Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: UNIPROT:P01185 (vasopressin)
23,126 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Neuronal expression of vasopressin messenger RNA (mRNA) and peptide has been shown to be estrogen dependent. A 5.5-kb genomic DNA fragment, 5' of the AVP coding region, was used in luciferase reporter assays to measure transcriptional activation by either estrogen receptor alpha or beta in response to various treatments. ER alpha and ER beta displayed differential regulation of the AVP promoter. SK-N-SH cells transfected with ER alpha exhibited increased luciferase activity in response to estrogen, and the selective estrogen receptor modulators (SERMs), Tamoxifen, and ICI 182,780. Cells transfected with ER beta exhibited a high constitutive activity, which is unchanged by exposure to SERMs but can be inhibited by estrogen. Deletion of 1.5 kb from the 5' end or mutation of a single estrogen response element (ERE)-like sequence resulted in loss of estrogen-dependent induction by ER alpha and increased the ability of estrogen to inhibit the high constitutive activity of ER beta. The distal ERE-containing 1.5-kb fragment, when coupled to luciferase, is able to support both ER alpha and ER beta mediated activation of transcription by estrogen. These results suggest that a single ERE in the distal 1.5-kb portion of the 5.5-kb fragment contains the primary positive estrogen responsive sequences for ER alpha and ER beta. The data also suggest that sequences proximal to this element serve to inhibit transcription mediated by ER beta.
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PMID:Differential transcriptional regulation of rat vasopressin gene expression by estrogen receptor alpha and beta. 1108 36

1. The major side effects of the immunosuppressive drug cyclosporin A (CsA) are hypertension and nephrotoxicity. It is likely that both are caused by local vasoconstriction. 2. We have shown previously that 20 h treatment of rat vascular smooth muscle cells (VSMC) with therapeutically relevant CsA concentrations increased the cellular response to [Arg8]vasopressin (AVP) by increasing about 2 fold the number of vasopressin receptors. 3. Displacement experiments using a specific antagonist of the vasopressin V1A receptor (V1AR) showed that the vasopressin binding sites present in VSMC were exclusively receptors of the V1A subtype. 4. Receptor internalization studies revealed that CsA (10(-6) M) did not significantly alter AVP receptor trafficking. 5. V1AR mRNA was increased by CsA, as measured by quantitative polymerase chain reaction. Time-course studies indicated that the increase in mRNA preceded cell surface expression of the receptor, as measured by hormone binding. 6. A direct effect of CsA on the V1AR promoter was investigated using VSMC transfected with a V1AR promoter-luciferase reporter construct. Surprisingly, CsA did not increase, but rather slightly reduced V1AR promoter activity. This effect was independent of the cyclophilin-calcineurin pathway. 7. Measurement of V1AR mRNA decay in the presence of the transcription inhibitor actinomycin D revealed that CsA increased the half-life of V1AR mRNA about 2 fold. 8. In conclusion, CsA increased the response of VSMC to AVP by upregulating V1AR expression through stabilization of its mRNA. This could be a key mechanism in enhanced vascular responsiveness induced by CsA, causing both hypertension and, via renal vasoconstriction, reduced glomerular filtration.
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PMID:Upregulation of vasopressin V1A receptor mRNA and protein in vascular smooth muscle cells following cyclosporin A treatment. 1118 32

