timing of HSN axon elongation.

Sequencing of DC-SIGN promoter indicates an association between promoter variation and risk of nasopharyngeal carcinoma in cantonese

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Liu Wen-Sheng

2010-11-01

Full Text Available Abstract Background The dendritic cell-specific intercellular adhesion molecule 3 grabbing non-integrin (DC-SIGN is an important pathogen recognition receptor of the innate immune system. DC-SIGN promoter variants play important role in the susceptibility to various infectious diseases. Nasopharyngeal carcinoma (NPC is a malignancy that is common in southern China and whether DC-SIGN promoter variants have effects on susceptibility to NPC is still unknown. The aim of this study is to ascertain the potential involvement of DC-SIGN promoter single nucleotide polymorphisms (SNPs in NPC susceptibility. Methods We conducted a case control study based on Cantonese population including 444 NPC patients and 464 controls matched on age and sex. The 1041 bp of DC-SIGN promoter region was directly sequenced for all samples. Sequence alignment and SNP search were inspected using DNAStar analysis programs and haplotype frequencies were estimated in Haploview V 4.0. The associations between the SNPs and the risk of NPC were analyzed using chi-square test and non-conditional logistic regression analysis with SPSS 13.0 software. Results A total of six variants were observed in the DC-SIGN promoter region and DC-SIGN -139 GG and -939 AA were significantly associated with NPC risk with adjusted Odds Ratios (ORs of 2.10 (95% confidence interval [CI] = 1.23-3.59; P = 0.006 and 2.52 (1.29-4.93; P = 0.007 respectively and subjects carrying the risk allele DC-SIGN -871 G had 1.47-fold (95% CI = 1.14-1.90 increased risks of developing NPC (P = 0.003. Haplotype analysis revealed that h1 'AAAG' was significantly associated with protection against NPC (OR = 0.69; P = 0.0002 and the association was still significant when using 1000 permutation test runs (P = 0.001. Conclusions Our study indicated that DC-SIGN promoter variants appear to be involved in the susceptibility to NPC and the detailed mechanism of this effect need further studies.

Targets of DNA-binding proteins in bacterial promoter regions present enhanced probabilities for spontaneous thermal openings

International Nuclear Information System (INIS)

Apostolaki, Angeliki; Kalosakas, George

2011-01-01

We mapped promoter regions of double-stranded DNA with respect to the probabilities of appearance of relatively large bubble openings exclusively due to thermal fluctuations at physiological temperatures. We analyzed five well-studied promoter regions of procaryotic type and found a spatial correlation between the binding sites of transcription factors and the position of peaks in the probability pattern of large thermal openings. Other distinct peaks of the calculated patterns correlate with potential binding sites of DNA-binding proteins. These results suggest that a DNA molecule would more frequently expose the bases that participate in contacts with proteins, which would probably enhance the probability of the latter to reach their targets. It also stands for using this method as a means to analyze DNA sequences based on their intrinsic thermal properties

Transcription control and neuronal differentiation by agents that activate the LXR nuclear receptor family.

Science.gov (United States)

Schmidt, A; Vogel, R; Holloway, M K; Rutledge, S J; Friedman, O; Yang, Z; Rodan, G A; Friedman, E

1999-09-10

LXR and PPAR receptors belong to the nuclear receptor superfamily of transcriptional activating factors. Using ligand-dependent transcription assays, we found that 5-tetradecyloxy-2-furancarboxylic acid (TOFA) transactivates chimeric receptors composed of the glucocorticoid receptor DNA binding domain and the ligand binding regions of PPARalpha, PPARbeta (NUC-1) and LXRbeta (NER) receptors. In the same assays, ligands for PPARs (oleic acid, WY-14643 and L-631,033) and LXRs (hydroxycholesterols) maintain their respective receptor selectivity. TOFA and hydroxycholesterols also stimulate transcription from a minimal fibrinogen promoter that is under the control of AP-1 or NF-kappaB transcription factor binding sites. In addition to their effects on transcription, these LXRbeta activators induce neuronal differentiation in rat pheochromocytoma cells. TOFA and the natural LXR agonist, 22 (R)-hydroxycholesterol, stimulate neurite outgrowth in 55 and 28% of cells, respectively. No neurite outgrowth was induced by the related 22(S)-hydroxycholesterol, which does not activate the LXR family. These results suggest that the hydroxycholesterol signaling pathway has a complex effect on transcription that mediates the activity of TOFA and hydroxycholesterol on neuronal differentiation in pheochromocytoma cells.

GABA-A Receptors Mediate Tonic Inhibition and Neurosteroid Sensitivity in the Brain.

Science.gov (United States)

Reddy, Doodipala Samba

2018-01-01

Neurosteroids like allopregnanolone (AP) are positive allosteric modulators of synaptic and extrasynaptic GABA-A receptors. AP and related neurosteroids exhibit a greater potency for δ-containing extrasynaptic receptors. The δGABA-A receptors, which are expressed extrasynaptically in the dentate gyrus and other regions, contribute to tonic inhibition, promoting network shunting as well as reducing seizure susceptibility. Levels of endogenous neurosteroids fluctuate with ovarian cycle. Natural and synthetic neurosteroids maximally potentiate tonic inhibition in the hippocampus and provide robust protection against a variety of limbic seizures and status epilepticus. Recently, a consensus neurosteroid pharmacophore model has been proposed at extrasynaptic δGABA-A receptors based on structure-activity relationship for functional activation of tonic currents and seizure protection. Aside from anticonvulsant actions, neurosteroids have been found to be powerful anxiolytic and anesthetic agents. Neurosteroids and Zn 2+ have preferential affinity for δ-containing receptors. Thus, Zn 2+ can prevent neurosteroid activation of extrasynaptic δGABA-A receptor-mediated tonic inhibition. Recently, we demonstrated that Zn 2+ selectively inhibits extrasynaptic δGABA-A receptors and thereby fully prevents AP activation of tonic inhibition and seizure protection. We confirmed that neurosteroids exhibit greater sensitivity at extrasynaptic δGABA-A receptors. Overall, extrasynaptic GABA-A receptors are primary mediators of tonic inhibition in the brain and play a key role in the pathophysiology of epilepsy and other neurological disorders. © 2018 Elsevier Inc. All rights reserved.

The role of GH receptor tyrosine phosphorylation in Stat5 activation

DEFF Research Database (Denmark)

Hansen, J A; Hansen, L H; Wang, X

1997-01-01

Stimulation of GH receptors leads to rapid activation of Jak2 kinase and subsequent tyrosine phosphorylation of the GH receptor. Three specific tyrosines located in the C-terminal domain of the GH receptor have been identified as being involved in GH-stimulated transcription of the Spi 2.1 promoter....... Mutated GH receptors lacking all but one of these three tyrosines are able to mediate a transcriptional response when transiently transfected into CHO cells together with a Spi 2.1 promoter/luciferase construct. Similarly, these GH receptors were found to be able to mediate activation of Stat5 DNA......-binding activity, whereas the GH receptor mutant lacking all intracellular tyrosines was not. Synthetic tyrosine phosphorylated peptides corresponding to the GH receptor sequence around the three tyrosines inhibited Stat5 DNA-binding activity while their non-phosphorylated counterparts were ineffective. Tyrosine...

Topical administration of adrenergic receptor pharmaceutics and nerve growth factor

Directory of Open Access Journals (Sweden)

Jena J Steinle

2010-06-01

Full Text Available Jena J SteinleDepartments of Ophthalmology and Anatomy and Neurobiology, Hamilton Eye Institute, University of Tennessee Health Science Center, Memphis, TN 38163, USAAbstract: Topical application of nerve growth factor (NGF and adrenergic receptor pharmaceutics are currently in use for corneal ulcers and glaucoma. A recent interest in the neuroprotective abilities of NGF has led to a renewed interest in NGF as a therapeutic for retinal and choroidal diseases. NGF can promote cell proliferation through actions of the TrkA receptor or promote apoptosis through receptor p75NTR. This understanding has led to novel interest in the role of NGF for diseases of the posterior eye. The role of β-adrenergic receptor agonists and antagonists for treatments of glaucoma, diabetic retinopathy, and their potential mechanisms of action, are still under investigation. This review discusses the current knowledge and applications of topical NGF and adrenergic receptor drugs for ocular disease.Keywords: NGF, β-adrenergic receptor agents, α-adrenergic receptor agents, retina, cornea, glaucoma

Distribution of seratonin-1A receptors in the monkey and the postmortem human hippocampal region. A quantitative autoradiographic study using the selective agonist (3H)8-PH-DPAT

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Koehler, Christer; Radesaeter, A.C.; Lang, Walter; Chan-Palay, Victoria

1986-01-01

Serotonin-1A receptors were visualized and their anatomical distribution mapped within the monkey and the human hippocampus by using in vitro receptor autoradiography of the selective agonist ( 3 H)8-OH-N, N-dipropyl-2-aminotetralin (( 3 H)8-OH-DPAT). The results show high densities of serotonin-1A receptors hetergeneously distributed in different subfields and layers of the monkey and the human hippocampal region. High densities are found in the molecular layer of area dentata, all layers of regio superior and the subiculum, parasubiculum, and layers 2, and 4 through 6 of the entorhinal area. In the human hippocampus, a distinct band of ( 3 H)8-OH-DPAT binding sites is present in the subgranular zone of the area dentata. The similar anatomical distribution of ( 3 H)8-OH-DPAT binding sites in the monkey and the human hippocampal region suggests that the serotonin-1A receptor is phylogenetically well preserved and indicates that this receptor may mediate action(s) of serotonin in the primate, including the human hippocampal region. (author)

Region-specific associations between sex, social status, and oxytocin receptor density in the brains of eusocial rodents.

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Mooney, S J; Coen, C W; Holmes, M M; Beery, A K

2015-09-10

Naturally occurring variations in neuropeptide receptor distributions in the brain contribute to numerous mammalian social behaviors. In naked mole-rats, which live in large social groups and exhibit remarkable reproductive skew, colony-related social behaviors vary with reproductive status. Here we examined whether variation in social status is associated with variations in the location and/or density of oxytocin binding in this species. Autoradiography was performed to assess forebrain oxytocin receptor (OTR) densities in breeding and non-breeding naked mole-rats of both sexes. Overall, males exhibited higher OTR binding in the medial amygdala in comparison to females. While there were no main effects of reproductive status in any region, a sex difference in OTR binding in the nucleus accumbens was mediated by status. Specifically, breeding males tended to have more OTR binding than breeding females in the nucleus accumbens, while no sex difference was observed in subordinates. These effects suggest that oxytocin may act in a sex- and region-specific way that corresponds to reproductive status and associated social behaviors. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

Transcriptional targets shared by estrogen receptor- related receptors (ERRs) and estrogen receptor (ER) alpha, but not by ERbeta.

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Vanacker, J M; Pettersson, K; Gustafsson, J A; Laudet, V

1999-01-01

The physiological activities of estrogens are thought to be mediated by specific nuclear receptors, ERalpha and ERbeta. However, certain tissues, such as the bone, that are highly responsive to estrogens only express a low level of these receptors. Starting from this apparent contradiction, we have evaluated the potentials of two related receptors ERRalpha and ERRbeta to intervene in estrogen signaling. ERalpha, ERRalpha and ERRbeta bind to and activate transcription through both the classical estrogen response element (ERE) and the SF-1 response element (SFRE). In contrast, ERbeta DNA-binding and transcriptional activity is restricted to the ERE. Accordingly, the osteopontin gene promoter is stimulated through SFRE sequences, by ERRalpha as well as by ERalpha, but not by ERbeta. Analysis of the cross-talk within the ER/ERR subgroup of nuclear receptors thus revealed common targets but also functional differences between the two ERs. PMID:10428965

Sonic Hedgehog promotes the survival of neural crest cells by limiting apoptosis induced by the dependence receptor CDON during branchial arch development.

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Delloye-Bourgeois, Céline; Rama, Nicolas; Brito, José; Le Douarin, Nicole; Mehlen, Patrick

2014-09-26

Cell-adhesion molecule-related/Downregulated by Oncogenes (CDO or CDON) was identified as a receptor for the classic morphogen Sonic Hedgehog (SHH). It has been shown that, in cell culture, CDO also behaves as a SHH dependence receptor: CDO actively triggers apoptosis in absence of SHH via a proteolytic cleavage in CDO intracellular domain. We present evidence that CDO is also pro-apoptotic in the developing neural tube where SHH is known to act as a survival factor. SHH, produced by the ventral foregut endoderm, was shown to promote survival of facial neural crest cells (NCCs) that colonize the first branchial arch (BA1). We show here that the survival activity of SHH on neural crest cells is due to SHH-mediated inhibition of CDO pro-apoptotic activity. Silencing of CDO rescued NCCs from apoptosis observed upon SHH inhibition in the ventral foregut endoderm. Thus, the pair SHH/dependence receptor CDO may play an important role in neural crest cell survival during the formation of the first branchial arch. Copyright © 2014 Elsevier Inc. All rights reserved.

P2X7, NMDA and BDNF receptors converge on GSK3 phosphorylation and cooperate to promote survival in cerebellar granule neurons.

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Ortega, Felipe; Pérez-Sen, Raquel; Morente, Verónica; Delicado, Esmerilda G; Miras-Portugal, Maria Teresa

2010-05-01

Glycogen synthase kinase-3 (GSK3) is a key player in the regulation of neuronal survival. Herein, we report evidence of an interaction between P2X7 receptors with NMDA and BDNF receptors at the level of GSK3 signalling and neuroprotection. The activation of these receptors in granule neurons led to a sustained pattern of GSK3 phosphorylation that was mainly PKC-dependent. BDNF was the most potent at inducing GSK3 phosphorylation, which was also dependent on PI3K. The P2X7 agonist, BzATP, exhibited additive effects with both NMDA and BDNF to rescue granule neurons from cell death induced by PI3K inhibition. This survival effect was mediated by the PKC-dependent GSK3 pathway. In addition, ERK1/2 proteins were also involved in BDNF protective effect. These results show the function of ATP in amplifying neuroprotective actions of glutamate and neurotrophins, and support the role of GSK3 as an important convergence point for these survival promoting factors in granule neurons.

Honokiol promotes non-rapid eye movement sleep via the benzodiazepine site of the GABA(A) receptor in mice.

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Qu, Wei-Min; Yue, Xiao-Fang; Sun, Yu; Fan, Kun; Chen, Chang-Rui; Hou, Yi-Ping; Urade, Yoshihiro; Huang, Zhi-Li

2012-10-01

Decoctions of the Chinese herb houpu contain honokiol and are used to treat a variety of mental disorders, including depression. Depression commonly presents alongside sleep disorders and sleep disturbances, which appear to be a major risk factor for depression. Here, we have evaluated the somnogenic effect of honokiol and the mechanisms involved. Honokiol was administered i.p. at 20:00 h in mice. Flumazenil, an antagonist at the benzodiazepine site of the GABA(A) receptor, was administered i.p. 15 min before honokiol. The effects of honokiol were measured by EEG and electromyogram (EMG), c-Fos expression and in vitro electrophysiology. Honokiol (10 and 20 mg·kg⁻¹) significantly shortened the sleep latency to non-rapid eye movement (non-REM, NREM) sleep and increased the amount of NREM sleep. Honokiol increased the number of state transitions from wakefulness to NREM sleep and, subsequently, from NREM sleep to wakefulness. However, honokiol had no effect on either the amount of REM sleep or EEG power density of both NREM and REM sleep. Honokiol increased c-Fos expression in ventrolateral preoptic area (VLPO) neurons, as examined by immunostaining, and excited sleep-promoting neurons in the VLPO by whole-cell patch clamping in the brain slice. Pretreatment with flumazenil abolished the somnogenic effects and activation of the VLPO neurons by honokiol. Honokiol promoted NREM sleep by modulating the benzodiazepine site of the GABA(A) receptor, suggesting potential applications in the treatment of insomnia, especially for patients who experience difficulty in falling and staying asleep. © 2012 The Authors. British Journal of Pharmacology © 2012 The British Pharmacological Society.

A distal region of the human TGM1 promoter is required for expression in transgenic mice and cultured keratinocytes

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Lu Ying

2004-04-01

Full Text Available Abstract Background TGM1(transglutaminase 1 is an enzyme that crosslinks the cornified envelope of mature keratinocytes. Appropriate expression of the TGM1 gene is crucial for proper keratinocyte function as inactivating mutations lead to the debilitating skin disease, lamellar ichthyosis. TGM1 is also expressed in squamous metaplasia, a consequence in some epithelia of vitamin A deficiency or toxic insult that can lead to neoplasia. An understanding of the regulation of this gene in normal and abnormal differentiation states may contribute to better disease diagnosis and treatment. Methods In vivo requirements for expression of the TGM1 gene were studied by fusing various lengths of promoter DNA to a reporter and injecting the DNA into mouse embryos to generate transgenic animals. Expression of the reporter was ascertained by Western blotting and immunohistochemistry. Further delineation of a transcriptionally important distal region was determined by transfections of progressively shortened or mutated promoter DNA into cultured keratinocytes. Results In vivo analysis of a reporter transgene driven by the TGM1 promoter revealed that 1.6 kilobases, but not 1.1 kilobases, of DNA was sufficient to confer tissue-specific and cell layer-specific expression. This same region was responsible for reporter expression in tissues undergoing squamous metaplasia as a response to vitamin A deprivation. Mutation of a distal promoter AP1 site or proximal promoter CRE site, both identified as important transcriptional elements in transfection assays, did not prevent appropriate expression. Further searching for transcriptional elements using electrophoretic mobility shift (EMSA and transfection assays in cultured keratinocytes identified two Sp1 elements in a transcriptionally active region between -1.6 and -1.4 kilobases. While mutation of either Sp1 site or the AP1 site singly had only a small effect, mutation of all three sites eliminated nearly all the

Functionally biased signalling properties of 7TM receptors - opportunities for drug development for the ghrelin receptor

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Sivertsen, B; Holliday, N; Madsen, A N

2013-01-01

UNLABELLED: The ghrelin receptor is a 7 transmembrane (7TM) receptor involved in a variety of physiological functions including growth hormone secretion, increased food intake and fat accumulation as well as modulation of reward and cognitive functions. Because of its important role in metabolism...... and energy expenditure, the ghrelin receptor has become an important therapeutic target for drug design and the development of anti-obesity compounds. However, none of the compounds developed so far have been approved for commercial use. Interestingly, the ghrelin receptor is able to signal through several...... review, we have described how ligands and mutations in the 7TM receptor may bias the receptors to favour either one G-protein over another or to promote G-protein independent signalling pathways rather than G-protein-dependent pathways. For the ghrelin receptor, both agonist and inverse agonists have...

Triclocarban mediates induction of xenobiotic metabolism through activation of the constitutive androstane receptor and the estrogen receptor alpha.

