Abstract 840: MicroRNAs Modulate Fos Protein Expression in Response to Angiotensin II
MicroRNAs (miRNAs) are a novel family of small RNA molecules which modulate gene expression through mRNA cleavage and/or translation repression. Recent evidence suggests that miRNAs are important regulators of normal development, cell growth and differentiation. miRNAs are expressed in the cardiovascular system, however the putative roles of these miRNAs in vascular biology is completely unknown. In order to identify miRNAs potentially modulated in vessels, we performed a comprehensive screen of differentially expressed miRNAs in response to Angiotensin II (Ang II) in rat aortic vascular smooth muscle cells (VSMCs). VSMCs were treated with Ang II (1x10−5M) or vehicle for 4, 8 or 24 hrs. As a result of this screen we identified a decrease in the expression of miRNA miR-7b, at 4 and 8 hrs of treatment with Ang II in comparison to the basal level. A bioinformatic analysis to identify the targets of this miRNA revealed that the 3′UTR of the Fos gene harbors two putative binding sites for miR-7b. To validate the potential targeting of Fos by miR-7b, the Fos3′UTR sequence was cloned downstream from the Renilla luciferase coding sequence. Transient co-transfection of miR-7b with this reporter Renilla luciferase vector demonstrated down-regulation of translation by miR-7b through the Fos3′UTR, whereas a miRNA control have no effect on the reporter translation. Furthermore, in VSMCs, we demonstrate induction in Fos protein expression after Ang II stimulation; in the same conditions, overexpression of miR-7b leads to decrease in the protein levels of Fos compared to a miRNA control. In additional experiments, we found that the transfection of miR-7b has no effect in the Fos mRNA expression levels in response to Ang II, suggesting an alternative effect of miR-7b on Fos translation repression. These results show a novel level of regulation of Ang II-induced gene expression by rapidly reducing the expression of miRNAs and subsequent increase in the levels of Fos proteins. Our results are the first to describe a novel miRNA-dependent mechanism by which Fos proteins may be regulated in response to Ang II. These results may have significant impact in our understanding of Ang II signaling in health and disease.