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B.S. 2008, New Jersey Institute of Technology
Thesis Advisor: Hongyu Qiu, M.D., Ph.D.
Department of Cell Biology and Molecular Medicine
Friday, April 26, 2013
1:00 P.M., MSB G609
The heart is one of the most energy demanding tissues relying largely on energy produced by oxidative phosphorylation. Consequently the mitochondria, which is responsible for producing most of the necessary ATP, is a critical organelle in the cardiac myocyte. H11 kinase/Hsp22 (Hsp22) is a stress-inducible small heat-shock protein that confers cardioprotection via an inducible nitric oxide synthase (iNOS) effect that is equivalent to ischemic preconditioning, the “gold standard” of cardiac protection. The goal of this present proposal is to further define the function of Hsp22 in the mitochondria in the heart. The overall hypothesis of this study is that the cardioprotective effects of Hsp22 is dependent upon its mitochondrial translocation and mediated by the induction and mitochondrial distribution of inducible nitric oxide synthase (iNOS) through the activation of the Valosin-containing protein (VCP).
Accordingly, this thesis has three components: in the first part, we tested our sub-hypothesis 1 that the mitochondrial transfer of Hsp22 in mammalian myocytes is necessary for its cardioprotective effects, and that this translocation is N-terminal dependent. We first determined the location of Hsp22 in the mammalian mitochondria and identified that Hsp22 is primarily bound to the inner membrane of the submitochondria. Next, we identified that the first 20 amino acids of the N-terminus are responsible for the intra-mitochondrial localization of Hsp22 in cardiac myocytes. Furthermore, we found that translocation of Hsp22 to the mitochondrial is necessary for stimulation of the oxidative phosphorylation chain and protection against cardiomyocyte death. In the second part of our study, we tested our sub-hypothesis 2 that the role of Hsp22 in the regulation of mitochondrial respiration is mediated by the induction and mitochondrial translocation of iNOS. This hypothesis was based on a previous study in my mentor’s lab showing that Hsp22 confers cardiac protection against ischemia that is equivalent to that conferred by the second window of preconditioning (SWOP) by an NO-dependent mechanism. In the present study, we found that overexpression of Hsp22 increased the translocation of iNOS into the mitochondria. We also showed that upon iNOS inhibition, the stimulatory effect of Hsp22 on mitochondrial respiration was blocked. In the third part, we tested our third hypothesis that VCP is a crucial mediator of mitochondrial function of Hsp22 through iNOS and a subsequent regulator of mitochondrial protein S-nitrosylation. We found in this study that overexpression of Hsp22 increased VCP abundance in the mitochondria. We also showed that VCP promotes mitochondrial respiration in an iNOS dependent manner through the regulation of S-nitrosylation of mitochondrial proteins. Thus, we have shown that the mitochondrial translocation of Hsp22 may be a critical mechanism by which its cardioprotective effects are mediated; thus allowing the myocytes to better adapt during situation of cardiac stress and consequently increase their survival rate.