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Mei-Fei Yueh

Full Text Available Triclocarban (3,4,4'-trichlorocarbanilide, TCC is used as a broad-based antimicrobial agent that is commonly added to personal hygiene products. Because of its extensive use in the health care industry and resistance to degradation in sewage treatment processes, TCC has become a significant waste product that is found in numerous environmental compartments where humans and wildlife can be exposed. While TCC has been linked to a range of health and environmental effects, few studies have been conducted linking exposure to TCC and induction of xenobiotic metabolism through regulation by environmental sensors such as the nuclear xenobiotic receptors (XenoRs. To identify the ability of TCC to activate xenobiotic sensors, we monitored XenoR activities in response to TCC treatment using luciferase-based reporter assays. Among the XenoRs in the reporter screening assay, TCC promotes both constitutive androstane receptor (CAR and estrogen receptor alpha (ERα activities. TCC treatment to hUGT1 mice resulted in induction of the UGT1A genes in liver. This induction was dependent upon the constitutive active/androstane receptor (CAR because no induction occurred in hUGT1Car(-/- mice. Induction of the UGT1A genes by TCC corresponded with induction of Cyp2b10, another CAR target gene. TCC was demonstrated to be a phenobarbital-like activator of CAR in receptor-based assays. While it has been suggested that TCC be classified as an endocrine disruptor, it activates ERα leading to induction of Cyp1b1 in female ovaries as well as in promoter activity. Activation of ERα by TCC in receptor-based assays also promotes induction of human CYP2B6. These observations demonstrate that TCC activates nuclear xenobiotic receptors CAR and ERα both in vivo and in vitro and might have the potential to alter normal physiological homeostasis. Activation of these xenobiotic-sensing receptors amplifies gene expression profiles that might represent a mechanistic base for

Down syndrome critical region 2 protein inhibits the transcriptional activity of peroxisome proliferator-activated receptor β in HEK293 cells

International Nuclear Information System (INIS)

Song, Hae Jin; Park, Joongkyu; Seo, Su Ryeon; Kim, Jongsun; Paik, Seung R.; Chung, Kwang Chul

2008-01-01

Down syndrome is mainly caused by a trisomy of chromosome 21. The Down syndrome critical region 2 (DSCR2) gene is located within a part of chromosome 21, the Down syndrome critical region (DSCR). To investigate the function of DSCR2, we sought to identify DSCR2-interacting proteins using yeast two-hybrid assays. A human fetal brain cDNA library was screened, and DSCR2 was found to interact with a member of the nuclear receptor superfamily, peroxisome proliferator-activated receptor β, (PPARβ). A co-immunoprecipitation assay demonstrated that DSCR2 physically interacts with PPARβ in mammalian HEK293 cells. DSCR2 also inhibited the ligand-induced transcriptional activity of PPARβ. Furthermore, PPARβ also decreased the solubility of DSCR2, which increased levels of insoluble DSCR2

Discovery and Evaluation of Polymorphisms in the and Promoter Regions for Risk of Korean Lung Cancer

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Jae Sook Sung

2012-09-01

Full Text Available AKT is a signal transduction protein that plays a central role in the tumorigenesis. There are 3 mammalian isoforms of this serine/threonine protein kinase-AKT1, AKT2, and AKT3-showing a broad tissue distribution. We first discovered 2 novel polymorphisms (AKT2 -9826 C/G and AKT3 -811 A/G, and we confirmed 6 known polymorphisms (AKT2 -9473 C/T, AKT2 -9151 C/T, AKT2 -9025 C/T, AKT2 -8618G/A, AKT3 -675 A/-, and AKT3 -244 C/T of the AKT2 and AKT3 promoter region in 24 blood samples of Korean lung cancer patients using direct sequencing. To evaluate the role of AKT2 and AKT3 polymorphisms in the risk of Korean lung cancer, genotypes of the AKT2 and AKT3 polymorphisms (AKT2 -9826 C/G, AKT2 -9473 C/T, AKT2 -9151 C/T, AKT2 -9025 C/T, AKT2 -8618G/A, and AKT3 -675 A/- were determined in 360 lung cancer patients and 360 normal controls. Statistical analyses revealed that the genotypes and haplotypes in the AKT2 and AKT3 promoter regions were not significantly associated with the risk of lung cancer in the Korean population. These results suggest that polymorphisms of the AKT2 and AKT3 promoter regions do not contribute to the genetic susceptibility to lung cancer in the Korean population.

Impact of cell type and epitope tagging on heterologous expression of G protein-coupled receptor: a systematic study on angiotensin type II receptor.

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Lili Jiang

Full Text Available Despite heterologous expression of epitope-tagged GPCR is widely adopted for functional characterization, there is lacking of systematic analysis of the impact of expression host and epitope tag on GPCR expression. Angiotensin type II (AT2 receptor displays agonist-dependent and -independent activities, coupling to a spectrum of signaling molecules. However, consensus has not been reached on the subcellular distributions, signaling cascades and receptor-mediated actions. To examine the contributions of host cell and epitope tag on receptor expression and activity, epitope-tagged AT2 receptor variants were transiently or stably expressed in HEK293, CHO-K1 and PC12 cells. The epitope-tagged AT2 receptor variants were detected both on the cell membrane and in the perinuclear region. In transiently transfected HEK293 cells, Myc-AT2 existed predominantly as monomer. Additionally, a ladder of ubiquitinated AT2 receptor proteins was detected. By contrast, stably expressed epitope-tagged AT2 receptor variants existed as both monomer and high molecular weight complexes, and the latter was enriched in cell surface. Glycosylation promoted cell surface expression of Myc-AT2 but had no effect on AT2-GFP in HEK293 cells. In cells that stably expressed Myc-AT2, serum starvation induced apoptosis in CHO-K1 cells but not in HEK293 or PC12 cells. Instead, HEK293 and PC12 cells stably expressing Myc-AT2 exhibited partial cell cycle arrest with cells accumulating at G1 and S phases, respectively. Taken together, these results suggest that expression levels, subcellular distributions and ligand-independent constitutive activities of AT2 receptor were cell type-dependent while posttranslational processing of nascent AT2 receptor protein was modulated by epitope tag and mode of expression.

Sensitivity of MLL-rearranged AML cells to all-trans retinoic acid is associated with the level of H3K4me2 in the RARα promoter region

International Nuclear Information System (INIS)

Sakamoto, K; Imamura, T; Yano, M; Yoshida, H; Fujiki, A; Hirashima, Y; Hosoi, H

2014-01-01

All-trans retinoic acid (ATRA) is well established as differentiation therapy for acute promyelocytic leukemia (APL) in which the PML–RARα (promyelocytic leukemia-retinoic acid receptor α) fusion protein causes blockade of the retinoic acid (RA) pathway; however, in types of acute myeloid leukemia (AML) other than APL, the mechanism of RA pathway inactivation is not fully understood. This study revealed the potential mechanism of high ATRA sensitivity of mixed-lineage leukemia (MLL)-AF9-positive AML compared with MLL-AF4/5q31-positive AML. Treatment with ATRA induced significant myeloid differentiation accompanied by upregulation of RARα, C/EBPα, C/EBPε and PU.1 in MLL-AF9-positive but not in MLL-AF4/5q31-positive cells. Combining ATRA with cytarabine had a synergistic antileukemic effect in MLL-AF9-positive cells in vitro. The level of dimethyl histone H3 lysine 4 (H3K4me2) in the RARα gene-promoter region, PU.1 upstream regulatory region (URE) and RUNX1+24/+25 intronic enhancer was higher in MLL-AF9-positive cells than in MLL-AF4-positive cells, and inhibiting lysine-specific demethylase 1, which acts as a histone demethylase inhibitor, reactivated ATRA sensitivity in MLL-AF4-positive cells. These findings suggest that the level of H3K4me2 in the RARα gene-promoter region, PU.1 URE and RUNX1 intronic enhancer is determined by the MLL-fusion partner. Our findings provide insight into the mechanisms of ATRA sensitivity in AML and novel treatment strategies for ATRA-resistant AML

RAGE, receptor of advanced glycation endoproducts, negatively regulates chondrocytes differentiation.

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Tatsuya Kosaka

Full Text Available RAGE, receptor for advanced glycation endoproducts (AGE, has been characterized as an activator of osteoclastgenesis. However, whether RAGE directly regulates chondrocyte proliferation and differentiation is unclear. Here, we show that RAGE has an inhibitory role in chondrocyte differentiation. RAGE expression was observed in chondrocytes from the prehypertrophic to hypertrophic regions. In cultured cells, overexpression of RAGE or dominant-negative-RAGE (DN-RAGE demonstrated that RAGE inhibited cartilaginous matrix production, while DN-RAGE promoted production. Additionally, RAGE regulated Ihh and Col10a1 negatively but upregulated PTHrP receptor. Ihh promoter analysis and real-time PCR analysis suggested that downregulation of Cdxs was the key for RAGE-induced inhibition of chondrocyte differentiation. Overexpression of the NF-κB inhibitor I-κB-SR inhibited RAGE-induced NF-κB activation, but did not influence inhibition of cartilaginous matrix production by RAGE. The inhibitory action of RAGE was restored by the Rho family GTPases inhibitor Toxin B. Furthermore, inhibitory action on Ihh, Col10a1 and Cdxs was reproduced by constitutively active forms, L63RhoA, L61Rac, and L61Cdc42, but not by I-κB-SR. Cdx1 induced Ihh and Col10a1 expressions and directly interacted with Ihh promoter. Retinoic acid (RA partially rescued the inhibitory action of RAGE. These data combined suggests that RAGE negatively regulates chondrocyte differentiation at the prehypertrophic stage by modulating NF-κB-independent and Rho family GTPases-dependent mechanisms.

Regional Differences in Correlates of Daily Walking among Middle Age and Older Australian Rural Adults: Implications for Health Promotion

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James Dollman

2016-01-01

Full Text Available Rural Australians are less physically active than their metropolitan counterparts, and yet very little is known of the candidate intervention targets for promoting physical activity in rural populations. As rural regions are economically, socially and environmentally diverse, drivers of regular physical activity are likely to vary between regions. This study explored the region-specific correlates of daily walking among middle age and older adults in rural regions with contrasting dominant primary industries. Participants were recruited through print and electronic media, primary care settings and community organisations. Pedometers were worn by 153 adults for at least four days, including a weekend day. A questionnaire identified potential intra-personal, social and environmental correlates of physical activity, according to a social ecological framework. Regression modelling identified independent correlates of daily walking separately in the two study regions. In one region, there were independent correlates of walking from all levels of the social ecological framework. In the other region, significant correlates of daily walking were almost all demographic (age, education and marital status. Participants living alone were less likely to be physically active regardless of region. This study highlights the importance of considering region-specific factors when designing strategies for promoting regular walking among rural adults.

Glucocorticoids facilitate the transcription from the human cytomegalovirus major immediate early promoter in glucocorticoid receptor- and nuclear factor-I-like protein-dependent manner

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Inoue-Toyoda, Maki; Kato, Kohsuke; Nagata, Kyosuke; Yoshikawa, Hiroyuki

2015-01-01

Human cytomegalovirus (HCMV) is a common and usually asymptomatic virus agent in healthy individuals. Initiation of HCMV productive infection depends on expression of the major immediate early (MIE) genes. The transcription of HCMV MIE genes is regulated by a diverse set of transcription factors. It was previously reported that productive HCMV infection is triggered probably by elevation of the plasma hydroxycorticoid level. However, it is poorly understood whether the transcription of MIE genes is directly regulated by glucocorticoid. Here, we found that the dexamethasone (DEX), a synthetic glucocorticoid, facilitates the transcription of HCMV MIE genes through the MIE promoter and enhancer in a glucocorticoid receptor (GR)-dependent manner. By competitive EMSA and reporter assays, we revealed that an NF-I like protein is involved in DEX-mediated transcriptional activation of the MIE promoter. Thus, this study supports a notion that the increased level of hydroxycorticoid in the third trimester of pregnancy reactivates HCMV virus production from the latent state. - Highlights: • DEX facilitates the transcription from the HCMV MIE promoter. • GR is involved in DEX-dependent transcription from the HCMV MIE promoter. • A 17 bp repeat is responsible for the HCMV MIE promoter activation by DEX. • An NF-I-like protein is involved in the HCMV MIE promoter activation by DEX

Neurotrophin receptors expression and JNK pathway activation in human astrocytomas

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Assimakopoulou, Martha; Kondyli, Maria; Gatzounis, George; Maraziotis, Theodore; Varakis, John

2007-01-01

grade-dependent manner (p < 0.05). Interestingly, a statistically significant (p < 0.05) reverse relationship between Trk receptors LIs and pc-Jun/pJNK LIs was noted in some glioblastomas multiforme. In the context of astrocytomas, Trk receptors (TrkA, TrkB, TrkC) expression may promote tumor growth independently of grade. Furthermore, activation of JNK pathway may contribute to progression towards malignancy. Considering the fact that regional tumor heterogeneity may be a limiting factor for immunohistochemical studies, the significance of the reverse relationship between Trk receptors and pc-Jun/pJNK LIs with respect to biological behavior of human astrocytomas requires further evaluation

Transcriptional Inhibition of Matrix Metal loproteinase 9 (MMP-9 Activity by a c-fos/Estrogen Receptor Fusion Protein is Mediated by the Proximal AP-1 Site of the MMP-9 Promoter and Correlates with Reduced Tumor Cell Invasion

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David L. Crowe

1999-10-01

Full Text Available Tumor cell invasion of basement membranes is one of the hallmarks of malignant transformation. Tumor cells secrete proteolytic enzymes known as matrix metalloproteinases (MMPs which degrade extracellular matrix molecules. Increased expression of MMP-9 has been associated with acquisition of invasive phenotype in many tumors. However, multiple mechanisms for regulation of MMP-9 gene expression by tumor cell lines have been proposed. A number of transcription factor binding sites have been characterized in the upstream regulatory region of the MMP-9 gene, including those for AP-1. To determine how a specific AP-1 family member, c-fos, regulates MMP-9 promoter activity through these sites, we used an expression vector containing the c-fos coding region fused to the estrogen receptor (ER ligand binding domain. This construct is activated upon binding estradiol. Stable expression of this construct in ER negative squamous cell carcinoma (SCC lines produced an estradiol dependent decrease in the number of cells that migrated through a reconstituted basement membrane. This decreased invasiveness was accompanied by estradiol dependent downregulation of MMP-9 activity as determined by gelatin zymography. Estradiol also produced transcriptional downregulation of an MMP-9 promoter construct in cells transiently transfected with the c-fosER expression vector. This downregulation was mediated by the AP-1 site at —79 by in the MMP-9 promoter. We concluded that the proximal AP-1 site mediated the transcriptional downregulation of the MMP-9 promoter by a conditionally activated c-fos fusion protein.

A var gene promoter implicated in severe malaria nucleates silencing and is regulated by 3' untranslated region and intronic cis-elements.

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Muhle, Rebecca A; Adjalley, Sophie; Falkard, Brie; Nkrumah, Louis J; Muhle, Michael E; Fidock, David A

2009-11-01

Questions surround the mechanism of mutually exclusive expression by which Plasmodium falciparum mediates activation and silencing of var genes. These encode PfEMP1 proteins, which function as cytoadherent and immunomodulatory molecules at the surface of parasitised erythrocytes. Current evidence suggests that promoter silencing by var introns might play a key role in var gene regulation. To evaluate the impact of cis-acting regulatory regions on var silencing, we generated P. falciparum lines in which luciferase was placed under the control of an UpsA var promoter. By utilising the Bxb1 integrase system, these reporter cassettes were targeted to a genomic region that was not in apposition to var subtelomeric domains. This eliminated possible effects from surrounding telomeric elements and removed the variability inherent in episomal systems. Studies with highly synchronised parasites revealed that the UpsA element possessed minimal activity in comparison with a heterologous (hrp3) promoter. This may result from the integrated UpsA promoter being largely silenced by the neighbouring cg6 promoter. Our analyses also revealed that the DownsA 3' untranslated region further decreased the luciferase activity from both cassettes, whereas the var A intron repressed the UpsA promoter specifically. By applying multivariate analysis over the entire cell cycle, we confirmed the significance of these cis-elements and found the parasite stage to be the major factor regulating UpsA-promoter activity. Additionally, we observed that the UpsA promoter was capable of nucleating reversible silencing that spread to a downstream promoter. We believe these studies are the first to analyse promoter activity of Group A var genes, which have been implicated in severe malaria, and support the model that var introns can further suppress var expression. These data also suggest an important suppressive role for the DownsA terminator. Our findings imply the existence of multiple levels of var

Genetic recombination is targeted towards gene promoter regions in dogs.

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Auton, Adam; Rui Li, Ying; Kidd, Jeffrey; Oliveira, Kyle; Nadel, Julie; Holloway, J Kim; Hayward, Jessica J; Cohen, Paula E; Greally, John M; Wang, Jun; Bustamante, Carlos D; Boyko, Adam R

2013-01-01

The identification of the H3K4 trimethylase, PRDM9, as the gene responsible for recombination hotspot localization has provided considerable insight into the mechanisms by which recombination is initiated in mammals. However, uniquely amongst mammals, canids appear to lack a functional version of PRDM9 and may therefore provide a model for understanding recombination that occurs in the absence of PRDM9, and thus how PRDM9 functions to shape the recombination landscape. We have constructed a fine-scale genetic map from patterns of linkage disequilibrium assessed using high-throughput sequence data from 51 free-ranging dogs, Canis lupus familiaris. While broad-scale properties of recombination appear similar to other mammalian species, our fine-scale estimates indicate that canine highly elevated recombination rates are observed in the vicinity of CpG rich regions including gene promoter regions, but show little association with H3K4 trimethylation marks identified in spermatocytes. By comparison to genomic data from the Andean fox, Lycalopex culpaeus, we show that biased gene conversion is a plausible mechanism by which the high CpG content of the dog genome could have occurred.

Receptor autoradiography in the hippocampus of man and rat

International Nuclear Information System (INIS)

Zilles, K.

1988-01-01

This chapter deals with the following questions: regional distribution of binding sites for 5-HT, glutamate, and acetylcholine in Ammon's horn and the dentate gyrus of rat and human brain; comparison of receptor distribution and neuronal pathways with identified transmitters; correlation of region-specific densities between different receptors and receptor subtypes (colocalization of different receptors on the level of hippocampal layers) and comparison of receptor distribution in human and rat hippocampus

Crystal structure of the urokinase receptor in a ligand-free form

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Xu, Xiang; Gårdsvoll, Henrik; Yuan, Cai

2012-01-01

The urokinase receptor urokinase-type plasminogen activator receptor (uPAR) is a surface receptor capable of not only focalizing urokinase-type plasminogen activator (uPA)-mediated fibrinolysis to the pericellular micro-environment but also promoting cell migration and chemotaxis. Consistent...

New twist on neuronal insulin receptor signaling in health, disease, and therapeutics.

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Wada, Akihiko; Yokoo, Hiroki; Yanagita, Toshihiko; Kobayashi, Hideyuki

2005-10-01

Long after the pioneering studies documenting the existence of insulin (year 1967) and insulin receptor (year 1978) in brain, the last decade has witnessed extraordinary progress in the understanding of brain region-specific multiple roles of insulin receptor signalings in health and disease. In the hypothalamus, insulin regulates food intake, body weight, peripheral fat deposition, hepatic gluconeogenesis, reproductive endocrine axis, and compensatory secretion of counter-regulatory hormones to hypoglycemia. In the hippocampus, insulin promotes learning and memory, independent of the glucoregulatory effect of insulin. Defective insulin receptor signalings are associated with the dementia in normal aging and patients with age-related neurodegenerative diseases (e.g., Alzheimer's disease); the cognitive impairment can be reversed with systemic administration of insulin in the euglycemic condition. Intranasal administration of insulin enhances memory and mood and decreases body weight in healthy humans, without causing hypoglycemia. In the hypothalamus, insulin-induced activation of the phosphoinositide 3-kinase pathway followed by opening of ATP-sensitive K+ channel has been shown to be related to multiple effects of insulin. However, the precise molecular mechanisms of insulin's pleiotropic effects still remain obscure. More importantly, much remains unknown about the quality control mechanisms ensuring correct conformational maturation of the insulin receptor, and the cellular mechanisms regulating density of cell surface functional insulin receptors.

CINPA1 Is an Inhibitor of Constitutive Androstane Receptor That Does Not Activate Pregnane X Receptor

OpenAIRE

Cherian, Milu T; Lin, Wenwei; Wu, Jing; Chen, Taosheng

2015-01-01

Constitutive androstane receptor (CAR) and pregnane X receptor (PXR) are xenobiotic sensors that enhance the detoxification and elimination of xenobiotics and endobiotics by modulating the expression of genes encoding drug-metabolizing enzymes and transporters. Elevated levels of drug-metabolizing enzymes and efflux transporters, resulting from CAR activation in various cancers, promote the elimination of chemotherapeutic agents, leading to reduced therapeutic effectiveness and acquired drug ...

