Characterization of a novel IRE1 substrate pact and interacting miRNAS

Abstract

The double-stranded RNA-dependent protein kinase activator A (PACT) anchors the RNAinduced silencing complex (RISC) to the endoplasmic reticulum (ER)’s membranous platform where RISC nucleation occurs and thus, PACT plays a key role in microRNA (miR)-mediated translational repression. Previous studies have shown that ER stress leads to PACT phosphorylation while simultaneously inducing changes in the expression of many miRs. Here, we demonstrate that PACT is phosphorylated by the ER-resident Inositol-requiring enzyme-1 (IRE1), a bifunctional kinase/endoribonuclease (RNase), both under ER stress and no stress conditions. While the role of IRE1 as a stress-induced RNase driving the unfolded protein response (UPR) is well understood, the function or the target(s) of its kinase activity have remained unexplored. Findings of this thesis show that IRE1- mediated phosphorylation of PACT regulates mature miR-181c levels, which suppresses the expression of key regulators of mitochondrial biogenesis (mitobiogenesis). Phosphorylation by IRE1 causes PACT-mediated suppression of mitobiogenesis and respiration. Partial PACT-deficiency in mice leads to enhanced mitobiogenesis during brown fat activation in cells and mice. Furthermore, cardiopulmonary bypass-induced ischemia/reperfusion injury downregulates PACT protein expression in human hearts while simultaneously inducing mitobiogenesis. Collectively, these findings demonstrate PACTmiR- 181c signaling axis is a key regulator of mitochondrial biogenesis and energetics.