microRNA regulation of endothelin-1 mRNA in renal collecting duct cells

Abstract

Aims Recently, microRNAs (miRNAs) have been implicated in control of Edn1 mRNA in several tissues. Here we examined the role of miRNA action on Edn1 mRNA expression in renal distal collecting duct cells. Main methods A microarray study was conducted to provide a comprehensive assessment of miRNAs present in a murine inner medullary collecting duct (mIMCD-3) cell line. The experiment was designed as a comparison between mIMCD-3 cells grown in the presence and absence of aldosterone. Argonaute (Ago) immunoprecipitation experiments were used to investigate binding of the RNA induced silencing complex (RISC) to Edn1 mRNA. Key findings Thirty-four miRNAs were detected in very high abundance in mIMCD-3 cells, and a large number of others were present at lower levels. The microarray experiments were validated by quantitative PCR analysis of selected miRNAs. The microarray data, in combination with in silico examination of the Edn1 3’ UTR provided a panel of candidate miRNAs that could act upon the Edn1 expression. Edn1 mRNA was co-immunoprecipitated with an Argonaute protein antibody, and this interaction was blocked by anti-miR-709 oligonucleotides. Significance These results define the miRNA landscape of the mIMCD-3 cell line. Moreover, Edn1 was shown to interact with Argonaute protein suggesting that it is a target of the RNA induced silencing complex (RISC).

Gene targets of mouse miR-709: regulation of distinct pools

“…miR-709 is an abundant miRNA expressed in multiple mouse tissues, including brain, thymus, heart, lung, liver, spleen, kidney, adipose tissue, and testes 14 15 16 17 . miR-709 is embedded in intron 8 of the Regulatory Factor X1 ( Rfx1 ) gene, a member of the winged-helix subfamily of helix-turn-helix transcription factors with activation as well as repression activity 18 .…”

MicroRNA-709 Mediates Acute Tubular Injury through Effects on Mitochondrial Function

“…[34][35][36][37] In addition, miR-709 acts on multiple genes, including Egr2, c-Jun, Sox-2, c-Myc, Akt, Ras-GRF1, GSK3b, ET-1, and Pctp, and plays different roles in various organs, such as promoting carcinoma migration and invasiveness, inhibiting adipocyte differentiation, and inducing the macrophage inflammatory response. 21,35,38 Thus far, the knowledge of miR-709 in the kidney is limited to cultured murine inner medullary collecting duct cells, where it might affect endothelin 1 expression, 34 although its role in kidney injury remains unknown. In this study, we took advantage of a mouse cisplatin-induced AKI model and first reported a prominent upregulation of miR-709 primarily in the renal tubular cells after acute injury.…”

SGK1 regulation by miR-466g in cortical collecting duct cells

“…Human embryonic kidney-293 cells (passages [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20] were transfected in 24-well cell culture plates with 37.5 ng of luciferase reporter plasmid, 25 ng ␤-galactosidase control plasmid, and 5-100 ϫ 10 Ϫ9 M of scrambled or precursor miR-466g (Applied Biosystems). Lipofectamine 2000 (Invitrogen, Carlsbad, CA) was used as a transfection reagent in all experiments.…”

“…Aldosterone, by activating mineralocorticoid receptors, may similarly signal through miRNAs to control expression of genes whose products regulate ENaC activity in the CCD. Recent studies have examined aldosterone-induced changes in miRNA expression in the collecting duct; however, these studies focused on changes in miRNA expression during the late phase of aldosterone induction (12,13,20). To date, no studies have identified early-phase, aldosterone-regulated miRNAs in the CCD; nor have any studies manipulated expression of specific miRNAs in CCD cells to examine functional effects of early-phase, aldosterone-regulated miRNAs on Na ϩ transport.…”