Exploring DNA methylation changes in promoter, intragenic, and intergenic regions as early and late events in breast cancer formation

International Nuclear Information System (INIS)

Rauscher, Garth H.; Kresovich, Jacob K.; Poulin, Matthew; Yan, Liying; Macias, Virgilia; Mahmoud, Abeer M.; Al-Alem, Umaima; Kajdacsy-Balla, Andre; Wiley, Elizabeth L.; Tonetti, Debra; Ehrlich, Melanie

2015-01-01

Breast cancer formation is associated with frequent changes in DNA methylation but the extent of very early alterations in DNA methylation and the biological significance of cancer-associated epigenetic changes need further elucidation. Pyrosequencing was done on bisulfite-treated DNA from formalin-fixed, paraffin-embedded sections containing invasive tumor and paired samples of histologically normal tissue adjacent to the cancers as well as control reduction mammoplasty samples from unaffected women. The DNA regions studied were promoters (BRCA1, CD44, ESR1, GSTM2, GSTP1, MAGEA1, MSI1, NFE2L3, RASSF1A, RUNX3, SIX3 and TFF1), far-upstream regions (EN1, PAX3, PITX2, and SGK1), introns (APC, EGFR, LHX2, RFX1 and SOX9) and the LINE-1 and satellite 2 DNA repeats. These choices were based upon previous literature or publicly available DNA methylome profiles. The percent methylation was averaged across neighboring CpG sites. Most of the assayed gene regions displayed hypermethylation in cancer vs. adjacent tissue but the TFF1 and MAGEA1 regions were significantly hypomethylated (p ≤0.001). Importantly, six of the 16 regions examined in a large collection of patients (105 – 129) and in 15-18 reduction mammoplasty samples were already aberrantly methylated in adjacent, histologically normal tissue vs. non-cancerous mammoplasty samples (p ≤0.01). In addition, examination of transcriptome and DNA methylation databases indicated that methylation at three non-promoter regions (far-upstream EN1 and PITX2 and intronic LHX2) was associated with higher gene expression, unlike the inverse associations between cancer DNA hypermethylation and cancer-altered gene expression usually reported. These three non-promoter regions also exhibited normal tissue-specific hypermethylation positively associated with differentiation-related gene expression (in muscle progenitor cells vs. many other types of normal cells). The importance of considering the exact DNA region analyzed and the

Promoting survival: A grounded theory study of consequences of modern health practices in Ouramanat region of Iranian Kurdistan.

Science.gov (United States)

Mohammadpur, Ahmad; Rezaei, Mehdi; Sadeghi, Rasoul

2010-05-14

The aim of this qualitative study is to explore the way people using modern health care perceive its consequences in Ouraman-e-Takht region of Iranian Kurdistan. Ouraman-e-Takht is a rural, highly mountainous and dry region located in the southwest Kurdistan province of Iran. Recently, modern health practices have been introduced to the region. The purpose of this study was to investigate, from the Ouramains' point of view, the impact that modern health services and practices have had on the Ouraman traditional way of life. Interview data from respondents were analyzed by using grounded theory. Promoting survival was the core category that explained the impact that modern health practices have had on the Ouraman region. The people of Ouraman interpreted modern health practices as increasing their quality of life and promoting their survival. Results are organized around this core category in a paradigm model consisting of conditions, interactions, and consequences. This model can be used to understand the impact of change from the introduction of modern health on a traditional society.

Promoting survival: A grounded theory study of consequences of modern health practices in Ouramanat region of Iranian Kurdistan

Science.gov (United States)

Mohammadpur, Ahmad; Rezaei, Mehdi; Sadeghi, Rasoul

2010-01-01

The aim of this qualitative study is to explore the way people using modern health care perceive its consequences in Ouraman-e-Takht region of Iranian Kurdistan. Ouraman-e-Takht is a rural, highly mountainous and dry region located in the southwest Kurdistan province of Iran. Recently, modern health practices have been introduced to the region. The purpose of this study was to investigate, from the Ouramains' point of view, the impact that modern health services and practices have had on the Ouraman traditional way of life. Interview data from respondents were analyzed by using grounded theory. Promoting survival was the core category that explained the impact that modern health practices have had on the Ouraman region. The people of Ouraman interpreted modern health practices as increasing their quality of life and promoting their survival. Results are organized around this core category in a paradigm model consisting of conditions, interactions, and consequences. This model can be used to understand the impact of change from the introduction of modern health on a traditional society. PMID:20640020

Caffeine promotes wakefulness via dopamine signaling in Drosophila

Science.gov (United States)

Nall, Aleksandra H.; Shakhmantsir, Iryna; Cichewicz, Karol; Birman, Serge; Hirsh, Jay; Sehgal, Amita

2016-01-01

Caffeine is the most widely-consumed psychoactive drug in the world, but our understanding of how caffeine affects our brains is relatively incomplete. Most studies focus on effects of caffeine on adenosine receptors, but there is evidence for other, more complex mechanisms. In the fruit fly Drosophila melanogaster, which shows a robust diurnal pattern of sleep/wake activity, caffeine reduces nighttime sleep behavior independently of the one known adenosine receptor. Here, we show that dopamine is required for the wake-promoting effect of caffeine in the fly, and that caffeine likely acts presynaptically to increase dopamine signaling. We identify a cluster of neurons, the paired anterior medial (PAM) cluster of dopaminergic neurons, as the ones relevant for the caffeine response. PAM neurons show increased activity following caffeine administration, and promote wake when activated. Also, inhibition of these neurons abrogates sleep suppression by caffeine. While previous studies have focused on adenosine-receptor mediated mechanisms for caffeine action, we have identified a role for dopaminergic neurons in the arousal-promoting effect of caffeine. PMID:26868675

Arrestin scaffolds NHERF1 to the P2Y12 receptor to regulate receptor internalization.

Science.gov (United States)

Nisar, Shaista P; Cunningham, Margaret; Saxena, Kunal; Pope, Robert J; Kelly, Eamonn; Mundell, Stuart J

2012-07-13

We have recently shown in a patient with mild bleeding that the PDZ-binding motif of the platelet G protein-coupled P2Y(12) receptor (P2Y(12)R) is required for effective receptor traffic in human platelets. In this study we show for the first time that the PDZ motif-binding protein NHERF1 exerts a major role in potentiating G protein-coupled receptor (GPCR) internalization. NHERF1 interacts with the C-tail of the P2Y(12)R and unlike many other GPCRs, NHERF1 interaction is required for effective P2Y(12)R internalization. In vitro and prior to agonist stimulation P2Y(12)R/NHERF1 interaction requires the intact PDZ binding motif of this receptor. Interestingly on receptor stimulation NHERF1 no longer interacts directly with the receptor but instead binds to the receptor via the endocytic scaffolding protein arrestin. These findings suggest a novel model by which arrestin can serve as an adaptor to promote NHERF1 interaction with a GPCR to facilitate effective NHERF1-dependent receptor internalization.

Laminar pattern of cholinergic and adrenergic receptors in rat visual cortex using quantitative receptor autoradiography

International Nuclear Information System (INIS)

Schliebs, R.; Walch, C.

1989-01-01

The laminar distribution of muscarinic acetylcholine receptors, including the M1-receptor subtype, of beta-adrenergic receptors, and noradrenaline uptake sites, was studied in the adult rat visual, frontal, somatosensory and motor cortex, using quantitative receptor autoradiography. In the visual cortex, the highest density of muscarinic acetylcholine receptors was found in layer I. From layer II/III to layer V binding decreases continueously reaching a constant binding level in layers V and VI. This laminar pattern of muscarinic receptor density differs somewhat from that observed in the non-visual cortical regions examined: layer II/III contained the highest receptor density followed by layer I and IV: lowest density was found in layer V and VI. The binding profile of the muscarinic cholinergic M1-subtype through the visual cortex shows a peak in cortical layer II and in the upper part of layer VI, whereas in the non-visual cortical regions cited the binding level was high in layer II/III, moderate in layer I and IV, and low in layer VI. Layers I to IV of the visual cortex contained the highest beta-adrenergic receptor densities, whereas only low binding levels were observed in the deeper layers. A similar laminar distribution was found also in the frontal, somatosensory and motor cortex. The density of noradrenaline uptake sites was high in all layers of the cortical regions studied, but with noradrenaline uptake sites somewhat more concentrated in the superficial layers than in deeper ones. The distinct laminar pattern of cholinergic and noradrenergic receptor sites indicates a different role for acetylcholine and noradrenaline in the functional anatomy of the cerebral cortex, and in particular, the visual cortex. (author)

Laminar pattern of cholinergic and adrenergic receptors in rat visual cortex using quantitative receptor autoradiography

Energy Technology Data Exchange (ETDEWEB)

Schliebs, R; Walch, C [Leipzig Univ. (German Democratic Republic). Bereich Medizin; Stewart, M G [Open Univ., Milton Keynes (UK)

1989-01-01

The laminar distribution of muscarinic acetylcholine receptors, including the M1-receptor subtype, of beta-adrenergic receptors, and noradrenaline uptake sites, was studied in the adult rat visual, frontal, somatosensory and motor cortex, using quantitative receptor autoradiography. In the visual cortex, the highest density of muscarinic acetylcholine receptors was found in layer I. From layer II/III to layer V binding decreases continueously reaching a constant binding level in layers V and VI. This laminar pattern of muscarinic receptor density differs somewhat from that observed in the non-visual cortical regions examined: layer II/III contained the highest receptor density followed by layer I and IV: lowest density was found in layer V and VI. The binding profile of the muscarinic cholinergic M1-subtype through the visual cortex shows a peak in cortical layer II and in the upper part of layer VI, whereas in the non-visual cortical regions cited the binding level was high in layer II/III, moderate in layer I and IV, and low in layer VI. Layers I to IV of the visual cortex contained the highest beta-adrenergic receptor densities, whereas only low binding levels were observed in the deeper layers. A similar laminar distribution was found also in the frontal, somatosensory and motor cortex. The density of noradrenaline uptake sites was high in all layers of the cortical regions studied, but with noradrenaline uptake sites somewhat more concentrated in the superficial layers than in deeper ones. The distinct laminar pattern of cholinergic and noradrenergic receptor sites indicates a different role for acetylcholine and noradrenaline in the functional anatomy of the cerebral cortex, and in particular, the visual cortex. (author).

Differential subcellular localization of insulin receptor substrates depends on C-terminal regions and importin β

International Nuclear Information System (INIS)

Kabuta, Tomohiro; Take, Kazumi; Kabuta, Chihana; Hakuno, Fumihiko; Takahashi, Shin-Ichiro

2008-01-01

Insulin receptor substrates (IRSs) play essential roles in signal transduction of insulin and insulin-like growth factors. Previously, we showed that IRS-3 is localized to the nucleus as well as the cytosol, while IRS-1 and 2 are mainly localized to the cytoplasm. In the present study, we found that importin β directly interacts with IRS-3 and is able to mediate nuclear transport of IRS-3. Importin β interacted with the pleckstrin homology domain, the phosphotyrosine binding domain and the C-terminal region of IRS-3; indeed all of these fragments exhibited predominant nuclear localization. By contrast, almost no interaction of importin β with IRS-1 and -2 was observed, and their C-terminal regions displayed discrete spotty images in the cytosol. In addition, using chimeric proteins between IRS-1 and IRS-3, we revealed that the C-terminal regions are the main determinants of the differing subcellular localizations of IRS-1 and IRS-3.

Growth and gene expression are predominantly controlled by distinct regions of the human IL-4 receptor.

Science.gov (United States)

Ryan, J J; McReynolds, L J; Keegan, A; Wang, L H; Garfein, E; Rothman, P; Nelms, K; Paul, W E

1996-02-01

IL-4 causes hematopoietic cells to proliferate and express a series of genes, including CD23. We examined whether IL-4-mediated growth, as measured by 4PS phosphorylation, and gene induction were similarly controlled. Studies of M12.4.1 cells expressing human IL-4R truncation mutants indicated that the region between amino acids 557-657 is necessary for full gene expression, which correlated with Stat6 DNA binding activity. This region was not required for 4PS phosphorylation. Tyrosine-to-phenylalanine mutations in the interval between amino acids 557-657 revealed that as long as one tyrosine remained unmutated, CD23 was fully induced. When all three tyrosines were mutated, the receptor was unable to induce CD23. The results indicate that growth regulation and gene expression are principally controlled by distinct regions of IL-4R.

Decreased expression of thyroid receptor-associated protein 220 in temporal lobe tissue of patients with refractory epilepsy

International Nuclear Information System (INIS)

Li Jinmei; Wang Xuefeng; Xi Zhiqin; Gong Yun; Liu Fengying; Sun Jijun; Wu Yuan; Luan Guoming; Wang Yuping; Li Yunlin; Zhang Jianguo; Lu Yong; Li Hongwei

2006-01-01

Purpose: TRAP220 (thyroid hormone receptor-associated protein) functions as a coactivator for nuclear receptors and stimulates transcription by recruiting the TRAP mediator complex to hormone responsive promoter regions. Thus, TRAP220 enhances the function of thyroid/steroid hormone receptors such as thyroid hormone and oestrogen receptors. This study investigated the expression of TRAP220 mRNA and protein level in epileptic brains comparing with human control. Methods: We examined the expression of TRAP220 mRNA and protein levels in temporal lobes from patients with chronic pharmacoresistant epilepsy who have undergone surgery. Results: Expression of TRAP220 mRNA and protein was shown to be decreased significantly in the temporal cortex of the patients with epilepsy. Conclusions: Our work showed that a decrease in TRAP220 mRNA and protein levels may be involved in the pathophysiology of epilepsy and may be associated with impairment of the brain caused by frequent seizures

The Receptor-Binding Domain in the VP1u Region of Parvovirus B19.

Science.gov (United States)

Leisi, Remo; Di Tommaso, Chiarina; Kempf, Christoph; Ros, Carlos

2016-02-24

Parvovirus B19 (B19V) is known as the human pathogen causing the mild childhood disease erythema infectiosum. B19V shows an extraordinary narrow tissue tropism for erythroid progenitor cells in the bone marrow, which is determined by a highly restricted uptake. We have previously shown that the specific internalization is mediated by the interaction of the viral protein 1 unique region (VP1u) with a yet unknown cellular receptor. To locate the receptor-binding domain (RBD) within the VP1u, we analyzed the effect of truncations and mutations on the internalization capacity of the recombinant protein into UT7/Epo cells. Here we report that the N-terminal amino acids 5-80 of the VP1u are necessary and sufficient for cellular binding and internalization; thus, this N-terminal region represents the RBD required for B19V uptake. Using site-directed mutagenesis, we further identified a cluster of important amino acids playing a critical role in VP1u internalization. In silico predictions and experimental results suggest that the RBD is structured as a rigid fold of three α-helices. Finally, we found that dimerization of the VP1u leads to a considerably enhanced cellular binding and internalization. Taken together, we identified the RBD that mediates B19V uptake and mapped functional and structural motifs within this sequence. The findings reveal insights into the uptake process of B19V, which contribute to understand the pathogenesis of the infection and the neutralization of the virus by the immune system.

Allelic polymorphisms in the repeat and promoter regions of the interleukin-4 gene and malaria severity in Ghanaian children

DEFF Research Database (Denmark)

Gyan, B A; Goka, B; Cvetkovic, J T

2004-01-01

Immunoglobulin E has been associated with severe malaria suggesting a regulatory role for interleukin (IL)-4 and/or IgE in the pathogenesis of severe malaria. We have investigated possible associations between polymorphisms in the IL-4 repeat region (intron 3) and promoter regions (IL-4 +33CT and...

Transient receptor potential ankyrin 1 channel localized to non-neuronal airway cells promotes non-neurogenic inflammation

DEFF Research Database (Denmark)

Nassini, Romina; Pedretti, Pamela; Moretto, Nadia

2012-01-01

The transient receptor potential ankyrin 1 (TRPA1) channel, localized to airway sensory nerves, has been proposed to mediate airway inflammation evoked by allergen and cigarette smoke (CS) in rodents, via a neurogenic mechanism. However the limited clinical evidence for the role of neurogenic...... inflammation in asthma or chronic obstructive pulmonary disease raises an alternative possibility that airway inflammation is promoted by non-neuronal TRPA1.By using Real-Time PCR and calcium imaging, we found that cultured human airway cells, including fibroblasts, epithelial and smooth muscle cells express...... functional TRPA1 channels. By using immunohistochemistry, TRPA1 staining was observed in airway epithelial and smooth muscle cells in sections taken from human airways and lung, and from airways and lung of wild-type, but not TRPA1-deficient mice. In cultured human airway epithelial and smooth muscle cells...

Cloning of human genes encoding novel G protein-coupled receptors

Energy Technology Data Exchange (ETDEWEB)

Marchese, A.; Docherty, J.M.; Heiber, M. [Univ. of Toronto, (Canada)] [and others

1994-10-01

We report the isolation and characterization of several novel human genes encoding G protein-coupled receptors. Each of the receptors contained the familiar seven transmembrane topography and most closely resembled peptide binding receptors. Gene GPR1 encoded a receptor protein that is intronless in the coding region and that shared identity (43% in the transmembrane regions) with the opioid receptors. Northern blot analysis revealed that GPR1 transcripts were expressed in the human hippocampus, and the gene was localized to chromosome 15q21.6. Gene GPR2 encoded a protein that most closely resembled an interleukin-8 receptor (51% in the transmembrane regions), and this gene, not expressed in the six brain regions examined, was localized to chromosome 17q2.1-q21.3. A third gene, GPR3, showed identity (56% in the transmembrane regions) with a previously characterized cDNA clone from rat and was localized to chromosome 1p35-p36.1. 31 refs., 5 figs., 1 tab.

Biological sex influences learning strategy preference and muscarinic receptor binding in specific brain regions of prepubertal rats.

Science.gov (United States)

Grissom, Elin M; Hawley, Wayne R; Hodges, Kelly S; Fawcett-Patel, Jessica M; Dohanich, Gary P

2013-04-01

According to the theory of multiple memory systems, specific brain regions interact to determine how the locations of goals are learned when rodents navigate a spatial environment. A number of factors influence the type of strategy used by rodents to remember the location of a given goal in space, including the biological sex of the learner. We recently found that prior to puberty male rats preferred a striatum-dependent stimulus-response strategy over a hippocampus-dependent place strategy when solving a dual-solution task, while age-matched females showed no strategy preference. Because the cholinergic system has been implicated in learning strategy and is known to be sexually dimorphic prior to puberty, we explored the relationship between learning strategy and muscarinic receptor binding in specific brain regions of prepubertal males and female rats. We confirmed our previous finding that at 28 days of age a significantly higher proportion of prepubertal males preferred a stimulus-response learning strategy than a place strategy to solve a dual-solution visible platform water maze task. Equal proportions of prepubertal females preferred stimulus-response or place strategies. Profiles of muscarinic receptor binding as assessed by autoradiography varied according to strategy preference. Regardless of biological sex, prepubertal rats that preferred stimulus-response strategy exhibited lower ratios of muscarinic receptor binding in the hippocampus relative to the dorsolateral striatum compared to rats that preferred place strategy. Importantly, much of the variance in this ratio was related to differences in the ventral hippocampus to a greater extent than the dorsal hippocampus. The ratios of muscarinic receptors in the hippocampus relative to the basolateral amygdala also were lower in rats that preferred stimulus-response strategy over place strategy. Results confirm that learning strategy preference varies with biological sex in prepubertal rats with males

Carbetocin is a Functional Selective Gq Agonist That Does Not Promote Oxytocin Receptor Recycling After Inducing β-Arrestin-Independent Internalisation.

Science.gov (United States)

Passoni, I; Leonzino, M; Gigliucci, V; Chini, B; Busnelli, M

2016-04-01

Carbetocin, a long-acting oxytocin analogue, has been reported to elicit interesting and peculiar behavioural effects. The present study investigated the molecular pharmacology of carbetocin, aiming to better understand the molecular basis of its action in the brain. Using bioluminescence resonance energy transfer biosensors, we characterised the effects of carbetocin on the three human oxytocin/vasopressin receptors expressed in the nervous system: the oxytocin receptor (OXTR) and the vasopressin V1a (V1aR) and V1b (V1bR) receptors. Our results indicate that (i) carbetocin activates the OXTR but not the V1aR and V1bR at which it may act as an antagonist; (ii) carbetocin selectively activates only the OXTR/Gq pathway displaying a strong functional selectivity; (iii) carbetocin is a partial agonist at the OXTR/Gq coupling; (iv) carbetocin promotes OXTR internalisation via a previously unreported β-arrestin-independent pathway; and (v) carbetocin does not induce OXTR recycling to the plasma membrane. Altogether, these molecular pharmacology features identify carbetocin as a substantially different analogue compared to the endogenous oxytocin and, consequently, carbetocin is not expected to mimic oxytocin in the brain. Whether these unique features of carbetocin could be exploited therapeutically remains to be established. © 2016 The Authors. Journal of Neuroendocrinology published by John Wiley & Sons Ltd on behalf of British Society for Neuroendocrinology.

