Identification of vasopressin‐responsive miRNAs and AQP2‐targeting miRNAs in kidney collecting duct cells (1137.6)

Abstract

MicroRNA (miRNA), a small RNA synthesized from non‐coding RNA, acts as an important post‐transcriptional regulator. We aimed to identify the vasopressin‐responsive miRNAs from kidney inner medullary collecting duct (IMCD) cells, and particularly to demonstrate aquaporin‐2 (AQP2)‐targeting miRNAs. Microarray chip assay (Affymetrix GeneChip miRNA 3.0 Array) was carried out in IMCD tubule suspension of rat kidney in the absence or the presence of dDAVP stimulation (10‐8 M, 2 h). The results demonstrated 19 miRNAs, including both precursors and mature miRNAs, which showed 1.3‐fold changes in the expression in response to dDAVP stimulation (P<0.05). Additionally, vasopressin‐responsive miRNA networks were predicted by in silico analysis, which exhibited potential target genes. To identify AQP2‐targeting miRNAs, in silico analysis using TargetScan 6.2 was performed. Four miRNAs (miR‐32, miR‐137, miR‐216a, and miR‐216b) targeting 3’UTR of rat AQP2 mRNA were predicted. Target seed regions of miR‐32 and miR‐137 were conserved at the 3’UTR (476 ‐ 483) of rat AQP2 mRNA. RT‐qPCR and immunoblot analysis demonstrated that dDAVP‐induced AQP2 up‐regulation was significantly attenuated in mpkCCDc14 cells, when the cells were transfected by miRNA‐mimic of miR‐32 or miR‐137. This study provides a novel insight on the regulation of AQP2 protein expression via RNA interference, i.e., AQP2‐targeting miRNAs.