Collecting Duct Gene Expression in a Mouse Vasopressin-Excess Model without Hyponatremia
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PROVIDER: PRJNA1284937 | ENA |
REPOSITORIES: ENA
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Project description:Introduction. States of vasopressin excess can exist either with or without hyponatremia, depending on water intake. Here, we set up a mouse model of vasopressin-excess without hyponatremia to address the consequences of vasopressin excess that are independent of serum sodium. Methods. A mouse vasopressin-excess model was created by infusing desmopressin for 5 days without a forced water load. Control mice were infused with vehicle. Results. RNA-seq in microdissected cortical collecting ducts (CCDs) from vasopressin-excess mice showed significant changes in 153 transcripts including the expected increases in aquaporin-2 (AQP2) and AQP3 along with increases in several other targets of V2-receptor-mediated signaling in principal cells. In addition, there were decreases in abundances of many transcripts known to be targets of regulation by the Wnt/frizzled/β-catenin pathway. The vasopressin-excess model also manifested marked increases in phosphorylation of β-catenin at a known protein kinase A (PKA) site, viz. Ser552. CRISPR-mediated mutation of this PKA-target site in mice resulted in extensive changes in the transcriptomes of microdissected CCDs after desmopressin-infusion, heavily weighted toward increases in known transcriptional targets of the Wnt/β-catenin pathway. ATAC-seq in microdissected CCDs from DDAVP-infused mice revealed decreases in chromatin accessibility at promoters of many genes regulated in the Wnt/β-catenin pathway. Conclusion. We conclude that vasopressin-excess is associated with vasopressin/PKA-dependent repression of Wnt/Frizzled/β-catenin target gene expression, mediated in part by PKA-dependent phosphorylation of Ser552 of β-catenin. The repression is associated with decreasing chromatin accessibility in the promoter-TSS regions of target genes. The findings suggest that normonatremic vasopressin-excess is not necessarily benign.
2025-07-03 | GSE301404 | GEO
Project description:Water permeability of the kidney collecting ducts is regulated in part by the amount of the molecular water channel protein aquaporin-2 (AQP2), whose expression, in turn, is regulated by the pituitary peptide hormone vasopressin. We previously showed that stable glucocorticoid receptor knockdown diminished the vasopressin-induced Aqp2 gene expression in the collecting duct cell model mpkCCD. Here, we investigated the pathways regulated by the glucocorticoid receptor by comparing transcriptomes of the mpkCCD cells with or without stable glucocorticoid receptor knockdown. Glucocorticoid receptor knockdown downregulated 5,394 transcripts associated with 55 KEGG pathways including “vasopressin-regulated water reabsorption,” indicative of positive regulatory roles of these pathways in the vasopressin-induced Aqp2 gene expression. Quantitative RT-PCR confirmed the downregulation of the vasopressin V2 receptor transcript upon glucocorticoid receptor knockdown. Glucocorticoid receptor knockdown upregulated 3,785 transcripts associated with 42 KEGG pathways including the “TNF signaling pathway” and “TGFβ signaling pathway,” suggesting the negative regulatory roles of these pathways in the vasopressin-induced Aqp2 gene expression. Quantitative RT-PCR confirmed the upregulation of TNF and TGFβ receptor transcripts upon glucocorticoid receptor knockdown. TNF or TGFβ inhibitor alone, in the absence of vasopressin, did not induce Aqp2 gene transcription. However, TNF or TGFβ blunted the vasopressin-induced Aqp2 gene expression. In particular, TGFβ reduced vasopressin-induced increases in Akt phosphorylation without inducing epithelial-to-mesenchymal transition or interfering with vasopressin-induced apical AQP2 trafficking. In summary, our RNA-seq transcriptomic comparison revealed positive and negative regulatory pathways maintained by the glucocorticoid receptor for the vasopressin-induced Aqp2 gene expression.
2022-05-16 | GSE194044 | GEO
Project description:Purpose: The goal of this study is to identify vasopressin-regulated genes in mouse kidney collecting duct cell line mpkCCD. To explore dynamic regulation of vasoressin-mediated transcriptional regulation, transcriptome of vasopressin-treated cells at four different time points (3h, 6h, 12h, and 24h) were profiled and analyzed. Methods: Total RNAs were isolated from vasopressin-treated mpkCCD cells at different time points (3h, 6h, 12h, and 24h). Three replicates for vehicle- or vasopressin-treated group were generated at each tested time point. cDNA libraries were prepared using a Nextera DNA library preparation protocol. The sequence reads from Illumina HiSeq3000 platform were qualified and quantified at the transcript level using Salmon (0.14.1). Differential expression analysis were performed using edgeR. Results: mRNA profiles of mouse kidney collecting duct mpkCCD cells treated with vasopressin analog (dDAVP) for four different time potins (3h, 6h, 12h, and 24h) were generated using an optimized RNA-Seq workflow. Transcript level quantification using a pseudo-alignment quantification method (Salmon) was performed to calculate transcript abundance in each sample. Then differential expression analysis identified the differentially expressed genes including Aqp2 gene at each time point comparison (dDAVP vs vehicle, FDR < 0.05). In addition, several new vasopressin-responsive genes that have not been elucidated before were identified. Conclusions: This study revealed dynamic changes of vasopressin-responsive gene expression within 24 hours in mouse kidney collecting duct cells. The results from time-course transcriptome profiling identified the known vasopressin-responsive genes and several novel gene that are regulated temporally.