Synergistic Action of Presynaptic Muscarinic Acetylcholine Receptors and Adenosine Receptors in Developmental Axonal Competition at the Neuromuscular Junction.

Science.gov (United States)

Nadal, Laura; Garcia, Neus; Hurtado, Erica; Simó, Anna; Tomàs, Marta; Lanuza, Maria Angel; Cilleros, Victor; Tomàs, Josep Maria

2016-01-01

The development of the nervous system involves the initial overproduction of synapses, which promotes connectivity. Hebbian competition between axons with different activities leads to the loss of roughly half of the overproduced elements and this refines connectivity. We used quantitative immunohistochemistry to investigate, in the postnatal day 7 (P7) to P9 neuromuscular junctions, the involvement of muscarinic receptors (muscarinic acetylcholine autoreceptors and the M1, M2, and M4 subtypes) and adenosine receptors (A1 and A2A subtypes) in the control of axonal elimination after the mouse levator auris longus muscle had been exposed to selective antagonists in vivo. In a previous study we analyzed the role of each of the individual receptors. Here we investigate the additive or occlusive effects of their inhibitors and thus the existence of synergistic activity between the receptors. The main results show that the A2A, M1, M4, and A1 receptors (in this order of ability) delayed axonal elimination at P7. M4 produces some occlusion of the M1 pathway and some addition to the A1 pathway, which suggests that they cooperate. M2 receptors may modulate (by allowing a permissive action) the other receptors, mainly M4 and A1. The continued action of these receptors (now including M2 but not M4) finally promotes axonal loss at P9. All 4 receptors (M2, M1, A1, and A2A, in this order of ability) are necessary. The M4 receptor (which in itself does not affect axon loss) seems to modulate the other receptors. We found a synergistic action between the M1, A1, and A2A receptors, which show an additive effect, whereas the potent M2 effect is largely independent of the other receptors (though can be modulated by M4). At P9, there is a full mutual dependence between the A1 and A2A receptors in regulating axon loss. In summary, postnatal axonal elimination is a regulated multireceptor mechanism that involves the cooperation of several muscarinic and adenosine receptor subtypes. �

A var gene promoter implicated in severe malaria nucleates silencing and is regulated by 3’ untranslated region and intronic cis-elements

Science.gov (United States)

Muhle, Rebecca A.; Adjalley, Sophie; Falkard, Brie; Nkrumah, Louis J.; Muhle, Michael E.; Fidock, David A.

2009-01-01

Questions surround the mechanism of mutually exclusive expression by which Plasmodium falciparum mediates activation and silencing of var genes. These encode PfEMP1 proteins, which function as cytoadherent and immunomodulatory molecules at the surface of parasitized erythrocytes. Current evidence suggests that promoter silencing by var introns might play a key role in var gene regulation. To evaluate the impact of cis-acting regulatory regions on var silencing, we generated P. falciparum lines in which luciferase was placed under the control of an UpsA var promoter. By utilizing the Bxb1 integrase system, these reporter cassettes were targeted to a genomic region that was not in apposition to var sub-telomeric domains. This eliminated possible effects from surrounding telomeric elements and removed the variability inherent in episomal systems. Studies with highly synchronized parasites revealed that the UpsA element possessed minimal activity in comparison with a heterologous (hrp3) promoter. This may well result from the integrated UpsA promoter being largely silenced by the neighboring cg6 promoter. Our analyses also revealed that the DownsA 3’ untranslated region further decreased the luciferase activity from both cassettes, whereas the var A intron repressed the UpsA promoter specifically. By applying multivariate analysis over the entire cell cycle, we confirmed the significance of these cis-elements and found the parasite stage to be the major factor regulating UpsA promoter activity. Additionally, we observed that the UpsA promoter was capable of nucleating reversible silencing that spread to a downstream promoter. We believe these studies are the first to analyze promoter activity of Group A var genes which have been implicated in severe malaria, and support the model that var introns can further suppress var expression. These data also suggest an important suppressive role for the DownsA terminator. Our findings imply the existence of multiple levels of

Cloning, mapping and molecular characterization of porcine progesterone receptor membrane component 2 (PGRMC2 gene

Directory of Open Access Journals (Sweden)

Congying Chen

2010-01-01

Full Text Available Progesterone plays an important role in sow reproduction by stimulating classic genomic pathways via nuclear receptors and non-genomic pathways via membrane receptors such a progesterone receptor membrane component 2 (PGRMC2. In this work, we used radiation hybrid mapping to assign PGRMC2 to pig chromosome 8 and observed that this receptor has two transcripts in pigs. The full-length cDNA of the large transcript is 1858 bp long and contains a 669-bp open reading frame (ORF encoding a protein of 223 amino acids. The shorter transcript encodes a protein of 170 amino acids. The porcine PGRMC2 gene consists of three exons 446 bp, 156 bp and 1259 bp in length. The promoter sequence is GC-rich and lacks a typical TATA box. Several putative cis-regulatory DNA motifs were identified in the 208-bp upstream genomic region. Five single nucleotide polymorphisms (SNPs were detected in introns* and the 3' UTR. RT-PCR indicated that the PGRMC2 gene is expressed ubiquitously in all pig tissues examined.

Methylation of Promoter Regions of Genes of the Human Intrauterine Renin Angiotensin System and Their Expression

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Shane D. Sykes

2015-01-01

Full Text Available The intrauterine renin angiotensin system (RAS is implicated in placentation and labour onset. Here we investigate whether promoter methylation of RAS genes changes with gestation or labour and if it affects gene expression. Early gestation amnion and placenta were studied, as were term amnion, decidua, and placenta collected before labour (at elective caesarean section or after spontaneous labour and delivery. The expression and degree of methylation of the prorenin receptor (ATP6AP2, angiotensin converting enzyme (ACE, angiotensin II type 1 receptor (AGTR1, and two proteases that can activate prorenin (kallikrein, KLK1, and cathepsin D, CTSD were measured by qPCR and a DNA methylation array. There was no effect of gestation or labour on the methylation of RAS genes and CTSD. Amnion and decidua displayed strong correlations between the percent hypermethylation of RAS genes and CTSD, suggestive of global methylation. There were no correlations between the degree of methylation and mRNA abundance of any genes studied. KLK1 was the most methylated gene and the proportion of hypermethylated KLK1 alleles was lower in placenta than decidua. The presence of intermediate methylated alleles of KLK1 in early gestation placenta and in amnion after labour suggests that KLK1 methylation is uniquely dynamic in these tissues.

Mode of action and dose-response framework analysis for receptor-mediated toxicity : The aryl hydrocarbon receptor as a case study

NARCIS (Netherlands)

Budinsky, R. A.; Schrenk, D.; Simon, T.; Van Den Berg, M.; Reichard, J. F.; Silkworth, J. B.; Aylward, L. L.; Brix, A.; Gasiewicz, T.; Kaminski, N.; Perdew, G.; Starr, T. B.; Walker, N. J.; Rowlands, J. C.

2014-01-01

Dioxins and dioxin-like compounds are tumor promoters that cause liver cancer in rats and mice. The aryl hydrocarbon receptor (AHR) has been implicated as a key component in this tumor promotion response. Despite extensive knowledge of the toxicology of dioxins, no mode of action (MOA) hypothesis

Androgen Receptor Signaling in Bladder Cancer

Science.gov (United States)

Li, Peng; Chen, Jinbo; Miyamoto, Hiroshi

2017-01-01

Emerging preclinical findings have indicated that steroid hormone receptor signaling plays an important role in bladder cancer outgrowth. In particular, androgen-mediated androgen receptor signals have been shown to correlate with the promotion of tumor development and progression, which may clearly explain some sex-specific differences in bladder cancer. This review summarizes and discusses the available data, suggesting the involvement of androgens and/or the androgen receptor pathways in urothelial carcinogenesis as well as tumor growth. While the precise mechanisms of the functions of the androgen receptor in urothelial cells remain far from being fully understood, current evidence may offer chemopreventive or therapeutic options, using androgen deprivation therapy, in patients with bladder cancer. PMID:28241422

Androgen Receptor Signaling in Bladder Cancer

Directory of Open Access Journals (Sweden)

Peng Li

2017-02-01

Full Text Available Emerging preclinical findings have indicated that steroid hormone receptor signaling plays an important role in bladder cancer outgrowth. In particular, androgen-mediated androgen receptor signals have been shown to correlate with the promotion of tumor development and progression, which may clearly explain some sex-specific differences in bladder cancer. This review summarizes and discusses the available data, suggesting the involvement of androgens and/or the androgen receptor pathways in urothelial carcinogenesis as well as tumor growth. While the precise mechanisms of the functions of the androgen receptor in urothelial cells remain far from being fully understood, current evidence may offer chemopreventive or therapeutic options, using androgen deprivation therapy, in patients with bladder cancer.

Characterization of the single transmembrane domain of human receptor activity-modifying protein 3 in adrenomedullin receptor internalization

International Nuclear Information System (INIS)

Kuwasako, Kenji; Kitamura, Kazuo; Nagata, Sayaka; Nozaki, Naomi; Kato, Johji

2012-01-01

Highlights: ► RAMP3 mediates CLR internalization much less effectively than does RAMP2. ► The RAMP3 TMD participates in the negative regulation of CLR/RAMP3 internalization. ► A new strategy of promoting internalization and resensitization of the receptor was found. -- Abstract: Two receptor activity-modifying proteins (RAMP2 and RAMP3) enable calcitonin receptor-like receptor (CLR) to function as two heterodimeric receptors (CLR/RAMP2 and CLR/RAMP3) for adrenomedullin (AM), a potent cardiovascular protective peptide. Following AM stimulation, both receptors undergo rapid internalization through a clathrin-dependent pathway, after which CLR/RAMP3, but not CLR/RAMP2, can be recycled to the cell surface for resensitization. However, human (h)RAMP3 mediates CLR internalization much less efficiently than does hRAMP2. Therefore, the molecular basis of the single transmembrane domain (TMD) and the intracellular domain of hRAMP3 during AM receptor internalization was investigated by transiently transfecting various RAMP chimeras and mutants into HEK-293 cells stably expressing hCLR. Flow cytometric analysis revealed that substituting the RAMP3 TMD with that of RAMP2 markedly enhanced AM-induced internalization of CLR. However, this replacement did not enhance the cell surface expression of CLR, [ 125 I]AM binding affinity or AM-induced cAMP response. More detailed analyses showed that substituting the Thr 130 –Val 131 sequence in the RAMP3 TMD with the corresponding sequence (Ile 157 –Pro 158 ) from RAMP2 significantly enhanced AM-mediated CLR internalization. In contrast, substituting the RAMP3 target sequence with Ala 130 –Ala 131 did not significantly affect CLR internalization. Thus, the RAMP3 TMD participates in the negative regulation of CLR/RAMP3 internalization, and the aforementioned introduction of the Ile–Pro sequence into the RAMP3 TMD may be a strategy for promoting receptor internalization/resensitization.

Promoter motifs required for c-mpl gene expression induced by thrombopoietin in CMK cells.

Science.gov (United States)

Sunohara, Masataka; Sato, Iwao; Morikawa, Shigeru

2017-11-30

Thrombopoietin (TPO) and its receptor, c-Mpl, are the central regulators of megakaryocyte development and platelet production and are also crucial to regulate megakaryocytopoiesis. TPO remarkably elevated c-mpl promoter activity, while the protein kinase C (PKC) inhibitors, GF109203, H7 and Calphostin C, clearly reduced the steady level of its promoter activity.  In the present study, motifs crucial for c-mpl promoter activity induced by TPO treatment have been analyzed using a human megakaryoblastic cell line, CMK. Destruction of the -107Sp1 and the -57Sp1 sites in the c-mpl promoter enhancer region resulted in decrease of the promoter activity by 53.1% and 64.4%, respectively, and destruction of -69Ets and -28Ets elements dramatically decreased the promoter activity by 96.4% and 87.8%, respectively, while mutation of -77GATA moderately reduced the activity by 31.4%. The result was in agreement with our previous report that showed the crucial motifs in the c-mpl promoter for the promoter activity induced by PMA-treatment. This indicates that TPO-induced activation of the c-mpl promoter activity is fully modulated by transcription through a PKC-dependent pathway and the two Sp1 and two Ets motifs are crucial for the activation of the c-mpl promoter activity rather than a GATA motif in the c-mpl promoter of CMK cells.

Palytoxin: exploiting a novel skin tumor promoter to explore signal transduction and carcinogenesis.

Science.gov (United States)

Wattenberg, Elizabeth V

2007-01-01

Palytoxin is a novel skin tumor promoter, which has been used to help probe the role of different types of signaling mechanisms in carcinogenesis. The multistage mouse skin model indicates that tumor promotion is an early, prolonged, and reversible phase of carcinogenesis. Understanding the molecular mechanisms underlying tumor promotion is therefore important for developing strategies to prevent and treat cancer. Naturally occurring tumor promoters that bind to specific cellular receptors have proven to be useful tools for investigating important biochemical events in multistage carcinogenesis. For example, the identification of protein kinase C as the receptor for the prototypical skin tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) (also called phorbol 12-myristate 13-acetate, PMA) provided key evidence that tumor promotion involves the aberrant modulation of signaling cascades that govern cell fate and function. The subsequent discovery that palytoxin, a marine toxin isolated from zoanthids (genus Palythoa), is a potent skin tumor promoter yet does not activate protein kinase C indicated that investigating palytoxin action could help reveal new aspects of tumor promotion. Interestingly, the putative receptor for palytoxin is the Na(+),K(+)-ATPase. This review focuses on palytoxin-stimulated signaling and how palytoxin has been used to investigate alternate biochemical mechanisms by which important targets in carcinogenesis can be modulated.

Altered promoter methylation of PDK4, IL1 B, IL6, and TNF after Roux-en Y gastric bypass

DEFF Research Database (Denmark)

Kirchner, Henriette; Nylen, Carolina; Laber, Samantha

2014-01-01

methylation of selected promoter regions was measured in whole blood before and after VLCD. A subgroup of seven patients was studied 1–2 days and 12± 3 months after RYGB. Promoter methylation was measured using methylated DNA capture and quantitative real-time polymerase chain reaction (PCR). Results VLCD....... The objective of this study was to test whether promoter methylation of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PPARGC1 A), pyruvate dehydrogenase kinase isozyme-4 (PDK4), transcription factor A (TFAM), interleukin-1 beta (IL1 B), interleukin-6 (IL6) and tumor necrosis factor...... decreased promoter methylation of PPARGC1 A. Methylation of PPARGC1 A, TFAM, IL1 B, IL6, and TNF promoters was changed two days after RYGB. Similar changes were also seen on day one after cholecystectomy. Moreover, methylation increased in PDK4, IL1 B, IL6, and TNF promoters 12 months after RYGB. Conclusion...

Hypermethylation of the DPYD promoter region is not a major predictor of severe toxicity in 5-fluorouracil based chemotherapy

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Aebi Stefan

2008-10-01

Full Text Available Abstract Background The activity of dihydropyrimidine dehydrogenase (DPD, the key enzyme of pyrimidine catabolism, is thought to be an important determinant for the occurrence of severe toxic reactions to 5-fluorouracil (5-FU, which is one of the most commonly prescribed chemotherapeutic agents for the treatment of solid cancers. Genetic variation in the DPD gene (DPYD has been proposed as a main factor for variation in DPD activity in the population. However, only a small proportion of severe toxicities in 5-FU based chemotherapy can be explained with such rare deleterious DPYD mutations resulting in severe enzyme deficiencies. Recently, hypermethylation of the DPYD promoter region has been proposed as an alternative mechanism for DPD deficiency and thus as a major cause of severe 5-FU toxicity. Methods Here, the prognostic significance of this epigenetic marker with respect to severe 5-FU toxicity was assessed in 27 cancer patients receiving 5-FU based chemotherapy, including 17 patients experiencing severe toxic side effects following drug administration, none of which were carriers of a known deleterious DPYD mutation, and ten control patients. The methylation status of the DPYD promoter region in peripheral blood mononuclear cells was evaluated by analysing for each patient between 19 and 30 different clones of a PCR-amplified 209 base pair fragment of the bisulfite-modified DPYD promoter region. The fragments were sequenced to detect bisulfite-induced, methylation-dependent sequence differences. Results No evidence of DPYD promoter methylation was observed in any of the investigated patient samples, whereas in a control experiment, as little as 10% methylated genomic DNA could be detected. Conclusion Our results indicate that DYPD promoter hypermethylation is not of major importance as a prognostic factor for severe toxicity in 5-FU based chemotherapy.

Both mineralocorticoid and glucocorticoid receptors regulate emotional memory in mice

NARCIS (Netherlands)

Zhou, M.; Bakker, E.H.M.; Velzing, E.; Berger, S.; Oitzl, M.; Joëls, M.; Krugers, H.J.

2010-01-01

Corticosteroid hormones are thought to promote optimal behavioral adaptation under fearful conditions, primarily via glucocorticoid receptors (GRs). Here, we examined - using pharmacological and genetic approaches in mice - if mineralocorticoid receptors (MRs) also play a role in fearful memory

Boundary Dpp promotes growth of medial and lateral regions of the Drosophila wing.

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Barrio, Lara; Milán, Marco

2017-07-04

The gradient of Decapentaplegic (Dpp) in the Drosophila wing has served as a paradigm to characterize the role of morphogens in regulating patterning. However, the role of this gradient in regulating tissue size is a topic of intense debate as proliferative growth is homogenous. Here, we combined the Gal4/UAS system and a temperature-sensitive Gal80 molecule to induce RNAi-mediated depletion of dpp and characterise the spatial and temporal requirement of Dpp in promoting growth. We show that Dpp emanating from the AP compartment boundary is required throughout development to promote growth by regulating cell proliferation and tissue size. Dpp regulates growth and proliferation rates equally in central and lateral regions of the developing wing appendage and reduced levels of Dpp affects similarly the width and length of the resulting wing. We also present evidence supporting the proposal that graded activity of Dpp is not an absolute requirement for wing growth.

Impulsive traits and 5-HT2A receptor promoter polymorphism in alcohol dependents: possible association but no influence of personality disorders.

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Preuss, U W; Koller, G; Bondy, B; Bahlmann, M; Soyka, M

2001-01-01

Impulsive behavior in alcoholics puts them at serious risk of severer course of disease and has been related to the serotonergic neurotransmission dysfunction. The aim of this study is to investigate the association between impulsive aggression in alcohol dependents with regard to the G-1438A polymorphism in the promoter region of the 5-HT2A receptor gene. Furthermore, we investigated the statistical interaction between 5-HT2A alleles, antisocial personality disorder (APD) and impulsive aggression in alcohol dependents. Alcohol dependents were investigated because these personality disorders and impulsive behavior are very frequent in alcohol dependence anf of clinical relevance. One hundred and thirty-five patients of German descent meeting DSM-IV criteria of alcohol dependence were recruited. Blood samples were taken from alcohol dependents to determine 5-HT2A promoter polymorphisms using PCR (polymerase chain reaction) of lymphocyte DNA. Impulsive aggression was assessed using a German version of the Barratt Impulsiveness Scale which was translated and backtranslated. Alcohol dependents were subdivided into low- or high-impulsivity groups using a median split of the Barratt score. APD and borderline personality disorder (BPD) were assessed using the SCID-II interview. The low-impulsivity group was slightly older and showed a later age at alcoholism onset than the highly impulsive group. Alcohol dependents with high impulsive traits showed a significant association with 5-HT2A 1438 A alleles. After excluding alcohol dependents with APD or BPD from the analysis, this association remained significant. Furthermore, no association between APD, BPD and 5-HT2A alleles was noted. Inpatient alcohol dependents showed a significant association between 5-HT2A A alleles and impulsive traits, independent of the presence of APD or BPD. No association was noted between personality disorders and the polymorphism. This is the first report about an association of 5-HT2A promoter

AKT increases VEGF expression in tumor cells by transactivating the proximal VEGF promoter

International Nuclear Information System (INIS)

Pore, N.; Bernhard, E.J.; Shu, H.-K.; Li, B.; O'Rourke, D.M.; Maity, A.; Haas-Kogan, D.

