Effects of microRNA-1 on negatively regulating L-type calcium channel beta2 subunit gene expression during cardiac hypertrophy.
- Author:
Yang WU
1
;
Peng GENG
;
Yu-Qin WANG
;
Yan LIU
Author Information
- Publication Type:Journal Article
- MeSH: Animals; Atrial Natriuretic Factor; metabolism; Calcium Channels, L-Type; genetics; Cardiomegaly; genetics; Gene Expression Regulation; HEK293 Cells; Humans; MicroRNAs; genetics; Rats; Rats, Sprague-Dawley; Transfection; Ventricular Myosins; metabolism
- From: Chinese Journal of Applied Physiology 2012;28(4):304-308
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo investigate the negative regulation of microRNA-1 (miR-1) on L-type calcium channel beta2 subunit (Cavbeta 2) during cardiomyocyte hypertrophy and its mechanism.
METHODSCardiomyocyte hypertrophy was induced by isoproterenol (ISO). The cell surface area was measured by image analysis system (HJ2000). The targets of miR-1 were predicted by online database microCosm. The 3' untranslated region sequence of Cavbeta 2 was cloned into luciferase reporter vector and then transiently transfected into HEK293 cells. The luciferase activities of samples were measured to verify the expression of luciferase reporter vector. The expression of atrial natriuretic peptide (ANP), beta-myosin heavy chain (beta-MHC), miR-1 and the Cavbeta 2 mRNA were detected by qRT-PCR. The protein expression of Cavbeta 2 was detected by Western blot. The level of miR-1 was up-regulated by miR-1 mimic transfection and the expression level of Cavbeta 2 was down-regulated by RNAi, then effects of which on cardiomyocyte hypertrophy were investigated.
RESULTS(1) The expression of miR-1 was significantly reduced in cardiomyocyte hypertrophy. Upregulating the miR-1 level could suppress the increase of cell surface area, the expression of ANP and beta-MHC mRNA (P < 0.05). (2) Cavbeta 2 was the one of potential targets of miR-1 by prediction using online database microCosm. The luciferase activities of HEK293 cells with the plasmid containing miR-1 and wide type Cavbeta 3' UTR sequence was significantly decreased when compared with that of control group (P < 0.01). Up-regulation of the miR-1 level could suppress the protein expression of Cavbeta 2. (3) The expression of Cavbeta 2 was significantly increased in cardiomyocyte hypertrophy induced by ISO. Downregulation of Cavbeta by RNAi could markedly inhibit the increase of cell surface area, the expression of ANP and beta-MHC mRNA.
CONCLUSIONCavbeta2 is one of potential targets of miR-1 by bioinformatics prediction. The experiment data confirms that Cavbeta2 is truly the target of miR-1. MiR-1 can negatively regulate the expression of Cavbeta 2, resulting in the decrease of intracellular Ca2+ content and the attenuation of cardiomyocyte hypertrophy.
