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Session 8, Abstract 51

INVESTIGATING THE MITOCHONDRIAL DISULFIDE RELAY SYSTEM IN HUMAN PLURIPOTENT STEM CELLS WITH MITOBLOCK6

S.A. Kennedy*​1,3​ (D. Zhang​2​, C.M. Koehler​2​, C.S. Malone​1​, M.A. Teitell​3,4​) ​

1​Department of Biology, California State University, Northridge. ​2Department of Chemistry and Biochemistry, University of California, Los Angeles. ​3Department of Pathology and Laboratory Medicine, University of California at Los Angeles. ​4​Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research, Department of Pediatrics, Jonsson Comprehensive Cancer Center, California NanoSystems Institute, Molecular Biology Institute, and Department of Bioengineering, University of California at Los Angeles, Los Angeles, CA 

MitoBloCK6 (MB-6) was identified in a screen for inhibitors of the protein Augmenter of Liver Regeneration/Growth Factor Erv1-Like (ALR/GFER). ALR is a taxonomically conserved protein involved in the disulfide relay system, which facilitates import of cysteine rich proteins into the mitochondrial intermembrane space (IMS). Previous studies have shown that MB-6 causes apoptosis in human pluripotent stem cells (hPSCs) while leaving their differentiated derivatives intact. Our current work aims to determine the mechanism for MB-6 rapid cell death in hPSCs. Toward this goal, we have identified the time of differentiation, which differs based on lineage and tissue type, at which hPSC derivatives become resistant to MB-6 exposure. Antioxidants mitoTEMPO and N-acetylcysteine had little effect on hPSC death after MB-6 treatment, suggesting that reactive oxygen species (ROS) influence on the apoptotic mechanism is minimal. We hypothesize that MB-6 causes preferential death of hPSCs through differences in ALR protein import activity, and that these differences are mediated by differential protein interactions in pluripotent versus differentiated cell states.  To examine this hypothesis, we are utilizing immunoprecipitation in combination with mass spectrometry (IP-MS) to identify changes in ALR interactions between undifferentiated hPSCs and their differentiated progeny. We anticipate new insights into differences in mitochondrial function in pluripotent versus differentiated cells. In addition, MB-6 could become a tool to remove pluripotent cells that fail to differentiate into the desired cell types, furthering clinical stem cell applications.