2003-01-01

Vascular endothelial growth factor (VEGF) is overexpressed in many cancers including glioblastomas and may contribute to their growth. Epidermal growth factor receptor (EGFR) amplification and loss of PTEN, commonly found in glioblastomas leading to increase phosphatidylinositol-3-kinase (PI3K) activity and VEGF expression. In the current study we show that AKT, which is downstream of PI3K, regulates VEGF expression. U87MG human glioblastoma cells lack wildtype PTEN and express high levels of phosphorylated AKT. Over expression of AKT either by stable expression in immortalized human astrocytes or by transduction with adenovirus containing activated myristoylated AKT in SF188 glioblastoma cells increases VEGF expression. Moreover the elevation of angiogenesis by constitutively expressed AKT is further confirmed by in vivo matrigel plug assay in nude mice. The upregulation of VEGF by AKT is mediated through a region in the proximal promoter located between -88 and -70 (+1 is transcription start site). In transient transfection activity of a luciferase reporter containing the -88/+54 region of the VEGF promoter is increased by cotransfection with myristoylated AKT and downregulated by a dominant negative AKT expression vector. Mutation of the putative Sp1 binding sites located in the -88/-70 region we show that AKT acts through Sp1 to transactivate the VEGF promoter. Cotransfection of the VEGF promoter reporter with both Sp1 and myristoylated AKT expression vectors increases promoter activity to a greater extent than either Sp1 or Akt by itself. In vivo phosphate labeling of proteins reveals that AKT leads to increased Sp1 phosphorylation. Gel shift assays using a radio labeled probe corresponding to nucleotides -88 through -66 in the promoter show increased binding with nuclear extracts from cells transduced with adenovirus expressing myristoylated AKT. In conclusion, our results suggest that loss of PTEN leads to increased VEGF expression by increasing AKT

SH2-B promotes insulin receptor substrate 1 (IRS1)- and IRS2-mediated activation of the phosphatidylinositol 3-kinase pathway in response to leptin.

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Duan, Chaojun; Li, Minghua; Rui, Liangyou

2004-10-15

Leptin regulates energy homeostasis primarily by binding and activating its long form receptor (LRb). Deficiency of either leptin or LRb causes morbid obesity. Leptin stimulates LRb-associated JAK2, thus initiating multiple pathways including the Stat3 and phosphatidylinositol (PI) 3-kinase pathways that mediate leptin biological actions. Here we report that SH2-B, a JAK2-interacting protein, promotes activation of the PI 3-kinase pathway by recruiting insulin receptor substrate 1 (IRS1) and IRS2 in response to leptin. SH2-B directly bound, via its PH and SH2 domain, to both IRS1 and IRS2 both in vitro and in intact cells and mediated formation of a JAK2/SH2-B/IRS1 or IRS2 tertiary complex. Consequently, SH2-B dramatically enhanced leptin-stimulated tyrosine phosphorylation of IRS1 and IRS2 in HEK293 cells stably expressing LRb, thus promoting association of IRS1 and IRS2 with the p85 regulatory subunit of PI 3-kinase and phosphorylation and activation of Akt. SH2-B mutants with lower affinity for IRS1 and IRS2 exhibited reduced ability to promote association of JAK2 with IRS1, tyrosine phosphorylation of IRS1, and association of IRS1 with p85 in response to leptin. Moreover, deletion of the SH2-B gene impaired leptin-stimulated tyrosine phosphorylation of endogenous IRS1 in mouse embryonic fibroblasts (MEF), which was reversed by reintroduction of SH2-B. Similarly, SH2-B promoted growth hormone-stimulated tyrosine phosphorylation of IRS1 in both HEK293 and MEF cells. Our data suggest that SH2-B is a novel mediator of the PI 3-kinase pathway in response to leptin or other hormones and cytokines that activate JAK2.

Diphenylarsinic acid, a chemical warfare-related neurotoxicant, promotes liver carcinogenesis via activation of aryl hydrocarbon receptor signaling and consequent induction of oxidative DAN damage in rats

International Nuclear Information System (INIS)

Wei, Min; Yamada, Takanori; Yamano, Shotaro; Kato, Minoru; Kakehashi, Anna; Fujioka, Masaki; Tago, Yoshiyuki; Kitano, Mistuaki; Wanibuchi, Hideki

2013-01-01

Diphenylarsinic acid (DPAA), a chemical warfare-related neurotoxic organic arsenical, is present in the groundwater and soil in some regions of Japan due to illegal dumping after World War II. Inorganic arsenic is carcinogenic in humans and its organic arsenic metabolites are carcinogenic in animal studies, raising serious concerns about the carcinogenicity of DPAA. However, the carcinogenic potential of DPAA has not yet been evaluated. In the present study we found that DPAA significantly enhanced the development of diethylnitrosamine-induced preneoplastic lesions in the liver in a medium-term rat liver carcinogenesis assay. Evaluation of the expression of cytochrome P450 (CYP) enzymes in the liver revealed that DPAA induced the expression of CYP1B1, but not any other CYP1, CYP2, or CYP3 enzymes, suggesting that CYP1B1 might be the enzyme responsible for the metabolic activation of DPAA. We also found increased oxidative DNA damage, possibly due to elevated CYP1B1 expression. Induction of CYP1B1 has generally been linked with the activation of AhR, and we found that DPAA activates the aryl hydrocarbon receptor (AhR). Importantly, the promotion effect of DPAA was observed only at a dose that activated the AhR, suggesting that activation of AhR and consequent induction of AhR target genes and oxidative DNA damage plays a vital role in the promotion effects of DPAA. The present study provides, for the first time, evidence regarding the carcinogenicity of DPAA and indicates the necessity of comprehensive evaluation of its carcinogenic potential using long-term carcinogenicity studies. - Highlights: • DPAA, an environmental neurotoxicant, promotes liver carcinogenesis in rats. • DPAA is an activator of AhR signaling pathway. • DPAA promoted oxidative DNA damage in rat livers. • AhR target gene CYP 1B1 might be involved in the metabolism of DPAA

Diphenylarsinic acid, a chemical warfare-related neurotoxicant, promotes liver carcinogenesis via activation of aryl hydrocarbon receptor signaling and consequent induction of oxidative DAN damage in rats

Energy Technology Data Exchange (ETDEWEB)

Wei, Min; Yamada, Takanori; Yamano, Shotaro; Kato, Minoru; Kakehashi, Anna; Fujioka, Masaki; Tago, Yoshiyuki; Kitano, Mistuaki; Wanibuchi, Hideki, E-mail: wani@med.osaka-cu.ac.jp

2013-11-15

Diphenylarsinic acid (DPAA), a chemical warfare-related neurotoxic organic arsenical, is present in the groundwater and soil in some regions of Japan due to illegal dumping after World War II. Inorganic arsenic is carcinogenic in humans and its organic arsenic metabolites are carcinogenic in animal studies, raising serious concerns about the carcinogenicity of DPAA. However, the carcinogenic potential of DPAA has not yet been evaluated. In the present study we found that DPAA significantly enhanced the development of diethylnitrosamine-induced preneoplastic lesions in the liver in a medium-term rat liver carcinogenesis assay. Evaluation of the expression of cytochrome P450 (CYP) enzymes in the liver revealed that DPAA induced the expression of CYP1B1, but not any other CYP1, CYP2, or CYP3 enzymes, suggesting that CYP1B1 might be the enzyme responsible for the metabolic activation of DPAA. We also found increased oxidative DNA damage, possibly due to elevated CYP1B1 expression. Induction of CYP1B1 has generally been linked with the activation of AhR, and we found that DPAA activates the aryl hydrocarbon receptor (AhR). Importantly, the promotion effect of DPAA was observed only at a dose that activated the AhR, suggesting that activation of AhR and consequent induction of AhR target genes and oxidative DNA damage plays a vital role in the promotion effects of DPAA. The present study provides, for the first time, evidence regarding the carcinogenicity of DPAA and indicates the necessity of comprehensive evaluation of its carcinogenic potential using long-term carcinogenicity studies. - Highlights: • DPAA, an environmental neurotoxicant, promotes liver carcinogenesis in rats. • DPAA is an activator of AhR signaling pathway. • DPAA promoted oxidative DNA damage in rat livers. • AhR target gene CYP 1B1 might be involved in the metabolism of DPAA.

Genetic polymorphisms within tumor necrosis factor gene promoter region: a role for susceptibility to ventilator-associated pneumonia.

Science.gov (United States)

Kotsaki, Antigoni; Raftogiannis, Maria; Routsi, Christina; Baziaka, Fotini; Kotanidou, Anastasia; Antonopoulou, Anastasia; Orfanos, Stylianos E; Katsenos, Chrisostomos; Koutoukas, Pantelis; Plachouras, Diamantis; Mandragos, Konstantinos; Giamarellos-Bourboulis, Evangelos J

2012-08-01

Debatable findings exist among various studies regarding the impact of single nucleotide polymorphisms (SNPs) within the promoter region of the tumor necrosis factor (TNF) gene for susceptibility to infections. Their impact was investigated in a cohort of mechanically ventilated patients who developed ventilator-associated pneumonia (VAP). Two-hundred and thirteen mechanically ventilated patients who developed VAP were enrolled. Genomic DNA was extracted and SNPs at the -376, -308 and -238 position of the promoter region of the TNF gene were assessed by restriction fragment length polymorphisms. Monocytes were isolated from 47 patients when they developed sepsis and stimulated by bacterial endotoxin for the production of TNFα and of interleukin-6 (IL-6). Patients were divided into two groups; 166 patients bearing only wild-type alleles of all three studied polymorphisms; and 47 patients carrying at least one A allele of the three studied SNPs. Time between start of mechanical ventilation and advent of VAP was significantly shorter in the second group than in the first group (log-rank: 4.416, p: 0.041). When VAP supervened, disease severity did not differ between groups. Stimulation of TNFα and of IL-6 was much greater by monocytes for patients carrying A alleles. Carriage of at least one A allele of the three studied SNPs at the promoter region of the TNF-gene is associated with shorter time to development of VAP but it is not associated with disease severity. Findings may be related with a role of the studied SNPs in the production of pro-inflammatory cytokines. Copyright © 2012 Elsevier Ltd. All rights reserved.

Sensory Gating and Alpha-7 Nicotinic Receptor Gene Allelic Variants in Schizoaffective Disorder, Bipolar Type

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Martin, Laura F.; Leonard, Sherry; Hall, Mei-Hua; Tregellas, Jason R.; Freedman, Robert; Olincy, Ann

2011-01-01

Objectives Single nucleotide allelic variants in the promoter region of the chromosome 15 alpha-7 acetylcholine nicotinic receptor gene (CHRNA7) are associated with both schizophrenia and the P50 auditory evoked potential sensory gating deficit. The purpose of this study was to determine if CHRNA7 promoter allelic variants are also associated with abnormal P50 ratios in persons with schizoaffective disorder, bipolar type. Methods P50 auditory evoked potentials were recorded in a paired stimulus paradigm in 17 subjects with schizoaffective disorder, bipolar type. The P50 test to conditioning ratio was used as the measure of sensory gating. Mutation screening of the CHRNA7 promoter region was performed on the subjects’ DNA samples. Comparisons to previously obtained data from persons with schizophrenia and controls were made. Results Subjects with schizophrenia, regardless of allele status, had an abnormal mean P50 ratio. Subjects with schizoaffective disorder, bipolar type and a variant allele had an abnormal mean P50 ratio, whereas those schizoaffective subjects with the common alleles had a normal mean P50 ratio. Normal control subjects had a normal mean ratio, but controls with variant alleles had higher P50 ratios. Conclusions In persons with bipolar type schizoaffective disorder, CHRNA7 promoter region allelic variants are linked to the capacity to inhibit the P50 auditory evoked potential and thus are associated with a type of illness genetically and biologically more similar to schizophrenia. PMID:17192894

Membrane Receptor-Induced Changes of the Protein Kinases A and C Activity May Play a Leading Role in Promoting Developmental Synapse Elimination at the Neuromuscular Junction.

Science.gov (United States)

Tomàs, Josep M; Garcia, Neus; Lanuza, Maria A; Nadal, Laura; Tomàs, Marta; Hurtado, Erica; Simó, Anna; Cilleros, Víctor

2017-01-01

Synapses that are overproduced during histogenesis in the nervous system are eventually lost and connectivity is refined. Membrane receptor signaling leads to activity-dependent mutual influence and competition between axons directly or with the involvement of the postsynaptic cell and the associated glial cell/s. Presynaptic muscarinic acetylcholine (ACh) receptors (subtypes mAChR; M 1 , M 2 and M 4 ), adenosine receptors (AR; A 1 and A 2A ) and the tropomyosin-related kinase B receptor (TrkB), among others, all cooperate in synapse elimination. Between these receptors there are several synergistic, antagonic and modulatory relations that clearly affect synapse elimination. Metabotropic receptors converge in a limited repertoire of intracellular effector kinases, particularly serine protein kinases A and C (PKA and PKC), to phosphorylate protein targets and bring about structural and functional changes leading to axon loss. In most cells A 1 , M 1 and TrkB operate mainly by stimulating PKC whereas A 2A , M 2 and M 4 inhibit PKA. We hypothesize that a membrane receptor-induced shifting in the protein kinases A and C activity (inhibition of PKA and/or stimulation of PKC) in some nerve endings may play an important role in promoting developmental synapse elimination at the neuromuscular junction (NMJ). This hypothesis is supported by: (i) the tonic effect (shown by using selective inhibitors) of several membrane receptors that accelerates axon loss between postnatal days P5-P9; (ii) the synergistic, antagonic and modulatory effects (shown by paired inhibition) of the receptors on axonal loss; (iii) the fact that the coupling of these receptors activates/inhibits the intracellular serine kinases; and (iv) the increase of the PKA activity, the reduction of the PKC activity or, in most cases, both situations simultaneously that presumably occurs in all the situations of singly and paired inhibition of the mAChR, AR and TrkB receptors. The use of transgenic animals and

Membrane Receptor-Induced Changes of the Protein Kinases A and C Activity May Play a Leading Role in Promoting Developmental Synapse Elimination at the Neuromuscular Junction

Directory of Open Access Journals (Sweden)

Josep M. Tomàs

2017-08-01

Full Text Available Synapses that are overproduced during histogenesis in the nervous system are eventually lost and connectivity is refined. Membrane receptor signaling leads to activity-dependent mutual influence and competition between axons directly or with the involvement of the postsynaptic cell and the associated glial cell/s. Presynaptic muscarinic acetylcholine (ACh receptors (subtypes mAChR; M1, M2 and M4, adenosine receptors (AR; A1 and A2A and the tropomyosin-related kinase B receptor (TrkB, among others, all cooperate in synapse elimination. Between these receptors there are several synergistic, antagonic and modulatory relations that clearly affect synapse elimination. Metabotropic receptors converge in a limited repertoire of intracellular effector kinases, particularly serine protein kinases A and C (PKA and PKC, to phosphorylate protein targets and bring about structural and functional changes leading to axon loss. In most cells A1, M1 and TrkB operate mainly by stimulating PKC whereas A2A, M2 and M4 inhibit PKA. We hypothesize that a membrane receptor-induced shifting in the protein kinases A and C activity (inhibition of PKA and/or stimulation of PKC in some nerve endings may play an important role in promoting developmental synapse elimination at the neuromuscular junction (NMJ. This hypothesis is supported by: (i the tonic effect (shown by using selective inhibitors of several membrane receptors that accelerates axon loss between postnatal days P5–P9; (ii the synergistic, antagonic and modulatory effects (shown by paired inhibition of the receptors on axonal loss; (iii the fact that the coupling of these receptors activates/inhibits the intracellular serine kinases; and (iv the increase of the PKA activity, the reduction of the PKC activity or, in most cases, both situations simultaneously that presumably occurs in all the situations of singly and paired inhibition of the mAChR, AR and TrkB receptors. The use of transgenic animals and various

Cryptocephal, the Drosophila melanogaster ATF4, is a specific coactivator for ecdysone receptor isoform B2.

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Sebastien A Gauthier

Full Text Available The ecdysone receptor is a heterodimer of two nuclear receptors, the Ecdysone receptor (EcR and Ultraspiracle (USP. In Drosophila melanogaster, three EcR isoforms share common DNA and ligand-binding domains, but these proteins differ in their most N-terminal regions and, consequently, in the activation domains (AF1s contained therein. The transcriptional coactivators for these domains, which impart unique transcriptional regulatory properties to the EcR isoforms, are unknown. Activating transcription factor 4 (ATF4 is a basic-leucine zipper transcription factor that plays a central role in the stress response of mammals. Here we show that Cryptocephal (CRC, the Drosophila homolog of ATF4, is an ecdysone receptor coactivator that is specific for isoform B2. CRC interacts with EcR-B2 to promote ecdysone-dependent expression of ecdysis-triggering hormone (ETH, an essential regulator of insect molting behavior. We propose that this interaction explains some of the differences in transcriptional properties that are displayed by the EcR isoforms, and similar interactions may underlie the differential activities of other nuclear receptors with distinct AF1-coactivators.

The putative imidazoline receptor agonist, harmane, promotes intracellular calcium mobilisation in pancreatic beta-cells.

Science.gov (United States)

Squires, Paul E; Hills, Claire E; Rogers, Gareth J; Garland, Patrick; Farley, Sophia R; Morgan, Noel G

2004-10-06

beta-Carbolines (including harmane and pinoline) stimulate insulin secretion by a mechanism that may involve interaction with imidazoline I(3)-receptors but which also appears to be mediated by actions that are additional to imidazoline receptor agonism. Using the MIN6 beta-cell line, we now show that both the imidazoline I(3)-receptor agonist, efaroxan, and the beta-carboline, harmane, directly elevate cytosolic Ca(2+) and increase insulin secretion but that these responses display different characteristics. In the case of efaroxan, the increase in cytosolic Ca(2+) was readily reversible, whereas, with harmane, the effect persisted beyond removal of the agonist and resulted in the development of a repetitive train of Ca(2+)-oscillations whose frequency, but not amplitude, was concentration-dependent. Initiation of the Ca(2+)-oscillations by harmane was independent of extracellular calcium but was sensitive to both dantrolene and high levels (20 mM) of caffeine, suggesting the involvement of ryanodine receptor-gated Ca(2+)-release. The expression of ryanodine receptor-1 and ryanodine receptor-2 mRNA in MIN6 cells was confirmed using reverse transcription-polymerase chain reaction (RT-PCR) and, since low concentrations of caffeine (1 mM) or thimerosal (10 microM) stimulated increases in [Ca(2+)](i), we conclude that ryanodine receptors are functional in these cells. Furthermore, the increase in insulin secretion induced by harmane was attenuated by dantrolene, consistent with the involvement of ryanodine receptors in mediating this response. By contrast, the smaller insulin secretory response to efaroxan was unaffected by dantrolene. Harmane-evoked changes in cytosolic Ca(2+) were maintained by nifedipine-sensitive Ca(2+)-influx, suggesting the involvement of L-type voltage-gated Ca(2+)-channels. Taken together, these data imply that harmane may interact with ryanodine receptors to generate sustained Ca(2+)-oscillations in pancreatic beta-cells and that this effect

The cytoplasmic domain close to the transmembrane region of the glucagon-like peptide-1 receptor contains sequence elements that regulate agonist-dependent internalisation.