We cloned the Slc14a2 gene and determined the genomic organization of the rat urea transporter UT-A. Slc14a2, the gene encoding the rat UT-A transporter, extends for more that 300 kb. The four known rat mRNA isoforms: UT-A1, UT-A2, UT-A3, and UT-A4 are transcribed from 24 exons. The Slc14a2 genomic map also accounts for 3'-untranslated sequences expressed alternatively in UT-A1, UT-A2, and UT-A3. We previously identified a TATA-less, tonicity-responsive promoter controlling the transcription of UT-A1, UT-A3, and UT-A4 from a single initiation site in the 5'-flanking region of the gene. Here, we describe a second, internal promoter in intron 12, which controls the transcription of UT-A2 starting from exon 13. This region contains a TATA motif upstream from the UT-A2 transcription start site, and shows consensus sequences for the cAMP response element (CRE) and for the tonicity enhancer (TonE) motif. Stimulation by cAMP induces UT-A2 mRNA expression in mIMCD3 cells, and luciferase activity in mIMCD3 cells transfected with those pGL3 constructs including the CRE sequences. Although long-term exposure to hypertonicity induces UT-A2 expression in mIMCD3 cells, hypertonicity does not induce significantly the activity of the promoter in intron 12. In summary, we describe the genomic structure of the rat UT-A urea transporter, encoded by the Slc14a2 gene. Our findings suggest that two promoters regulate transcription of the four UT-A isoforms, and that stimulation of transcription by vasopressin, mediated by cAMP and CRE sequences, and controlled by an intronic promoter, may contribute to the increase in UT-A2 expression during water deprivation.
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PMID:Cloning of the rat Slc14a2 gene and genomic organization of the UT-A urea transporter. 1126 55

The V(1b) vasopressin receptor, expressed mainly in the corticotroph of the anterior pituitary, mediates the stimulatory effect of vasopressin on ACTH release. To clarify the regulation of receptor expression, we cloned, sequenced (up to approximately 5 kb from the translation start site), and characterized the 5'-flanking region of the rat V(1b) receptor gene. We identified the transcription start site by amplification of cDNA ends and found a new intron within the 5'-untranslated region (5'-UTR) by comparing the sequence with that of cDNA. We then confirmed that the obtained promoter indeed has transcriptional activity by use of the luciferase reporter in AtT-20 mouse corticotroph cells. Interestingly, there were five short upstream open reading frames (uORFs) located within the 5'-UTR that were found to suppress V(1b) expression. Subsequent mutational analyses showed that the two downstream uORFs have an inhibitory effect on expression in both homologous and heterologous contexts. Furthermore, the inhibition did not accompany a parallel decrease in mRNA, suggesting that the suppressive effect occurs at a level downstream of transcription. Taken together, our data strongly suggest that the expression of the V(1b) receptor is regulated at the posttranscriptional as well as transcriptional level through uORFs within the 5'-UTR region of the mRNA. Whether the uORF-mediated regulation of V(1b) expression is functionally linked to any intracellular and/or extracellular factor(s) awaits further research.
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PMID:Involvement of upstream open reading frames in regulation of rat V(1b) vasopressin receptor expression. 1128 61

The role of CT repeats (inverted GAGA box) in the rat vasopressin V1b receptor (V1bR) promoter in the transcriptional regulation of this gene was studied in H32 hypothalamic cells, which express endogenous V1bR. Transfection of a 2.5-kb V1bR fragment (2161 bp upstream and 377 bp downstream of the proximal transcriptional start point) into a luciferase vector (V1bRp2.5-Luc) results in promoter activity in these cells. The 670-bp proximal promoter fragment containing the GAGA box showed maximal promoter activity, whereas deletion of the GAGA box abolished transcription. Drosophila GAGA-binding protein increased V1bR promoter activity by 11-fold when cotransfected with V1bRp2.5-Luc and increased endogenous V1bR expression. Electrophoretic mobility shift assay showed specific binding of pituitary nuclear extracts to radiolabeled GAGA oligonucleotides, which increased following restraint stress in rats, a condition associated with V1bR up-regulation. DNA-binding activity involved a protein complex because it was abolished by deoxycholate. Size-exclusion column chromatography showed a complex of 127 kDa, which dissociated into approximately 70-kDa components after deoxycholate/Nonidet P-40 treatment. This study demonstrates that interactions of GAGA-binding proteins with the GAGA box of the V1bR promoter activate V1bR gene expression and provides a potential mechanism for physiological regulation of V1bR transcription.
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PMID:Transcriptional regulation of the pituitary vasopressin V1b receptor involves a GAGA-binding protein. 1202 77

