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MicroRNAs regulate renal tubule maturation through modulation of Pkd1.

ABSTRACT: MicroRNAs (miRNAs) contribute to the regulation of early kidney development, but their role during later stages of renal tubule maturation is not well understood. Here, we found that ablation of the miRNA-processing enzyme Dicer from maturing renal tubules produces tubular and glomerular cysts in mice. Inactivation of Dicer is associated with downregulation of miR-200, a kidney-enriched miRNA family, and upregulation of the polycystic kidney disease gene Pkd1. Inhibition of miR-200 in cultured renal epithelial cells disrupted tubulogenesis and led to upregulation of Pkd1. Using bioinformatic and in vitro approaches, we found that miR-200b/c/429 induce post-transcriptional repression of Pkd1 through two conserved binding sites in the 3'-Untranslated regions of Pkd1. Overexpression of PKD1 in renal epithelial cells was sufficient to disrupt tubulogenesis and produce cyst-like structures. In conclusion, miRNAs are essential for the maturation of renal tubules, and Pkd1 is a target of miR-200. These results also suggest that miRNAs may modulate PKD1 gene dosage and play a role in the initiation of cystogenesis.

SUBMITTER: Patel V

PROVIDER: S-EPMC3507362 | biostudies-literature | 2012 Dec

REPOSITORIES: biostudies-literature

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MicroRNAs regulate renal tubule maturation through modulation of Pkd1.

Patel Vishal V Hajarnis Sachin S Williams Darren D Hunter Ryan R Huynh Donovan D Igarashi Peter P

Journal of the American Society of Nephrology : JASN 20121108 12

MicroRNAs (miRNAs) contribute to the regulation of early kidney development, but their role during later stages of renal tubule maturation is not well understood. Here, we found that ablation of the miRNA-processing enzyme Dicer from maturing renal tubules produces tubular and glomerular cysts in mice. Inactivation of Dicer is associated with downregulation of miR-200, a kidney-enriched miRNA family, and upregulation of the polycystic kidney disease gene Pkd1. Inhibition of miR-200 in cultured r ...[more]

PMID: 23138483

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Project description:Crosstalk of renal epithelial cells with interstitial fibroblasts plays an important role in kidney pathophysiology. A previous study showed that crosstalk between renal epithelial cells and renal fibroblasts protects against acidosis-induced damage. In order to gain further mechanistic insight into this crosstalk, we investigated the effect of acidosis on the transcriptome of renal epithelial cells (NRK-52E) and renal fibroblasts (NRK-49F) in co-culture by RNASeq, bioinformatics analysis and experimental validation. Cells were exposed to acidic media or control media for 48 h. RNA and protein from whole cell lysate were isolated. In addition, cells were fractionated into cytosol, nucleus and chromatin. RNASeq data were analyzed for differential expression and pathway enrichment (ingenuity pathway analysis, IPA, QIAGEN). Total and phosphorylated protein expression was assessed by Western blot (WB). Transcription factor activity was assessed by luciferase reporter assay. Bioinformatic analysis using differentially expressed genes according to RNASeq (7834 for NRK-52E and 3197 for NRK-49F) predicted the antioxidant and cell-protective Nrf2 pathway as acidosis-induced in NRK-52E and NRK-49F cells. Activation of Nrf2 comprises enhanced Nrf2 phosphorylation, nuclear translocation, DNA binding and initiation of a cell protective transcriptional program. Our data show that acidosis enhances chromatin-associated Nrf2 expression and the abundance of phosphorylated Nrf2 in the chromatin fraction of NRK-52E cells in co-culture but not in monoculture. Furthermore, acidosis enhances the activity of a reporter for Nrf2 (ARE-luciferase). Despite the bioinformatics prediction, NRK-49F cells did not respond with Nrf2 activation. Transketolase (TKT) is an important regulator of antioxidant and homeostatic responses in the kidney and a canonical Nrf2 target gene. We show that protein and mRNA expression of TKT is increased in NRK-52E cells under co-culture but not under monoculture conditions. In conclusion, our data show that extracellular acidosis activates the cytoprotective transcription factor Nrf2 in renal epithelial cells co-cultivated with renal fibroblasts, thereby enhancing the expression of cytoprotective TKT. This protective response is not observed in monoculture. Activation of the Nrf2 pathway represents a co-operative cellular strategy of protection against acidosis.

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MicroRNAs regulate renal tubule maturation through modulation of Pkd1.

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MicroRNAs regulate renal tubule maturation through modulation of Pkd1.

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