Science.gov (United States)

Vázquez, Patricia; Roncero, Isabel; Blázquez, Enrique; Alvarez, Elvira

2005-07-01

In order to gain better insight into the molecular events involved in the signal transduction generated through glucagon-like peptide-1 (GLP-1) receptors, we tested the effect of deletions and point mutations within the cytoplasmic tail of this receptor with a view to establishing relationships between signal transduction desensitisation and receptor internalisation. Wild-type and truncated (deletion of the last 27 amino acids (GLPR 435R) and deletion of 44 amino acids (GLPR 418R)) GLP-1 receptors bound the agonist with similar affinity. Deletion of the last 27 amino acids decreased the internalisation rate by 78%, while deletion of 44 amino acids containing all the phosphorylation sites hitherto described in this receptor decreased the internalisation rate by only 47%. Binding of the ligand to both receptors stimulated adenylyl cyclase. In contrast, deletion of the region containing amino acids 419 to 435 (GLPR 419delta435) increased the internalisation rate by 268%, and the replacement of EVQ(408-410) by alanine (GLPR A(408-410)) increased this process to 296%. In both receptors, the efficacy in stimulating adenylate cyclase was decreased. All the receptors studied were internalised by coated pits, except for the receptor with a deletion of the last 44 amino acids, which also had a faster resensitisation rate. Our findings indicate that the neighbouring trans-membrane domain of the carboxyl-terminal tail of the GLP-1 receptor contains sequence elements that regulate agonist-dependent internalisation and transmembrane signalling.

Mutational studies reveal a complex set of positive and negative control elements within the chicken vitellogenin II promoter.

Science.gov (United States)

Seal, S N; Davis, D L; Burch, J B

1991-05-01

The endogenous chicken vitellogenin II (VTGII) gene is transcribed exclusively in hepatocytes in response to estrogen. We previously identified two estrogen response elements (EREs) upstream of this gene. We now present an analysis of the VTGII promoter activated by these EREs in response to estrogen. Chimeric VTGII-CAT genes were cotransfected into LMH chicken hepatoma cells along with an estrogen receptor expression vector, and transient CAT expression was assayed after culturing the cells in the absence or presence of estrogen. An analysis of constructs bearing deletions downstream of the more proximal ERE indicated that promoter elements relevant to transcription in LMH cells extend to between -113 and -96. The relative importance of sequences within the VTGII promoter was examined by using 10 contiguous linker scanner mutations spanning the region from -117 to -24. Although most of these mutations compromised VTGII promoter function, one dramatically increased expression in LMH cells and also rendered the VTGII promoter capable of being activated by cis-linked EREs in fibroblasts cotransfected with an estrogen receptor expression vector. Gel retardation and DNase I footprinting assays revealed four factor-binding sites within this promoter. We demonstrate that three of these sites bind C/EBP, SP1, and USF (or related factors), respectively; the fourth site binds a factor that we denote TF-V beta. The biological relevance of these findings is suggested by the fact that three of these binding sites map to sites previously shown to be occupied in vivo in response to estrogen.

Modelling the interdependence between the stoichiometry of receptor oligomerization and ligand binding for a coexisting dimer/tetramer receptor system.

Science.gov (United States)

Rovira, X; Vivó, M; Serra, J; Roche, D; Strange, P G; Giraldo, J

2009-01-01

Many G protein-coupled receptors have been shown to exist as oligomers, but the oligomerization state and the effects of this on receptor function are unclear. For some G protein-coupled receptors, in ligand binding assays, different radioligands provide different maximal binding capacities. Here we have developed mathematical models for co-expressed dimeric and tetrameric species of receptors. We have considered models where the dimers and tetramers are in equilibrium and where they do not interconvert and we have also considered the potential influence of the ligands on the degree of oligomerization. By analogy with agonist efficacy, we have considered ligands that promote, inhibit or have no effect on oligomerization. Cell surface receptor expression and the intrinsic capacity of receptors to oligomerize are quantitative parameters of the equations. The models can account for differences in the maximal binding capacities of radioligands in different preparations of receptors and provide a conceptual framework for simulation and data fitting in complex oligomeric receptor situations.

Similarities in transcription factor IIIC subunits that bind to the posterior regions of internal promoters for RNA polymerase III

Directory of Open Access Journals (Sweden)

Matsutani Sachiko

2004-08-01

Full Text Available Abstract Background In eukaryotes, RNA polymerase III (RNAP III transcribes the genes for small RNAs like tRNAs, 5S rRNA, and several viral RNAs, and short interspersed repetitive elements (SINEs. The genes for these RNAs and SINEs have internal promoters that consist of two regions. These two regions are called the A and B blocks. The multisubunit transcription factor TFIIIC is required for transcription initiation of RNAP III; in transcription of tRNAs, the B-block binding subunit of TFIIIC recognizes a promoter. Although internal promoter sequences are conserved in eukaryotes, no evidence of homology between the B-block binding subunits of vertebrates and yeasts has been reported previously. Results Here, I reported the results of PSI-BLAST searches using the B-block binding subunits of human and Shizosacchromyces pombe as queries, showing that the same Arabidopsis proteins were hit with low E-values in both searches. Comparison of the convergent iterative alignments obtained by these PSI-BLAST searches revealed that the vertebrate, yeast, and Arabidopsis proteins have similarities in their N-terminal one-third regions. In these regions, there were three domains with conserved sequence similarities, one located in the N-terminal end region. The N-terminal end region of the B-block binding subunit of Saccharomyces cerevisiae is tentatively identified as a HMG box, which is the DNA binding motif. Although I compared the alignment of the N-terminal end regions of the B-block binding subunits, and their homologs, with that of the HMG boxes, it is not clear whether they are related. Conclusion Molecular phylogenetic analyses using the small subunit rRNA and ubiquitous proteins like actin and α-tubulin, show that fungi are more closely related to animals than either is to plants. Interestingly, the results obtained in this study show that, with respect to the B-block binding subunits of TFIIICs, animals appear to be evolutionarily closer to plants

Similarities in transcription factor IIIC subunits that bind to the posterior regions of internal promoters for RNA polymerase III.

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Matsutani, Sachiko

2004-08-09

In eukaryotes, RNA polymerase III (RNAP III) transcribes the genes for small RNAs like tRNAs, 5S rRNA, and several viral RNAs, and short interspersed repetitive elements (SINEs). The genes for these RNAs and SINEs have internal promoters that consist of two regions. These two regions are called the A and B blocks. The multisubunit transcription factor TFIIIC is required for transcription initiation of RNAP III; in transcription of tRNAs, the B-block binding subunit of TFIIIC recognizes a promoter. Although internal promoter sequences are conserved in eukaryotes, no evidence of homology between the B-block binding subunits of vertebrates and yeasts has been reported previously. Here, I reported the results of PSI-BLAST searches using the B-block binding subunits of human and Shizosacchromyces pombe as queries, showing that the same Arabidopsis proteins were hit with low E-values in both searches. Comparison of the convergent iterative alignments obtained by these PSI-BLAST searches revealed that the vertebrate, yeast, and Arabidopsis proteins have similarities in their N-terminal one-third regions. In these regions, there were three domains with conserved sequence similarities, one located in the N-terminal end region. The N-terminal end region of the B-block binding subunit of Saccharomyces cerevisiae is tentatively identified as a HMG box, which is the DNA binding motif. Although I compared the alignment of the N-terminal end regions of the B-block binding subunits, and their homologs, with that of the HMG boxes, it is not clear whether they are related. Molecular phylogenetic analyses using the small subunit rRNA and ubiquitous proteins like actin and alpha-tubulin, show that fungi are more closely related to animals than either is to plants. Interestingly, the results obtained in this study show that, with respect to the B-block binding subunits of TFIIICs, animals appear to be evolutionarily closer to plants than to fungi.

Fiber mediated receptor masking in non-infected bystander cells restricts adenovirus cell killing effect but promotes adenovirus host co-existence.

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Johan Rebetz

Full Text Available The basic concept of conditionally replicating adenoviruses (CRAD as oncolytic agents is that progenies generated from each round of infection will disperse, infect and kill new cancer cells. However, CRAD has only inhibited, but not eradicated tumor growth in xenograft tumor therapy, and CRAD therapy has had only marginal clinical benefit to cancer patients. Here, we found that CRAD propagation and cancer cell survival co-existed for long periods of time when infection was initiated at low multiplicity of infection (MOI, and cancer cell killing was inefficient and slow compared to the assumed cell killing effect upon infection at high MOI. Excessive production of fiber molecules from initial CRAD infection of only 1 to 2% cancer cells and their release prior to the viral particle itself caused a tropism-specific receptor masking in both infected and non-infected bystander cells. Consequently, the non-infected bystander cells were inefficiently bound and infected by CRAD progenies. Further, fiber overproduction with concomitant restriction of adenovirus spread was observed in xenograft cancer therapy models. Besides the CAR-binding Ad4, Ad5, and Ad37, infection with CD46-binding Ad35 and Ad11 also caused receptor masking. Fiber overproduction and its resulting receptor masking thus play a key role in limiting CRAD functionality, but potentially promote adenovirus and host cell co-existence. These findings also give important clues for understanding mechanisms underlying the natural infection course of various adenoviruses.

Estradiol-promoted accumulation of receptor in nuclei of porcine endometrium cells. Immunogold electron microscopy of resting and estradiol-stimulated cells.

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Sierralta, W D; Jakob, F; Thole, H; Engel, P; Jungblut, P W

1992-01-01

Endometrium was collected by curettage from castrated pigs, either untreated or exposed to estradiol in vivo by intrauterine injection, and processed for electron microscopy. The resin LR Gold was used for embedding, and sections were floated on droplets of 10 nm diameter gold particles, coated with the immunoglobulin-G1 (IgG1) fraction or its Fab2 fragment of a monospecific polyclonal antiserum raised in goats against the C-terminal half of the estradiol receptor. On average, only one gold particle per microns 2 became attached in the cytoplasmic area of untreated cells, whereas four were found over the nuclear area. These figures rose to 2-3/microns 2 and 15-26/microns 2, respectively, within 10 min after exposure to estradiol. The labeling intensities of nuclei in cell clusters and of coprocessed nuclei released from cells ruptured during curettage were identical in all situations. Nuclear pores were frequently tagged after estradiol treatment. The proportions of tagging densities in nuclei of untreated and estradiol-exposed cells corresponded to those of receptor contents measured in extracts of isolated nuclei by ligand binding. This correlation was not seen for the cytoplasmic compartment of untreated cells, the scarce tagging of which is interpreted by hidden antigenic determinants. Our morphological analyses support the conclusions drawn from biochemical data (Sierralta et al., 1992) of an estradiol-promoted translocation of receptor from the cytoplasm into the nucleus.

Self-renewal of human embryonic stem cells requires insulin-like growth factor-1 receptor and ERBB2 receptor signaling

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Wang, Linlin; Schulz, Thomas C.; Sherrer, Eric S.; Dauphin, Derek S.; Shin, Soojung; Nelson, Angelique M.; Ware, Carol B.; Zhan, Mei; Song, Chao-Zhong; Chen, Xiaoji; Brimble, Sandii N.; McLean, Amanda; Galeano, Maria J.; Uhl, Elizabeth W.; D'Amour, Kevin A.; Chesnut, Jonathan D.; Rao, Mahendra S.

2007-01-01

Despite progress in developing defined conditions for human embryonic stem cell (hESC) cultures, little is known about the cell-surface receptors that are activated under conditions supportive of hESC self-renewal. A simultaneous interrogation of 42 receptor tyrosine kinases (RTKs) in hESCs following stimulation with mouse embryonic fibroblast (MEF) conditioned medium (CM) revealed rapid and prominent tyrosine phosphorylation of insulin receptor (IR) and insulin-like growth factor-1 receptor (IGF1R); less prominent tyrosine phosphorylation of epidermal growth factor receptor (EGFR) family members, including ERBB2 and ERBB3; and trace phosphorylation of fibroblast growth factor receptors. Intense IGF1R and IR phosphorylation occurred in the absence of MEF conditioning (NCM) and was attributable to high concentrations of insulin in the proprietary KnockOut Serum Replacer (KSR). Inhibition of IGF1R using a blocking antibody or lentivirus-delivered shRNA reduced hESC self-renewal and promoted differentiation, while disruption of ERBB2 signaling with the selective inhibitor AG825 severely inhibited hESC proliferation and promoted apoptosis. A simple defined medium containing an IGF1 analog, heregulin-1β (a ligand for ERBB2/ERBB3), fibroblast growth factor-2 (FGF2), and activin A supported long-term growth of multiple hESC lines. These studies identify previously unappreciated RTKs that support hESC proliferation and self-renewal, and provide a rationally designed medium for the growth and maintenance of pluripotent hESCs. PMID:17761519

GABA and benzodiazepine receptors in the gerbil brain after transient ischemia: demonstration by quantitative receptor autoradiography

International Nuclear Information System (INIS)

Onodera, H.; Sato, G.; Kogure, K.

1987-01-01

Quantitative receptor autoradiography was used to measure the binding of gamma-aminobutyric acid (GABA) and benzodiazepine receptors after ischemia by means of transient occlusion of bilateral common carotid arteries in the gerbil. [ 3 H]Muscimol was used to label the GABAA receptors and [ 3 H]flunitrazepam to label central type benzodiazepine receptors. In the superolateral convexities of the frontal cortices, [ 3 H]muscimol binding was increased in 60% of the animals killed 3 days after ischemia, and decreased in 67% of the animals killed 27 days after ischemia. Twenty-seven days after ischemia, [ 3 H]flunitrazepam binding in the substantia nigra pars reticulata increased to 252% of the control, though the increase in [ 3 H]muscimol binding was not significant. In the dorsolateral region of the caudate putamen, marked neuronal necrosis and depletion of both [ 3 H]muscimol and [ 3 H]flunitrazepam binding sites were observed 27 days after ischemia, the ventromedial region being left intact. In spite of the depletion of pyramidal cells in the CA1 region of the hippocampus, both [ 3 H]muscimol and [ 3 H]flunitrazepam binding sites were preserved 27 days after ischemia. Since our previous study revealed that adenosine A1 binding sites were depleted in the CA1 subfield of the hippocampus after ischemia correlating with neuronal damage, GABAA and benzodiazepine receptors may not be distributed predominantly on the pyramidal cells in the CA1 region

Melanocortin 1 Receptor Deficiency Promotes Atherosclerosis in Apolipoprotein E-/- Mice.

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Rinne, Petteri; Kadiri, James J; Velasco-Delgado, Mauricio; Nuutinen, Salla; Viitala, Miro; Hollmén, Maija; Rami, Martina; Savontaus, Eriika; Steffens, Sabine

2018-02-01

The MC1-R (melanocortin 1 receptor) is expressed by monocytes and macrophages where it mediates anti-inflammatory actions. MC1-R also protects against macrophage foam cell formation primarily by promoting cholesterol efflux through the ABCA1 (ATP-binding cassette transporter subfamily A member 1) and ABCG1 (ATP-binding cassette transporter subfamily G member 1). In this study, we aimed to investigate whether global deficiency in MC1-R signaling affects the development of atherosclerosis. Apoe -/- (apolipoprotein E deficient) mice were crossed with recessive yellow (Mc1r e/e ) mice carrying dysfunctional MC1-R and fed a high-fat diet to induce atherosclerosis. Apoe -/- Mc1r e/e mice developed significantly larger atherosclerotic lesions in the aortic sinus and in the whole aorta compared with Apoe -/- controls. In terms of plaque composition, MC1-R deficiency was associated with less collagen and smooth muscle cells and increased necrotic core, indicative of more vulnerable lesions. These changes were accompanied by reduced Abca1 and Abcg1 expression in the aorta. Furthermore, Apoe -/- Mc1r e/e mice showed a defect in bile acid metabolism that aggravated high-fat diet-induced hypercholesterolemia and hepatic lipid accumulation. Flow cytometric analysis of leukocyte profile revealed that dysfunctional MC1-R enhanced arterial accumulation of classical Ly6C high monocytes and macrophages, effects that were evident in mice fed a normal chow diet but not under high-fat diet conditions. In support of enhanced arterial recruitment of Ly6C high monocytes, these cells had increased expression of L-selectin and P-selectin glycoprotein ligand 1. The present study highlights the importance of MC1-R in the development of atherosclerosis. Deficiency in MC1-R signaling exacerbates atherosclerosis by disturbing cholesterol handling and by increasing arterial monocyte accumulation. © 2017 The Authors.

Nuclear proteins interacting with the promoter region of the human granulocyte/macrophage colony-stimulating factor gene

International Nuclear Information System (INIS)

Shannon, M.F.; Gamble, J.R.; Vadas, M.A.

1988-01-01

The gene for human granulocyte/macrophage colony-stimulating factor (GM-CSF) is expressed in a tissue-specific as well as an activation-dependent manner. The interaction of nuclear proteins with the promoter region of the GM-CSF gene that is likely to be responsible for this pattern of GM-CSF expression was investigated. The authors show that nuclear proteins interact with DNA fragments from the GM-CSF promoter in a cell-specific manner. A region spanning two cytokine-specific sequences, cytokine 1 (CK-1, 5', GAGATTCCAC 3') and cytokine 2 (CK-2, 5' TCAGGTA 3') bound two nuclear proteins from GM-CSF-expressing cells in gel retardation assays. NF-GMb was inducible with phorbol 12-myristate 13-acetate and accompanied induction of GM-CSF message. NF-GMb was absent in cell lines not producing GM-CSF, some of which had other distinct binding proteins. NF-GMa and NF-GMb eluted from a heparin-Sepharose column at 0.3 and 0.6 M KCl, respectively. They hypothesize that the sequences CK-1 and CK-2 bind specific proteins and regulate GM-CSF transcription

US-India Technical Collaboration to Promote Regional Stability

International Nuclear Information System (INIS)

Killinger, Mark H.; Griggs, James R.; Apt, Kenneth E.; Doyle, James E.

2001-01-01

Two US-India documents were signed in 2000 that provided new impetus for scientific and technical cooperation between the two countries. The first document is the US-India Science and Technology Agreement, which is aimed at 'promoting scientific and technological cooperation between the people of their two countries.' The second is the US-India Joint Statement on Energy and Environment, which states 'the United States and India believe that energy and environment could be one of the most important areas of cooperation between the two countries.' In addition to the work already underway as part of these two agreements, DOE has established a US-India Science and Technology Initiative to utilize the expertise of DOE national laboratories to conduct activities that support US policy objectives in South Asia. PNNL and LANL are working with US agencies and the Indian government and scientific communities to identify appropriate non-sensitive areas for US-Indian technical collaboration. The objectives of such collaboration are to address visible national/international problems, build trust between the United States and India, and contribute to regional stability in South Asia. This paper describes research done on the Indian scientific organization and infrastructure, potential areas for collaboration, the approach for this engagement, and current status of the initiative.

US-INDIA TECHNICAL COLLABORATION TO PROMOTE REGIONAL STABILITY

International Nuclear Information System (INIS)

Killinger, M.H.; Griggs, J.R.; Apt, Kenneth E.; Doyle, J.E.

2001-01-01

Two US-India documents were signed in 2000 that provided new impetus for scientific and technical cooperation between the two countries. The first document is the US-India Science and Technology Agreement, which is aimed at 'promoting scientific and technological cooperation between the people of their two countries.' The second is the US-India Joint Statement on Energy and Environment, which states 'the United States and India believe that energy and environment could be one of the most important areas of cooperation between the two countries.' In addition to the work already underway as part of these two agreements, the US Department of Energy (DOE) has established a US-India Science and Technology Initiative to utilize the expertise of DOE national laboratories to conduct activities that support US policy objectives in South Asia. PNNL and LANL are working with US government agencies to identify appropriate non-sensitive, non-nuclear areas for US-Indian technical collaboration. The objectives of such collaboration are to address visible national and international problems, build trust between the United States and India, and contribute to regional stability in South Asia. This paper describes the approach for this engagement, the Indian scientific organization and infrastructure, potential areas for collaboration, and current status of the initiative.

β-Adrenergic receptor antagonism prevents anxiety-like behavior and microglial reactivity induced by repeated social defeat.