The BMAL2 gene encodes a member of the basic helix-loop-helix PER-ARNT-SIM family of transcription factors, which control diverse physiological processes including circadian rhythms. We identified four novel human BMAL2 transcripts that differ by alternative splicing within their NH2-terminal regions. Divergent expression of these and previously reported transcripts was observed among human tissues. The functional consequences of alternative splicing for transcriptional activation by CLOCK:BMAL2 heterodimers were assessed using luciferase reporter gene constructs that contained one of three diurnally regulated promoters, namely, those of the mouse period1, mouse vasopressin, and human plasminogen activator inhibitor-1 genes. These studies revealed that alternative splicing generates BMAL2 isoforms possessing high, medium, low, or no transcriptional activity. Similar results were obtained with each promoter, suggesting that alternative splicing may influence the amplitudes of both central and peripheral oscillators. Indeed, alternative splicing of BMAL2 may provide tissues with a rheostat capable of regulating CLOCK:BMAL2 heterodimer function across a broad continuum of potential transcriptional activities to accommodate varied metabolic demands and physiological roles.
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PMID:Alternative splicing yields novel BMAL2 variants: tissue distribution and functional characterization. 1205 78

By binding to agonist-activated G protein-coupled receptors (GPCRs), beta-arrestins mediate homologous receptor desensitization and endocytosis via clathrin-coated pits. Recent data suggest that beta-arrestins also contribute to GPCR signaling by acting as scaffolds for components of the ERK mitogen-activated protein kinase cascade. Because of these dual functions, we hypothesized that the stability of the receptor-beta-arrestin interaction might affect the mechanism and functional consequences of GPCR-stimulated ERK activation. In transfected COS-7 cells, we found that angiotensin AT1a and vasopressin V2 receptors, which form stable receptor-beta-arrestin complexes, activated a beta-arrestin-bound pool of ERK2 more efficiently than alpha 1b and beta2 adrenergic receptors, which form transient receptor-beta-arrestin complexes. We next studied chimeric receptors in which the pattern of beta-arrestin binding was reversed by exchanging the C-terminal tails of the beta2 and V2 receptors. The ability of the V2 beta 2 and beta 2V2 chimeras to activate beta-arrestin-bound ERK2 corresponded to the pattern of beta-arrestin binding, suggesting that the stability of the receptor-beta-arrestin complex determined the mechanism of ERK2 activation. Analysis of covalently cross-linked detergent lysates and cellular fractionation revealed that wild type V2 receptors generated a larger pool of cytosolic phospho-ERK1/2 and less nuclear phospho-ERK1/2 than the chimeric V2 beta 2 receptor, consistent with the cytosolic retention of beta-arrestin-bound ERK. In stably transfected HEK-293 cells, the V2 beta 2 receptor increased ERK1/2-mediated, Elk-1-driven transcription of a luciferase reporter to a greater extent than the wild type V2 receptor. Furthermore, the V2 beta 2, but not the V2 receptor, was capable of eliciting a mitogenic response. These data suggest that the C-terminal tail of a GPCR, by determining the stability of the receptor-beta-arrestin complex, controls the extent of beta-arrestin-bound ERK activation, and influences both the subcellular localization of activated ERK and the physiologic consequences of ERK activation.
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PMID:The stability of the G protein-coupled receptor-beta-arrestin interaction determines the mechanism and functional consequence of ERK activation. 1247 60

