P-03/ BO-01 : Post-transcriptional regulation of 3-hydroxy-3-methyl glutaryl-coenzyme A reductase: crucial role of the microRNA miR-27a

posted Jan 26, 2017, 6:40 AM by sourav ghosh
Abrar A. Khan, Vinayak Gupta, Kalyani Ananthamohan, Vikas Arige, Anil Kiran Chokkalla, Nitish R. Mahapatra
Department of Biotechnology, Bhupat and Jyoti Mehta School of Bioscience, Indian Institute of Technology Madras, Chennai , India

Dyslipidemia is a strong predictor of cardiovascular diseases including Essential Hypertension (EH). 3-Hydroxy-3-Methyl Glutaryl-Coenzyme A reductase (Hmgcr), coding for the rate-limiting enzyme in the cholesterol biosynthesis, is an important candidate gene for EH. However, the regulation of Hmgcr, especially at the post-transcriptional level, is incompletely understood. We set out to explore the possible roles of miRNAs in the regulation of Hmgcr. In silico predictions coupled with systematic functional analysis revealed specific interaction of miR-27a with mouse Hmgcr 3’-UTR in mouse liver fibroblasts AML-12. Furthermore, miR-27a showed a significant negative correlation with Hmgcr expression in cultured AML-12 cells as well as rat liver, kidney and skeletal muscle tissues. Ribonucleoprotein Immunoprecipitation (RIP) assays using antibodies against Ago2 in human hepatocellular HuH-7 cells over-expressing miR-27a revealed enrichment of HMGCR in the Ago2-IP fraction. Next, we quantified the expression of miR-27a and Hmgcr protein levels in liver tissues of genetically hypertensive blood pressure high (BPH) and genetically hypotensive blood pressure low (BPL) mice. Liver tissues from BPH showed elevated Hmgcr protein levels and diminished miR-27a levels as compared to BPL. Interestingly, cholesterol depletion in AML-12 cells resulted in down-regulation of endogenous miR-27a while augmenting the Hmgcr protein levels. In corroboration, exogenous cholesterol treatment diminished Hmgcr protein levels while enhancing the miR-27a levels. In addition, miR-27a expression and Egr1 protein levels were elevated in AML-12 cells under hypoxic conditions. Interestingly, computational analysis of the 1 kb promoter region of mmu-miR-27a revealed multiple putative Egr1 binding sites. Moreover, over-expression/down-regulation of Egr1 augmented/diminished the miR-27a promoter activity in both AML-12 and HuH-7 cells respectively. This study highlights the role of miR-27a in the post-transcriptional regulation of Hmgcr, thereby implicating its plausible role in cholesterol homeostasis and hypoxia-like pathological conditions.