2021-07-22 | GSE163566 | GEO
Project description:Vasopressin is the major hormone that regulates renal water excretion. It does so by binding to a receptor in renal collecting duct cells, triggering signaling pathways that ultimately regulate the abundance, location, and activity of the water channel protein aquaporin 2. We took an advantage of quantitative large scale proteomic technologies and oligonucleotide microarrays to quantify steady state changes in protein and transcript abundances in response to vasopressin in a collecting duct cell line, mpkCCD clone 11 (Yu et al. PNAS 2009, 106:2441-2446). This cell line originally developed by Alan Vandewalle’s group recapitulates vasopressin-mediated AQP2 expression and phosphorylation as seen in native colleting duct cells.
2010-12-09 | GSE19938 | GEO
Project description:We sequenced mRNAs and mapping the binding of RNA polymerase 2 in collecting duct cells treat with Vasopressin or vehicle.
2016-07-01 | GSE79525 | GEO
Project description:Vasopressin is the major hormone that regulates renal water excretion. It does so by binding to a receptor in renal collecting duct cells, triggering signaling pathways that ultimately regulate the abundance, location, and activity of the water channel protein aquaporin 2. We took an advantage of quantitative large scale proteomic technologies and oligonucleotide microarrays to quantify steady state changes in protein and transcript abundances in response to vasopressin in a collecting duct cell line, mpkCCD clone 11 (Yu et al. PNAS 2009, 106:2441-2446). This cell line originally developed by Alan Vandewalle’s group recapitulates vasopressin-mediated AQP2 expression and phosphorylation as seen in native colleting duct cells. The mpkCCD cells were grown on membrane supports to permit polarization. Once transepithelial resistance reached 5kohm per centimeter square and higher, the cells were exposed to the vasopressin V2 receptor analog, dDAVP, at a physiological concentration, 0.1nM, for 5 days. Control experiments were done with cells exposed to vehicle alone. Total RNA was harvested and processed for transcript expression analysis using Affymetrix GeneChip Mouse Genome 430 2.0 Arrays. Each experimental treatment, vehicle and dDAVP, was repeated 3 times.
2010-12-09 | E-GEOD-19938 | biostudies-arrayexpress
Project description:We sequenced mRNAs and mapping the binding of RNA polymerase 2 in collecting duct cells treat with Vasopressin or vehicle. Confluent mpkCCD cell monolayers were maintained in serum-free medium for 24 h prior to treatment with or without 0.1 nM dDAVP in serum-free medium for 24 h. followed RNA polymerase 2 Chip seq.
2016-07-01 | E-GEOD-79525 | biostudies-arrayexpress
Project description:In mammals, the peptide hormone vasopressin controls renal water excretion, largely through regulation of the molecular water channel aquaporin-2 (AQP2) in the renal collecting duct. Regulatory mechanisms of AQP2 show: 1) short-term regulation by membrane trafficking; and 2) long-term regulation involving vasopressin-induced changes in the abundance of the aquaporin-2 protein. Vasopressin activates a G protein-coupled receptor (V2R) increasing cyclic AMP and activating protein kinase PKA. Crebbp and Ep300 are known targets of PKA. They are histone acetyltransferases that acetylate histone H3 lysine-27, a histone mark associated with open chromatin and increased transcription (Tie F et al. Development 2009). The translocation of CREBBP and Ep300 into the nucleus in response to vasopressin in the collecting duct cells, predicts that vasopressin, working through PKA, may increase histone H3K27 acetylation of some genes. We tested this by performing ChIP-Seq for this modification.
2018-01-02 | GSE95007 | GEO
Project description:Glucocorticoid receptor maintains vasopressin responses in kidney collecting duct cells
| PRJNA798899 | ENA
Project description:Vasopressin, a peptide hormone, controls renal water excretion, largely through regulation of water channel aquaporin-2 (AQP2) in the renal collecting duct. There are two regulatory mechanisms of AQP2: 1) short-term regulation by membrane trafficking of AQP2; and 2) long-term regulation involving vasopressin-induced changes of protein abundance of AQP2 through regulation of gene transcription and protein half-life. Vasopressin binds a G protein-coupled receptor (V2R) activating several downstream signaling pathways. At downstream of V2R activation, many of transcription factors involve gene transcription process associated with status of chromatin structure. ATAC-Seq (Assay for Transpoase-Accessible Chromatin using Sequencing) is a recent technique to study chromatin accessibility (Buenrostro et al. Nat Methods 2013). We carried out ATAC-Seq following standard an ATAC-Seq protocol in mpkCCD cells treated with vehicle or dDAVP for 30 minutes.
2018-03-26 | GSE108786 | GEO