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"Mitochondrial complex II is a source of the reserve respiratory capacity that is regulated by metabolic sensors and promotes cell survival"

by
Jessica Mary Pfleger
Cell Biology and Molecular Medicine Program
B.S. 2009, Syracuse University, NY




Thesis Advisor: Maha Abdellatif, M.D./Ph.D.
Professor
Department of Cell Biology and Molecular Medicine

Friday, June 26, 2015
12:30 P.M., MSB G609


Abstract

The survival of a cell depends on its ability to meet its energy requirements. We hypothesized that the mitochondrial reserve respiratory capacity (RRC) is a critical component of cellular bioenergetics that can be utilized during an increase in energy demand, thereby, enhancing viability. It was our goal to identify the elements that regulate and contribute to the development of RRC and its involvement in cell survival. Our results show that development of RRC is dependent on metabolic substrate availability, in a cell type-dependent manner. While the neonatal rat cardiac myocytes (NRCM) utilize glucose as their main substrate, developing RRC required fatty acids and glucose. In contrast, human iPSC-derived cardiac myocytes (iPSCCM) utilize fatty acids as their main substrate and for development of RRC, which is diminished with the addition of glucose. In both cell types, basal respiration and RRC are diminished after hypoxia/reoxygenation. Additionally, in NRCM, inhibition of either glucose or fatty acid oxidation, completely abrogated RRC. Conversely, RRC was enhanced through increasing glucose oxidation, via inhibiting pyruvate dehydrogenase kinases with dichloroacetate (DCA) in NRCM or iPSC-CM, or through increasing fatty acid oxidation, via activation of AMP-activated protein kinase with 5-aminoimidazole-4-carboxamide-1 -D-ribofuranosyl 5-monophosphate (AICAR) in NRCM. The latter was partly mediated by peroxisome proliferator-activated receptor alpha. Additionally, these DCA- or AICAR-induced increases in RRC are observed both before and after hypoxia/reoxygenation. An electron flow activity assay revealed that an increase in RRC correlated with an increase in complex II (CII) activity. Inhibiting CII activity or assembly completely abolished RRC, confirming it as the source of RRC. We also show that enhancing RRC, via AICAR treatment in NRCM, results in increased CII-dependent basal respiration and oxidative phosphorylation, accompanied by reduced superoxide production and enhanced cell survival, post-energy deprivation. Finally, we show that development of RRC also requires Sirtuin 3 (SIRT3), which may lead to CII deacetylation. Thus, for the first time, we show that manipulating metabolic sensors increases cellular RRC via activating CII in a SIRT3-dependent manner. This represents a general mitochondrial phenomenon that can be used to assess mitochondrial heath, and that can be exploited for increasing cell survival after hypoxia.


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