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ChIP-Seq analysis of transcription factors CREB and C/EBP beta in vasopressin-responsive mouse renal collecting duct mpkCCD cells

ABSTRACT: 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 a cyclic AMP/protein kinase A (PKA) signaling pathway. Sequentially, after activation of cAMP/PKA signaling, many of transcription factors involve gene transcription process. cAMP-response element binding protein (CREB) and cAMP-responsive transcription factor C/EBP beta are potential candidates for vaopressin-mediated regulation of Aqp2 gene transcription proviously reported. In the present study, genome-wide binding sites for two b-ZIP transcription factors CREB and C/EBP beta were identified in vasopressin-responsive mouse collecting duct mpkCCD cells using ChIP-Seq.

ORGANISM(S): Mus musculus

PROVIDER: GSE98076 | GEO | 2018/03/26

REPOSITORIES: GEO

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ChIP-Seq analysis of Histone H3 acetylation (K27) changes in response to vasopressin in mouse renal collecting duct mpkCCD cells

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2018-01-02 | GSE95007 | GEO

Mass spectrometry-based analysis of LRBA phosphorylation

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2022-07-23 | PXD033859 | Pride

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2022-07-23 | PXD033852 | Pride

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2018-03-26 | GSE108786 | GEO

Identification of PKA-dependent signaling network using CRISPR-Cas9 coupled with quantitative transcriptomics, proteomics and phosphoproteomics

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2018-01-02 | GSE95008 | GEO

Glucocorticoid receptor maintains vasopressin responses in kidney collecting duct cells

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

CRISPR-Cas9/phosphoproteomics identifies Myosin Light Chain Kinase dependent signaling in kidney epithelial cells

Project description:Prior studies have implicated myosin light chain kinase (MLCK) in the regulation of aqua-porin-2 (AQP2) in the renal collecting duct. To discover signaling targets of MLCK, we deleted the Mylk- gene to obtain MLCK-null mpkCCD cells and carried out comprehensive phosphopro-teomics using the SILAC method for quantification. Only 107 phosphopeptides were changed in abundance by MLCK deletion out of 1743 quantified over multiple replicates (29 decreased and 78 increased). One of the decreased phosphopeptides corresponded to the canonical target site in myosin regulatory light chain (MLC). Network analysis indicated that targeted phosphopro-teins clustered into distinct structural/functional groups: “acto-myosin,” “signaling,” “nuclear envelope,” “gene transcription,” “mRNA processing,” “energy metabolism,” “intermediate fila-ments,” “adherens junctions” and “tight junctions.” There was significant overlap between the derived MLCK signaling network and a previously determined PKA signaling network. Phalloidin labeling revealed that MLCK deletion inhibited the normal effect of vasopressin to depolymer-ize F-actin. Immunocytochemistry and electron microscopy demonstrated a defect in pro-cessing of early endosomes to late endosomes. In contrast, surface biotinylation studies showed no effect of MLCK deletion on AQP2 endocytosis. We conclude that MLCK is part of a multi-component signaling pathway that includes much more than simple regulation of conven-tional non-muscle myosins through MLC phosphorylation. Furthermore, vasopressin signaling in collecting duct cells involves both MLCK-dependent signaling and PKA-dependent signaling, which display significant overlap. Mapping MLCK targets revealed potential roles in both nu-cleus and cytoplasm. The latter includes proteins that regulate both actin polymerization and AQP2-endosomal processing from early to late endosomes.

2020-03-04 | PXD014985 | Pride

Expression data from mouse collecting duct cell, mpkCCD, in response to a peptide hormone vasopressin analog, dDAVP

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

Transcriptomic Changes in Microdissected CCDs after Three Hours Lipopolysaccharide (LPS) Administration in Rats

Project description:Ureteral obstruction is marked by disappearance of the vasopressin-dependent water channel aquaporin-2 (AQP2) in the renal collecting duct and polyuria upon reversal. Most studies of unilateral ureteral obstruction (UUO) models have examined late time points, obscuring the early signals that trigger loss of AQP2. Here, we performed RNA-Seq on microdissected rat cortical collecting ducts (CCDs) to identify early signaling pathways post-UUO. To address whether the induction of inflammatory signaling is a cause of loss of AQP2 expression, we directly induced inflammatory signaling in rats with lipopolysaccharide (LPS) administration

2022-07-20 | GSE200928 | GEO

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