Abstract 1784: Mir-199a Is A Key Regulator Of Hif-1alpha And An Inhibitor Of Hypoxia-induced Apoptosis
MicroRNAs (miRNA) are touted to be fundamental posttranscriptional regulators of gene expression. They are differentially regulated during health and disease states with a major impact on cellular functions. We found that miR-199a is upregulated (4.8 ± 1 fold) during compensatory cardiac hypertrophy, returning to basal levels in the failing heart, and is virtually undetectable during cardiac ischemia. The response of miR-199a to ischemia is rapid through posttranscriptional inhibition of its processing. To determine its targets we analyzed myocytes overexpressing miR-199a by using an antibody array analysis, the outcome of which unequivocally indicated an antiapoptotic function of this miRNA. Target prediction software revealed hypoxia-inducible factor 1 alpha (Hif-1α) as one of its potential targets. We confirmed this by protein expression, luciferase-miR-199a target reporter, and recombinant Hif-1α gene expression under hypoxic conditions in cardiac myocytes. The results show that miR-199a inhibited hypoxia-induced Hif-1α > 70%, and vice versa, knockdown of miR-199a by a miR-199a ‘eraser’ resulted in its upregulation (1.8 fold). Hif-1α, which is undetectable during normoxia and induced by hypoxia, mediates hypoxia-induced apoptosis through a p53-dependent pathway. MiR-199a overexpression in myocytes exposed to anoxia or ischemia resulted in inhibition of Hif-1α expression, as well as, >80% inhibition of the increase in p53. Moreover, miR-199a inhibited hypoxia-induced expression of caspase-6, caspase-9, caspase-12, and caspase-3 activity (~60%). To determine if the protective effect of miR-199a was indeed mediated through inhibition of Hif-1α, we co-delivered a miR-199a-indepndent Hif-1α construct to the myocytes. This resulted in complete reversal of the miR-199a effect on anoxia-induced p53 expression and its downstream caspase effectors. Thus, our results indicate that downregulation of miR-199a by hypoxia is necessary for enhancing expression of Hif-1α and its downstream apoptosis effector p53. Moreover, we show that replenishing miR-199a protects the myocytes against ischemia- or hypoxia-induced apoptosis.