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Wohleb, Eric S; Hanke, Mark L; Corona, Angela W; Powell, Nicole D; Stiner, La'Tonia M; Bailey, Michael T; Nelson, Randy J; Godbout, Jonathan P; Sheridan, John F

2011-04-27

Psychosocial stress is associated with altered immune function and development of psychological disorders including anxiety and depression. Here we show that repeated social defeat in mice increased c-Fos staining in brain regions associated with fear and threat appraisal and promoted anxiety-like behavior in a β-adrenergic receptor-dependent manner. Repeated social defeat also significantly increased the number of CD11b(+)/CD45(high)/Ly6C(high) macrophages that trafficked to the brain. In addition, several inflammatory markers were increased on the surface of microglia (CD14, CD86, and TLR4) and macrophages (CD14 and CD86) after social defeat. Repeated social defeat also increased the presence of deramified microglia in the medial amygdala, prefrontal cortex, and hippocampus. Moreover, mRNA analysis of microglia indicated that repeated social defeat increased levels of interleukin (IL)-1β and reduced levels of glucocorticoid responsive genes [glucocorticoid-induced leucine zipper (GILZ) and FK506 binding protein-51 (FKBP51)]. The stress-dependent changes in microglia and macrophages were prevented by propranolol, a β-adrenergic receptor antagonist. Microglia isolated from socially defeated mice and cultured ex vivo produced markedly higher levels of IL-6, tumor necrosis factor-α, and monocyte chemoattractant protein-1 after stimulation with lipopolysaccharide compared with microglia from control mice. Last, repeated social defeat increased c-Fos activation in IL-1 receptor type-1-deficient mice, but did not promote anxiety-like behavior or microglia activation in the absence of functional IL-1 receptor type-1. These findings indicate that repeated social defeat-induced anxiety-like behavior and enhanced reactivity of microglia was dependent on activation of β-adrenergic and IL-1 receptors.

A Participatory Regional Partnership Approach to Promote Nutrition and Physical Activity Through Environmental and Policy Change in Rural Missouri.

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Barnidge, Ellen K; Baker, Elizabeth A; Estlund, Amy; Motton, Freda; Hipp, Pamela R; Brownson, Ross C

2015-06-11

Rural residents are less likely than urban and suburban residents to meet recommendations for nutrition and physical activity. Interventions at the environmental and policy level create environments that support healthy eating and physical activity. Healthier Missouri Communities (Healthier MO) is a community-based research project conducted by the Prevention Research Center in St. Louis with community partners from 12 counties in rural southeast Missouri. We created a regional partnership to leverage resources and enhance environmental and policy interventions to improve nutrition and physical activity in rural southeast Missouri. Partners were engaged in a participatory action planning process that included prioritizing, implementing, and evaluating promising evidence-based interventions to promote nutrition and physical activity. Group interviews were conducted with Healthier MO community partners post intervention to evaluate resource sharing and sustainability efforts of the regional partnership. Community partners identified the benefits and challenges of resource sharing within the regional partnership as well as the opportunities and threats to long-term partnership sustainability. The partners noted that the regional participatory process was difficult, but the benefits outweighed the challenges. Regional rural partnerships may be an effective way to leverage relationships to increase the capacity of rural communities to implement environmental and policy interventions to promote nutrition and physical activity.

N-glycosylation of the β2 adrenergic receptor regulates receptor function by modulating dimerization.

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Li, Xiaona; Zhou, Mang; Huang, Wei; Yang, Huaiyu

2017-07-01

N-glycosylation is a common post-translational modification of G-protein-coupled receptors (GPCRs). However, it remains unknown how N-glycosylation affects GPCR signaling. β 2 adrenergic receptor (β 2 AR) has three N-glycosylation sites: Asn6, Asn15 at the N-terminus, and Asn187 at the second extracellular loop (ECL2). Here, we show that deletion of the N-glycan did not affect receptor expression and ligand binding. Deletion of the N-glycan at the N-terminus rather than Asn187 showed decreased effects on isoproterenol-promoted G-protein-dependent signaling, β-arrestin2 recruitment, and receptor internalization. Both N6Q and N15Q showed decreased receptor dimerization, while N187Q did not influence receptor dimerization. As decreased β 2 AR homodimer accompanied with reduced efficiency for receptor function, we proposed that the N-glycosylation of β 2 AR regulated receptor function by influencing receptor dimerization. To verify this hypothesis, we further paid attention to the residues at the dimerization interface. Studies of Lys60 and Glu338, two residues at the receptor dimerization interface, exhibited that the K60A/E338A showed decreased β 2 AR dimerization and its effects on receptor signaling were similar to N6Q and N15Q, which further supported the importance of receptor dimerization for receptor function. This work provides new insights into the relationship among glycosylation, dimerization, and function of GPCRs. Peptide-N-glycosidase F (PNGase F, EC 3.2.2.11); endo-β-N-acetylglucosaminidase A (Endo-A, EC 3.2.1.96). © 2017 Federation of European Biochemical Societies.

No evidence for promoter region methylation of the succinate dehydrogenase and fumarate hydratase tumour suppressor genes in breast cancer

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Dobrovic Alexander

2009-09-01

Full Text Available Abstract Background Succinate dehydrogenase (SDH and fumarate hydratase (FH are tricarboxylic acid (TCA cycle enzymes that are also known to act as tumour suppressor genes. Increased succinate or fumarate levels as a consequence of SDH and FH deficiency inhibit hypoxia inducible factor-1α (HIF-1α prolyl hydroxylases leading to sustained HIF-1α expression in tumours. Since HIF-1α is frequently expressed in breast carcinomas, DNA methylation at the promoter regions of the SDHA, SDHB, SDHC and SDHD and FH genes was evaluated as a possible mechanism in silencing of SDH and FH expression in breast carcinomas. Findings No DNA methylation was identified in the promoter regions of the SDHA, SDHB, SDHC, SDHD and FH genes in 72 breast carcinomas and 10 breast cancer cell lines using methylation-sensitive high resolution melting which detects both homogeneous and heterogeneous methylation. Conclusion These results show that inactivation via DNA methylation of the promoter CpG islands of SDH and FH is unlikely to play a major role in sporadic breast carcinomas.

Receptor tyrosine kinase-like orphan receptor 2 (ROR2) and Indian hedgehog regulate digit outgrowth mediated by the phalanx-forming region.

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Witte, Florian; Chan, Danny; Economides, Aris N; Mundlos, Stefan; Stricker, Sigmar

2010-08-10

Elongation of the digit rays resulting in the formation of a defined number of phalanges is a process poorly understood in mammals, whereas in the chicken distal mesenchymal bone morphogenetic protein (BMP) signaling in the so-called phalanx-forming region (PFR) or digit crescent (DC) seems to be involved. The human brachydactylies (BDs) are inheritable conditions characterized by variable degrees of digit shortening, thus providing an ideal model to analyze the development and elongation of phalanges. We used a mouse model for BDB1 (Ror2(W749X/W749X)) lacking middle phalanges and show that a signaling center corresponding to the chick PFR exists in the mouse, which is diminished in BDB1 mice. This resulted in a strongly impaired elongation of the digit condensations due to reduced chondrogenic commitment of undifferentiated distal mesenchymal cells. We further show that a similar BMP-based mechanism accounts for digit shortening in a mouse model for the closely related condition BDA1 (Ihh(E95K/E95K)), altogether indicating the functional significance of the PFR in mammals. Genetic interaction experiments as well as pathway analysis in BDB1 mice suggest that Indian hedgehog and WNT/beta-catenin signaling, which we show is inhibited by receptor tyrosine kinase-like orphan receptor 2 (ROR2) in distal limb mesenchyme, are acting upstream of BMP signaling in the PFR.

Molecular analysis of the nerve growth factor receptor

International Nuclear Information System (INIS)

Hempstead, B.; Patil, N.; Olson, K.; Chao, M.

1988-01-01

An essential molecule in the translocation of information by nerve growth factor (NGF) to responsive cells is the cell-surface receptor for NGF. This paper presents information on the genomic structure of the NGF receptor gene, NGF receptor models, and transfection of NGF receptors. Equilibrium binding of [ 125 I]NGF to cells reveals two distinct affinity states for the NGF receptor. The human NGF receptor gene is a single-copy gene, consisting of six exons that span 23 kb. The receptor gene is capable of being transferred to fibroblast cells from human genomic DNA and expressed at high levels. The constitutive nature of the receptor promoter sequence is a partial explanation of why this tissue-specific gene is expressed efficiently in a variety of nonneuronal cells after genomic gene transfer. The two kinetic forms of the NGF receptor appear to be encoded by the same protein, which is the product of a single gene

Quantifying Surface Coal-Mining Patterns to Promote Regional Sustainability in Ordos, Inner Mongolia

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Xiaoji Zeng

2018-04-01

Full Text Available Ordos became the new “coal capital” of China within a few decades since the country’s economic reform in 1978, as large-scale surface coal mining dramatically propelled its per capita GDP from being one of the lowest to one of the highest in China, exceeding Hong Kong in 2009. Surface coal-mining areas (SCMAs have continued to expand in this region during recent decades, resulting in serious environmental and socioeconomic consequences. To understand these impacts and promote regional sustainability, quantifying the spatiotemporal patterns of SCMAs is urgently needed. Thus, the main objectives of this study were to quantify the spatiotemporal patterns of SCMAs in the Ordos region from 1990 to 2015, and to examine some of the major environmental and socioeconomic impacts in the study region. We extracted the SCMAs using remote-sensing data, and then quantified their spatiotemporal patterns using landscape metrics. The loss of natural habitat and several socioeconomic indicators were examined in relation to surface coal mining. Our results show that the area of SCMAs increased from 7.12 km2 to 355.95 km2, an increase of nearly 49 times from 1990 to 2015 in the Ordos region. The number of SCMAs in this region increased from 82 to 651, a nearly seven-fold increase. In particular, Zhungeer banner (an administrative division, Yijinhuoluo banner, Dongsheng District and Dalate banner in the north-eastern part of the Ordos region had higher growth rates of SCMAs. The income gap between urban and rural residents increased along with the growth in SCMAs, undermining social equity in the Ordos region. Moreover, the rapid increase in SCMAs resulted in natural habitat loss (including grasslands, forests, and deserts across this region. Thus, we suggest that regional sustainability in Ordos needs to emphasize effective measures to curb large-scale surface coal mining in order to reduce the urban–rural income gap, and to restore degraded natural

Oxysterol Sensing through the Receptor GPR183 Promotes the Lymphoid-Tissue-Inducing Function of Innate Lymphoid Cells and Colonic Inflammation.

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Emgård, Johanna; Kammoun, Hana; García-Cassani, Bethania; Chesné, Julie; Parigi, Sara M; Jacob, Jean-Marie; Cheng, Hung-Wei; Evren, Elza; Das, Srustidhar; Czarnewski, Paulo; Sleiers, Natalie; Melo-Gonzalez, Felipe; Kvedaraite, Egle; Svensson, Mattias; Scandella, Elke; Hepworth, Matthew R; Huber, Samuel; Ludewig, Burkhard; Peduto, Lucie; Villablanca, Eduardo J; Veiga-Fernandes, Henrique; Pereira, João P; Flavell, Richard A; Willinger, Tim

2018-01-16

Group 3 innate lymphoid cells (ILC3s) sense environmental signals and are critical for tissue integrity in the intestine. Yet, which signals are sensed and what receptors control ILC3 function remain poorly understood. Here, we show that ILC3s with a lymphoid-tissue-inducer (LTi) phenotype expressed G-protein-coupled receptor 183 (GPR183) and migrated to its oxysterol ligand 7α,25-hydroxycholesterol (7α,25-OHC). In mice lacking Gpr183 or 7α,25-OHC, ILC3s failed to localize to cryptopatches (CPs) and isolated lymphoid follicles (ILFs). Gpr183 deficiency in ILC3s caused a defect in CP and ILF formation in the colon, but not in the small intestine. Localized oxysterol production by fibroblastic stromal cells provided an essential signal for colonic lymphoid tissue development, and inflammation-induced increased oxysterol production caused colitis through GPR183-mediated cell recruitment. Our findings show that GPR183 promotes lymphoid organ development and indicate that oxysterol-GPR183-dependent positioning within tissues controls ILC3 activity and intestinal homeostasis. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Identification of an estrogen response element in the 3'-flanking region of the murine c-fos protooncogene.

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Hyder, S M; Stancel, G M; Nawaz, Z; McDonnell, D P; Loose-Mitchell, D S

1992-09-05

We have used transient transfection assays with reporter plasmids expressing chloramphenicol acetyltransferase, linked to regions of mouse c-fos, to identify a specific estrogen response element (ERE) in this protooncogene. This element is located in the untranslated 3'-flanking region of the c-fos gene, 5 kilobases (kb) downstream from the c-fos promoter and 1.5 kb downstream of the poly(A) signal. This element confers estrogen responsiveness to chloramphenicol acetyltransferase reporters linked to both the herpes simplex virus thymidine kinase promoter and the homologous c-fos promoter. Deletion analysis localized the response element to a 200-base pair fragment which contains the element GGTCACCACAGCC that resembles the consensus ERE sequence GGTCACAGTGACC originally identified in Xenopus vitellogenin A2 gene. A synthetic 36-base pair oligodeoxynucleotide containing this c-fos sequence conferred estrogen inducibility to the thymidine kinase promoter. The corresponding sequence also induced reporter activity when present in the c-fos gene fragment 3 kb from the thymidine kinase promoter. Gel-shift experiments demonstrated that synthetic oligonucleotides containing either the consensus ERE or the c-fos element bind human estrogen receptor obtained from a yeast expression system. However, the mobility of the shifted band is faster for the fos-ERE-complex than the consensus ERE complex suggesting that the three-dimensional structure of the protein-DNA complexes is different or that other factors are differentially involved in the two reactions. When the 5'-GGTCA sequence present in the c-fos ERE is mutated to 5'-TTTCA, transcriptional activation and receptor binding activities are both lost. Mutation of the CAGCC-3' element corresponding to the second half-site of the c-fos sequence also led to the loss of receptor binding activity, suggesting that both half-sites of this element are involved in this function. The estrogen induction mediated by either the c-fos or

Structural diversity and evolution of the N-terminal isoform-specific region of ecdysone receptor-A and -B1 isoforms in insects

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Kubo Takeo

2010-02-01

Full Text Available Abstract Background The ecdysone receptor (EcR regulates various cellular responses to ecdysteroids during insect development. Insects have multiple EcR isoforms with different N-terminal A/B domains that contain the isoform-specific activation function (AF-1 region. Although distinct physiologic functions of the EcR isoforms have been characterized in higher holometabolous insects, they remain unclear in basal direct-developing insects, in which only A isoform has been identified. To examine the structural basis of the EcR isoform-specific AF-1 regions, we performed a comprehensive structural comparison of the isoform-specific region of the EcR-A and -B1 isoforms in insects. Results The EcR isoforms were newly identified in 51 species of insects and non-insect arthropods, including direct-developing ametabolous and hemimetabolous insects. The comprehensive structural comparison revealed that the isoform-specific region of each EcR isoform contained evolutionally conserved microdomain structures and insect subgroup-specific structural modifications. The A isoform-specific region generally contained four conserved microdomains, including the SUMOylation motif and the nuclear localization signal, whereas the B1 isoform-specific region contained three conserved microdomains, including an acidic activator domain-like motif. In addition, the EcR-B1 isoform of holometabolous insects had a novel microdomain at the N-terminal end. Conclusions Given that the nuclear receptor AF-1 is involved in cofactor recruitment and transcriptional regulation, the microdomain structures identified in the isoform-specific A/B domains might function as signature motifs and/or as targets for cofactor proteins that play essential roles in the EcR isoform-specific AF-1 regions. Moreover, the novel microdomain in the isoform-specific region of the holometabolous insect EcR-B1 isoform suggests that the holometabolous insect EcR-B1 acquired additional transcriptional

Receptor type I and type II binding regions and the peptidyl-prolyl isomerase site of cyclophilin B are required for enhancement of T-lymphocyte adhesion to fibronectin.

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Carpentier, Mathieu; Allain, Fabrice; Slomianny, Marie-Christine; Durieux, Sandrine; Vanpouille, Christophe; Haendler, Bernard; Spik, Geneviève

2002-04-23

Cyclophilin B (CyPB), a cyclosporin A (CsA) binding protein, interacts with two types of binding sites at the surface of T-lymphocytes. The type I sites correspond to functional receptors involved in endocytosis and the type II sites to sulfated glycosaminoglycans (GAGs). Mutational analysis of CyPB has revealed that W128, which is part of the CsA-binding pocket, is implicated in the binding to the functional type I receptors and that two amino acid clusters located in the N-terminus ensure the binding to GAGs. The peptidyl-prolyl isomerase activity of CyPB is not required for receptor binding. We have recently demonstrated that CyPB enhances adhesion of peripheral blood T-lymphocytes to fibronectin, a component of the extracellular matrix. We intended to identify additional amino acids involved in the binding of CyPB to its functional type I receptor and to determine regions responsible for the stimulation of peripheral blood T-lymphocyte adhesion. We determined that residues R76, G77, K132, D155, and D158 of the calcineurin (CN) interacting region were implicated in the recognition of type I receptor but not of GAGs. We also found that two different changes in the N-terminal extension that abated binding to GAGs prevented adhesion of peripheral blood T-lymphocytes to coated CyPB, whereas abbrogation of the PPIase activity had no effect. On the other hand, the adhesion of peripheral blood T-lymphocytes to coated fibronectin was not stimulated by CyPB mutants devoid of either type I receptor or GAGs binding activity or by mutants of the PPIase site. Altogether, the results demonstrate that different regions of CyPB are involved in peripheral blood T-lymphocyte activation and imply a novel important physiological function for peptidyl-prolyl isomerase activity.

Role for the disulfide-bonded region of human immunodeficiency virus type 1 gp41 in receptor-triggered activation of membrane fusion function

International Nuclear Information System (INIS)

Bellamy-McIntyre, Anna K.; Baer, Severine; Ludlow, Louise; Drummer, Heidi E.; Poumbourios, Pantelis

2010-01-01

The conserved disulfide-bonded region (DSR) of the human immunodeficiency virus type 1 (HIV-1) fusion glycoprotein, gp41, mediates association with the receptor-binding glycoprotein, gp120. Interactions between gp120, CD4 and chemokine receptors activate the fusion activity of gp41. The introduction of W596L and W610F mutations to the DSR of HIV-1 QH1549.13 blocked viral entry and hemifusion without affecting gp120-gp41 association. The fusion defect correlated with inhibition of CD4-triggered gp41 pre-hairpin formation, consistent with the DSR mutations having decoupled receptor-induced conformational changes in gp120 from gp41 activation. Our data implicate the DSR in sensing conformational changes in the gp120-gp41 complex that lead to fusion activation.

G Protein-Coupled Receptor 87 (GPR87 Promotes Cell Proliferation in Human Bladder Cancer Cells

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Xia Zhang

2015-10-01

Full Text Available G protein-coupled receptor 87 (GPR87 is a newly deorphanized member of the cell surface molecule G protein-coupled receptor family. GPR signaling was shown to play a role in promotion of cell growth and survival, metastasis, and drug resistance. The overexpression of GPR87 has also been reported in many malignant tumors including bladder cancer. The aim of the present study is to examine the effect of silencing GPR87 expression with a replication-deficient recombinant adenoviral vector expressing short hairpin RNA targeting GPR87 (Ad-shGPR87 and to explore the underlying molecular mechanisms in bladder cancer cells. Six GPR87-expressing human bladder cancer cells, HT1197, HT1376, J82, RT112, TCCSUP and UMUC3, were used. Infection with Ad-shGPR87 effectively downregulated the GPR87 expression, and significantly reduced the percentage of viable cells in 4 of 6 cell lines as detected by an MTT assay. Significant inhibition on cell proliferation with Ad-shGPR87 was observed in the wild-type p53 bladder cancer cell lines (HT1197, RT112, TCCSUP and UMUC3, but not in the mutant p53 cells (HT1376 and J82. As represented by a wild-type p53 RT112 cell, Ad-shGPR87 infection significantly enhanced p53 and p21 expression and caused caspase-dependent apoptosis. Furthermore, the treatment with Ad-shGPR87 exerted a significant antitumor effect against the GPR87-expressing RT112 xenografts. GPR87 appeared to be a promising target for gene therapy, and Ad-shGPR87 had strong antitumor effects, specifically anti-proliferative and pro-apoptotic effects, against GPR87-expressing human bladder cancer cells.