In BRET2 (Bioluminescence Resonance Energy Transfer), a Renilla luciferase (RLuc) is used as the donor protein, while a Green Fluorescent Protein (GFP2) is used as the acceptor protein. In the presence of the cell permeable substrate DeepBlueC, RLuc emits blue light at 395 nm. If the GFP2 is brought into close proximity to RLuc via a specific biomolecular interaction, the GFP2 will absorb the blue light energy and reemit green light at 510nm. BRET2 signals are therefore easily determined by measuring the ratio of green over blue light (510/395nm) using appropriate dual channel luminometry instruments (e.g., Fusion Universal Microplate Analyzer, Packard BioScience). Since no light source is required for BRET2 assays, the technology does not suffer from high fluorescent background or photobleaching, the common problems associated with standard FRET-based assays. Using BRET2, we developed a generic G Protein-Coupled Receptor (GPCR) assay based on the observation that activation of the majority of GPCRs by agonists leads to the interaction of beta-arrestin (a protein that is involved in receptor desensitization and sequestration) with the receptor. We established a cell line stably expressing the GFP2:beta-arrestin 2 fusion protein, and showed that it can be used to monitor the activation of various transiently expressed GPCRs, in BRET2/arrestin assays. In addition, using the HEK 293/GFP2:beta-arrestin 2 cell line as a recipient, we generated a double-stable line co-expressing the vasopressin 2 receptor (V2R) fused to RLuc (V2R:RLuc) and used it for the pharmacological characterization of compounds in BRET2/arrestin assays. This approach yields genuine pharmacology and supports the BRET2/arrestin assay as a tool that can be used with recombinant cell lines to characterize ligand-GPCR interactions which can be applied to ligand identification for orphan receptors.
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PMID:The BRET2/arrestin assay in stable recombinant cells: a platform to screen for compounds that interact with G protein-coupled receptors (GPCRS). 1250 39

The regulation of CRH promoter activity by cAMP was studied in two cell lines, the pituitary corticotroph cell line AtT-20 and the immortalized hypothalamic cell line 4B, which expresses CRH and vasopressin. In 4B cells transfected with a CRH promoter-luciferase construct, the adenylyl cyclase stimulator, forskolin, increased luciferase activity in parallel with increases in intracellular cAMP. In 4B cells, however, the phosphodiesterase inhibitor, isobutylmethylxanthine, potentiated forskolin-stimulated cAMP without affecting further increases in luciferase activity. In AtT-20 cells, forskolin plus isobutylmethylxanthine elevated cAMP only slightly, but increased luciferase activity to levels similar to those observed in 4B cells. AtT-20 cells were also unresponsive to 8-bromo-cAMP, due in part to higher phosphodiesterase (PDE) activities. Although both cells contained PDE1, -3, and -4, inhibition of either PDE4 or PDE1 potentiated luciferase activity stimulated by submaximal forskolin concentrations in 4B cells, while only simultaneous inhibition of PDE3 and PDE4 was effective in AtT-20 cells. The data show that minor elevations in intracellular cAMP are sufficient for full stimulation of CRH promoter activity regardless of the cell line. Furthermore, poor CRH promoter activation in AtT-20 cells appears to result from deficient cAMP production and rapid cAMP degradation by PDE.
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PMID:Cyclic adenosine 3',5'-monophosphate regulation of corticotropin-releasing hormone promoter activity in AtT-20 cells and in a transformed hypothalamic cell line. 1263 12

The vasopressin gene is expressed in the suprachiasmatic nucleus where the basic helix-loop-helix (bHLH)-PAS factors CLOCK and MOP3 regulate circadian expression through interactions with E-box sequences. We have examined vasopressin gene regulation by HIF-1alpha, a bHLH-PAS factor involved in responses to hypoxia. By transfecting Neuro-2A cells with 5' flanking regions of vasopressin gene driving a luciferase reporter, we have shown that CLOCK and HIF-1alpha cooperate in the induction of expression from 1000 bp and 350 bp of the vasopressin promoter but do not activate a 120-bp promoter fragment. The region between -191 and -128 contains an E-box A that appears to be essential for HIF-1alpha/CLOCK-mediated transcriptional activity. However, gel-shift analysis shows that the cooperative effect of HIF-1alpha and CLOCK results in MOP3 binding, but does not involve heterodimerization of HIF-1alpha/CLOCK, at E-box A. These data indicate that cross-talk between mediators of hypoxic and circadian pathways can regulate target genes.
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PMID:Cross-talk between hypoxic and circadian pathways: cooperative roles for hypoxia-inducible factor 1alpha and CLOCK in transcriptional activation of the vasopressin gene. 1269 40


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