Analysis of the Impact of the Flow of Migrant Workers on Regional Economy: Based on the Thought about the Promotion of Jiangxi Regional Economic Competitiveness

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Sun Yuping

2016-03-01

Full Text Available Labor resource is the necessary productive factor in regional economic development, and one of important indexes to evaluate regional economic competitiveness. The great economic achievement brought by the 30-year reform and opening up of China is due to the fact that China brought the backward advantage of “demographic dividend” into play, promoted the fast development of industrialization and urbanization, and became the second largest economy in the world. The entity of “demographic dividend” is the non-agricultural migrant population, i.e., migrant workers. The transfer employment of migrant workers has typical regional liquidity, and the imbalance of regional economy causes the flow of many migrant workers. In order to achieve harmonious development and coordinated development, underdeveloped areas must understand the character and regulation, adopt positive industrial policy and supportive policy, guide the reasonable flow of migrant workers, and realize the transfer of local employment and citizenization of migrant workers, which can enhance regional economic competitiveness

Regional increases in [11C]flumazenil binding after epilepsy surgery

International Nuclear Information System (INIS)

Savic, I.; Blomqvist, G.; Halldin, C.; Litton, J.E.; Gulyas, B.

1998-01-01

Introduction - Animal experiments suggest that epileptic seizures alter the expression of mRNA for neuro-receptors. PET measurements with [ 11 C]flumazenil show that patients with partial seizures have a reduced density of benzodiazepine (BZ) receptors in the epileptogenic regions (ER) and some of the target areas for seizure activity, the so called projection areas. Recent data suggest that the degree of BZ receptor reduction in ER is correlated to seizure frequency. We therefore hypothesized that seizure activity can alter the BZ receptor binding, and that some of these changes could normalize when the seizures disappeared. Methods - In 4 patients whose seizures were generated by mesial temporal lobe structures, BZ receptor density was measured with [ 11 C]flumazenil PET before, and 1 year after the epilepsy surgery and cessation of seizures. By use of a computerized anatomical brain atlas the same regions were analyzed in both PET scans, and the results related to data from 7 healthy controls. Results - Presurgical PET scans showed reductions in BZ receptor density in the epileptogenic regions and some of its primary projection areas. Other cortical regions had normal values. Postsurgically, the calculated BZ receptor density normalized (29±17% increase) in several of the affected projection areas, whereas the values in other cortical regions remained unaltered. Conclusion - Regional reductions in BZ receptor density may be dynamic and related to seizures. The present preliminary observations encourage further studies on seizure-related changes in regional receptor binding in humans. (au)

Opioid regulation of mu receptor internalisation: relevance to the development of tolerance and dependence.

Science.gov (United States)

Lopez-Gimenez, Juan F; Milligan, Graeme

2010-11-01

Internalisation of the mu opioid receptor from the surface of cells is generally achieved by receptor occupancy with agonist ligands of high efficacy. However, in many situations the potent analgesic morphine fails to promote internalisation effectively and whether there is a direct link between this and the propensity for the sustained use of morphine to result in both tolerance and dependence has been studied intensely. Although frequently described as a partial agonist, this characteristic appears insufficient to explain the poor capacity of morphine to promote internalisation of the mu opioid receptor. Experiments performed using both transfected cell systems and ex vivo/in vivo models have provided evidence that when morphine can promote internalisation of the mu receptor there is a decrease in the development of tolerance and dependence. Although aspects of this model are controversial, such observations suggest a number of approaches to further enhance the use of morphine as an analgesic.

Low 5-HT1B receptor binding in the migraine brain

DEFF Research Database (Denmark)

Deen, Marie; Hansen, Hanne D; Hougaard, Anders

2018-01-01

Background The pathophysiology of migraine may involve dysfunction of serotonergic signaling. In particular, the 5-HT1B receptor is considered a key player due to the efficacy of 5-HT1B receptor agonists for treatment of migraine attacks. Aim To examine the cerebral 5-HT1B receptor binding....... Patients who reported migraine brain regions involved in pain modulation as regions of interest and applied a latent variable model (LVM) to assess the group effect on binding across these regions. Results Our data...... support a model wherein group status predicts the latent variable ( p = 0.038), with migraine patients having lower 5-HT1B receptor binding across regions compared to controls. Further, in a whole-brain voxel-based analysis, time since last migraine attack correlated positively with 5-HT1B receptor...

P2X7 Receptor Function in Bone-Related Cancer

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Elena Adinolfi

2012-01-01

Full Text Available Modulation of tumor microenvironment by different mediators is central in determining neoplastic formation and progression. Among these molecules extracellular ATP is emerging as a good candidate in promoting cell growth, neovascularization, tumor-host interactions, and metastatization. This paper summarizes recent findings on expression and function of P2X7 receptor for extracellular ATP in primary and metastatic bone cancers. Search of mRNA expression microchip databases and literature analysis demonstrate a high expression of P2X7 in primary bone tumors as well as in other malignancies such as multiple myeloma, neuroblastoma, breast, and prostate cancer. Evidence that P2X7 triggers NFATc1, PI3K/Akt, ROCK, and VEGF pathways in osteoblasts promoting either primary tumor development or osteoblastic lesions is also reported. Moreover, P2X7 receptor is involved in osteoclast differentiation, RANKL expression, matrix metalloproteases and cathepsin secretion thus promoting bone resorption and osteolytic lesions. Taken together these data point to a pivotal role for the P2X7 receptor in bone cancer biology.

The role of COMESA in promoting intra-regional agricultural trade: Case study of Sudan

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Azharia Abdelbagi Elbushra

2011-06-01

Full Text Available African countries have created many regional trade agreements with the economic objectives of reducing trade barriers and encouraging economic growth. The COMESA is an example of regional integration singed on 1993 by 19 African countries including Sudan. COMESA represents a chance for member countries to enhance their economic and social relations through increasing intra-trade. The objective of this paper is to assess the role of COMESA in promoting intra-regional agricultural trade between Sudan and COMESA countries. A multi-market model with Armington non-linear specification was applied. The paper results showed that there is a great potential for Sudan to increase its agricultural exports to other COMESA countries. The domestic agricultural markets are expected to be hampered by imports surge and increase in competition, while the producers of agricultural export commodities will be better off. In order to compete and benefit from potential in the COMESA markets, the paper recommended improving efficiency in the Sudanese agricultural sector through increasing productivity, lowering cost of production, enhancing marketing services, attaining economies of scale and attracting foreign investment.

Potential mechanisms underlying estrogen-induced expression of the molluscan estrogen receptor (ER) gene

Energy Technology Data Exchange (ETDEWEB)

Tran, Thi Kim Anh [School of Environmental and Life Sciences, The University of Newcastle, Callaghan, NSW 2308 (Australia); Department of Agriculture, Forestry and Fisheries, Vinh University, 182 Le Duan St., Vinh City, Nghe An (Viet Nam); MacFarlane, Geoff R. [School of Environmental and Life Sciences, The University of Newcastle, Callaghan, NSW 2308 (Australia); Kong, Richard Yuen Chong [Department of Biology and Chemistry, City University of Hong Kong, Tat Chee Avenue, Kowloon, Hong Kong Special Administrative Region (China); O’Connor, Wayne A. [New South Wales Department of Primary Industries, Port Stephens Fisheries Institute, Taylors Beach, NSW 2316 (Australia); Yu, Richard Man Kit, E-mail: Richard.Yu@newcastle.edu.au [School of Environmental and Life Sciences, The University of Newcastle, Callaghan, NSW 2308 (Australia)

2016-10-15

Highlights: • This is the first report on the putative promoter sequence of a molluscan ER gene. • The gene promoter contains putative binding sites for direct and indirect interaction with ER. • E2 upregulates ER gene expression in the ovary in vitro and in vivo. • E2-induced gene expression may require a novel ligand-dependent receptor. • The ER proximal promoter is hypomethylated regardless of gene expression levels. - Abstract: In vertebrates, estrogens and estrogen mimicking chemicals modulate gene expression mainly through a genomic pathway mediated by the estrogen receptors (ERs). Although the existence of an ER orthologue in the mollusc genome has been known for some time, its role in estrogen signalling has yet to be deciphered. This is largely due to its constitutive (ligand-independent) activation and a limited mechanistic understanding of its regulation. To fill this knowledge gap, we cloned and characterised an ER cDNA (sgER) and the 5′-flanking region of the gene from the Sydney rock oyster Saccostrea glomerata. The sgER cDNA is predicted to encode a 477-amino acid protein that contains a DNA-binding domain (DBD) and a ligand-binding domain (LBD) typically conserved among both vertebrate and invertebrate ERs. A comparison of the sgER LBD sequence with those of other ligand-dependent ERs revealed that the sgER LBD is variable at several conserved residues known to be critical for ligand binding and receptor activation. Ligand binding assays using fluorescent-labelled E2 and purified sgER protein confirmed that sgER is devoid of estrogen binding. In silico analysis of the sgER 5′-flanking sequence indicated the presence of three putative estrogen responsive element (ERE) half-sites and several putative sites for ER-interacting transcription factors, suggesting that the sgER promoter may be autoregulated by its own gene product. sgER mRNA is ubiquitously expressed in adult oyster tissues, with the highest expression found in the ovary

Key region of laminin receptor 1 for interaction with human period 1

African Journals Online (AJOL)

GREGORY

2010-09-20

Sep 20, 2010 ... acids) through yeast two-hybrid system in the present study. And we identified the ... the cell surface and functions as a membrane receptor for the adhesive ..... Circadian modulation of dopamine receptor responsiveness in ...

NPY2-receptor variation modulates iconic memory processes.

Science.gov (United States)

Arning, Larissa; Stock, Ann-Kathrin; Kloster, Eugen; Epplen, Jörg T; Beste, Christian

2014-08-01

Sensory memory systems are modality-specific buffers that comprise information about external stimuli, which represent the earliest stage of information processing. While these systems have been the subject of cognitive neuroscience research for decades, little is known about the neurobiological basis of sensory memory. However, accumulating evidence suggests that the glutamatergic system and systems influencing glutamatergic neural transmission are important. In the current study we examine if functional promoter variations in neuropeptide Y (NPY) and its receptor gene NPY2R affect iconic memory processes using a partial report paradigm. We found that iconic memory decayed much faster in individuals carrying the rare promoter NPY2R G allele which is associated with increased expression of the Y2 receptor. Possibly this effect is due to altered presynaptic inhibition of glutamate release, known to be modulated by Y2 receptors. Altogether, our results provide evidence that the functionally relevant single nucleotide polymorphism (SNP) in the NPY2R promoter gene affect circumscribed processes of early sensory processing, i.e. only the stability of information in sensory memory buffers. This leads us to suggest that especially the stability of information in sensory memory buffers depends on glutamatergic neural transmission and factors modulating glutamatergic turnover. Copyright © 2014 Elsevier B.V. and ECNP. All rights reserved.

Effect of ARA9 on dioxin receptor mediated transcription

International Nuclear Information System (INIS)

Lees, M.J.; Whitelaw, M.L.

2002-01-01

The dioxin (Aryl hydrocarbon) receptor (DR) is a unique bHLH transcription factor which is activated by binding of planar aromatic hydrocarbons typified by dioxin (TCDD). The active receptor is key to metabolism of aryl hydrocarbon xenobiotics by being a potent inducer of CYP1A1 gene activity. Chlorinated dioxins are inert to metabolism and initiate multifarious toxicities, including potent tumour promotion. These ill-effects are mediated by the activated DR and we are studying the mechanisms by which the ligand binding domain of the DR controls activity of the protein. The DR ligand binding domain resides within a PAS (Per/Arnt/Sim homology) region which is contiguous with the bHLH. The latent bHLH/PAS dioxin receptor (DR) is found in the cytoplasm of most mammalian cell types in a complex with heat shock protein 90, a novel immunophilin like protein termed ARA9/XAP2/AIP, and the co-chaperone p23. Here we use antisense ARA9 constructs to reveal that in the absence of ARA9, the DR is unable to form a transcriptionally active complex. Co-expression of antisense ARA9 with a form of the DR which is constitutively targeted to the nucleus leads to dramatically decreased levels of the nuclear DR protein, implying that ARA9 may function beyond its currently proposed role in cytoplasmic retention of the latent DR

Influence of promoter/enhancer region haplotypes on MGMT transcriptional regulation: a potential biomarker for human sensitivity to alkylating agents.

Science.gov (United States)

Xu, Meixiang; Nekhayeva, Ilona; Cross, Courtney E; Rondelli, Catherine M; Wickliffe, Jeffrey K; Abdel-Rahman, Sherif Z

2014-03-01

The O6-methylguanine-DNA methyltransferase gene (MGMT) encodes the direct reversal DNA repair protein that removes alkyl adducts from the O6 position of guanine. Several single-nucleotide polymorphisms (SNPs) exist in the MGMT promoter/enhancer (P/E) region. However, the haplotype structure encompassing these SNPs and their functional/biological significance are currently unknown. We hypothesized that MGMT P/E haplotypes, rather than individual SNPs, alter MGMT transcription and can thus alter human sensitivity to alkylating agents. To identify the haplotype structure encompassing the MGMT P/E region SNPs, we sequenced 104 DNA samples from healthy individuals and inferred the haplotypes using the data generated. We identified eight SNPs in this region, namely T7C (rs180989103), T135G (rs1711646), G290A (rs61859810), C485A (rs1625649), C575A (rs113813075), G666A (rs34180180), C777A (rs34138162) and C1099T (rs16906252). Phylogenetics and Sequence Evolution analysis predicted 21 potential haplotypes that encompass these SNPs ranging in frequencies from 0.000048 to 0.39. Of these, 10 were identified in our study population as 20 paired haplotype combinations. To determine the functional significance of these haplotypes, luciferase reporter constructs representing these haplotypes were transfected into glioblastoma cells and their effect on MGMT promoter activity was determined. Compared with the most common (reference) haplotype 1, seven haplotypes significantly upregulated MGMT promoter activity (18-119% increase; P alkylating agents.

Cloning and functional analysis of promoters of three GnRH genes in a cichlid

International Nuclear Information System (INIS)

Kitahashi, Takashi; Sato, Hideki; Sakuma, Yasuo; Parhar, Ishwar S.

2005-01-01

Mechanisms regulating gonadotropin-releasing hormone (GnRH) types, a key molecule for reproductive physiology, remain unclear. In the present study, we cloned the promoters of GnRH1, GnRH2, and GnRH3 genes in the tilapia, Oreochromis niloticus; and found putative binding sites for glucocorticoid receptors, Sp1, C/EBP, GATA, and Oct-1, but not for androgen receptors in all three GnRH promoters using computer analysis. The presence of binding sites for progesterone receptors in GnRH1, estrogen receptors in GnRH1 and GnRH2, and thyroid hormone receptors in GnRH1 and GnRH3 suggests direct action of steroid hormones on GnRH types. Our observation of SOX and LINE-like sequences exclusively in GnRH1, COUP in GnRH2, and retinoid X receptors in GnRH3 suggests their role in sexual differentiation, midbrain segmentation, and visual cue integration, respectively. Thus, the characteristic binding sites for nuclear receptors and transcription factors support the notion that each GnRH type is regulated differently and has distinct physiological roles

Enhancement of insulin-like growth factor 2 receptors in glioblastoma

International Nuclear Information System (INIS)

Sara, V.; Prisell, Per; Sjoegren, Barbro; Enberg, Goesta

1986-01-01

The somatomedins (IGF-1/IGF-2) are a family of growth-promoting hormones which have been identified in the human central nervous system where their specific receptors are distributed. The present study identified somatomedin receptors in glioblastoma and compared them with those found in normal brain. A significant enhancement in the binding of 125 1-IGF-2 but not 125 1-IGF-1 to glioblastoma membranes was found. A fourfold increase in IGF-2 receptor concentration was observed. These findings indicate enhanced expression of the IGF-2 receptor in glioblastoma. (author)

Enhancement of insulin-like growth factor 2 receptors in glioblastoma

Energy Technology Data Exchange (ETDEWEB)

Sara, V; Prisell, P; Sjoegren, B; Persson, L; Boethius, J; Enberg, G

1986-09-01

The somatomedins (IGF-1/IGF-2) are a family of growth-promoting hormones which have been identified in the human central nervous system where their specific receptors are distributed. The present study identified somatomedin receptors in glioblastoma and compared them with those found in normal brain. A significant enhancement in the binding of /sup 125/1-IGF-2 but not /sup 125/1-IGF-1 to glioblastoma membranes was found. A fourfold increase in IGF-2 receptor concentration was observed. These findings indicate enhanced expression of the IGF-2 receptor in glioblastoma. 14 refs.

Autoradiographic localization of mu and delta opioid receptors in the mesocorticolimbic dopamine system

Energy Technology Data Exchange (ETDEWEB)

Dilts, R.P. Jr.

1989-01-01

In vitro autoradiographic techniques were coupled with selective chemical lesions of the A10 dopamine cells and intrinsic perikarya of the region to delineate the anatomical localization of mu and delta opioid receptors, as well as, neurotensin receptors. Mu opioid receptors were labeled with {sup 125}I-DAGO. Delta receptors were labeled with {sup 125}I-DPDPE. Neurotensin receptors were labeled with {sup 125}I-NT3. Unilateral lesions of the dopamine perikarya were produced by injections of 6-OHDA administered in the ventral mesencephalon. Unilateral lesions of intrinsic perikarya were induced by injections of quinolinic acid in to the A10 dopamine cell region. Unilateral lesions produced with 6-OHDA resulted in the loss of neurotensin receptors in the A10 region and within the terminal fields. Mu opioid receptors were unaffected by this treatment, but delta opioid receptors increased in the contralateral striatum and nucleus accumbens following 6-OHDA administration. Quinolinic acid produced a reduction of mu opioid receptors within the A10 region with a concomitant reduction in neurotensin receptors in both the cell body region and terminal fields. These results are consistent with a variety of biochemical and behavioral data which suggest the indirect modulation of dopamine transmission by the opioids. In contrast these results strongly indicate a direct modulation of the mesolimbic dopamine system by neurotensin.

Lectin-like oxidized low-density lipoprotein receptor-1 promotes endothelial dysfunction in LDL receptor knockout background.

Science.gov (United States)

Hofmann, Anja; Brunssen, Coy; Poitz, David M; Langbein, Heike; Strasser, Ruth H; Henle, Thomas; Ravens, Ursula; Morawietz, Henning

2017-11-01

Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is the major receptor for oxidized LDL in endothelial cells. LOX-1 is highly expressed in atherosclerotic plaques. The impact of LOX-1 on development of endothelial dysfunction in large vessels in absence or presence of atherosclerosis-prone conditions has not been studied to date. Mice with endothelial cell-specific LOX-1 overexpression (bLOX-1tg) were analyzed. Wild-type (WT) mice served as controls. In addition, bLOX-1tg mice were crossed with LDL receptor knockout (Ldlr -/- ) mice. All mice were fed a western-type diet (WD) or control diet (CD) for 20 weeks. Afterwards, endothelial function was analyzed ex vivo in thoracic aortas using a Mulvany myograph. WD induced hypertriglyceridemia (bLOX-1tg: 1.6-fold; WT: 1.4-fold) and hypercholesterolemia (P LDL-cholesterol (∼9-fold) compared to WT and bLOX-1tg mice on WD. Endothelial function in response to WD was impaired in bLOX-1tg/Ldlr -/- mice (Eff max : 56.7 ± 23.0%) compared to WT (Eff max : 88.2 ± 15.8%, P < 0.001), bLOX-1tg (Eff max : 76.7 ± 12.9%, P < 0.05) and Ldlr -/- mice (Eff max : 70.1 ± 13.1%, P < 0.05). No differences between WT, bLOX-1tg and Ldlr -/- mice were detectable when comparing all genotypes. Endothelial LOX-1 overexpression in an atherosclerosis-prone background impairs endothelial function, proving its importance in the development of atherosclerosis. Copyright © 2017 Elsevier B.V. All rights reserved.