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2013 Scientific Report

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Jeffrey P. MacKeigan,

Jeffrey P. MacKeigan, Ph.D. Laboratory of Systems Biology Dr. MacKeigan received his Ph.D. in microbiology and immunology at the University of North Carolina Lineberger Comprehensive Cancer Center in 2002, followed by a postdoctoral fellowship with John Blenis at Harvard Medical School. In 2004, he joined Novartis Institutes for Biomedical Research in Cambridge, Massachusetts, as an investigator and project leader in the Molecular and Developmental Pathways expertise platform. Dr. MacKeigan, who joined VARI in 2006, is an Associate Professor. From left: MacKeigan, Niemi, Burgenske, Martin, Westrate, Doppel, Lanning, Goodall, Looyenga, Fogg, May, Nelson, Karnes, Kauffman Staff Nicole Doppel, B.S. Vanessa Fogg, Ph.D. Audra Kauffman, M.S. Nate Lanning, Ph.D. Brendan Looyenga, Ph.D. Katie Martin, Ph.D. Brett May, B.S. Amy Nelson Students Dani Burgenske, B.S. Megan Goodall, B.S. Matt Harder Jonathan Karnes, M.S. Natalie Niemi, Ph.D. Anna Plantinga Aaron Sayfie Laura Westrate, B.A., B.S. Visiting Scientist Aaron Putzke, Ph.D. 35

Van Andel Research Institute | Scientific Report Research Interests “Systems biology” integrates multiple disciplines such as biochemistry, mathematics, and genetics to investigate unanswered biological questions. The Laboratory of Systems Biology focuses on identifying and understanding the genes and signaling pathways that, when mutated, contribute to the pathophysiology of cancer and neurodegeneration. The lab has two major research programs: cancer metabolism and the PI3K-mTOR autophagy signaling network. We employ tools such as RNA interference (RNAi), quantitative proteomics, and in silico screening to investigate the kinases and phosphatases that mediate the pro-apoptotic and cell survival functions of mitochondria, as well as those that regulate lipid signaling and autophagy. The laboratory’s primary scientific objectives are to investigate the molecular details of cancer and Parkinson’s disease; develop therapeutics for high-priority targets; and reposition drugs for use against cancer and neurodegenerative diseases. Cancer metabolism and cellular energetics Evasion of apoptosis is a significant problem in a variety of cancers. In order to identify novel regulators of apoptosis, the lab performed an RNAi screen against all kinases and phosphatases in the human genome. A possible regulator we identified was MK-STYX (encoded by the STYXL1 gene), a catalytically inactive phosphatase with homology to the MAPK phosphatases. Despite this homology, MK-STYX knockdown failed to modulate MAPK signaling in response to growth factors or apoptotic stimuli. Rather, RNAi-mediated knockdown of MK-STYX prevented cells from undergoing apoptosis induced by cellular stressors, activating mitochondrial-dependent apoptosis. This MK-STYX phenotype mimicked the loss of Bax and Bak, two potent guardians of mitochondrial apoptotic potential: cells without MK-STYX expression were unable to release cytochrome c. The overexpression of pro-apoptotic Bcl-2 proteins was unable to trigger cytochrome c release in MK-STYX knockdown cells, placing the apoptotic deficiency at the level of mitochondrial outer membrane permeabilization (MOMP). MK-STYX localizes to the mitochondria, but it is neither released from the mitochondria upon apoptotic stress nor localized proximal to the machinery currently known to control MOMP. Thus, MK-STYX regulates the chemoresistance potential of cancer cells through the control of MOMP, but in distinct fashion from currently characterized mechanisms. Additionally, we have determined that MK-STYX interacts with a mitochondrial phosphatase, PTPMT1. The loss of PTPMT1 in MK-STYX knockdown cells resensitizes the cells to chemotherapy and cytochrome c release, demonstrating a genetic interaction between these two proteins. Ongoing studies are focused on characterizing this MK-STYX–PTPMT1 interaction and on gaining insight into the metabolic and apoptotic capacity of cancer cells. A wealth of experimental evidence clearly connects the regulation of cellular metabolism with the development of cancer. Metabolic changes in cancer cells are considered a key event in the transition from a normal cell to a cancer cell. Such changes cause cancer cells to be metabolically reprogrammed to provide the fuel and energy required for rapid proliferation. To identify genes crucial for cancer cell metabolism, we developed a novel, high-throughput method to comprehensively screen all known nuclear-encoded genes whose protein products localize to mitochondria. Our screen also included other metabolic genes and used cellular ATP levels as a readout. The screen was performed under both glycolytic and oxidative phosphorylation-restricted conditions to define genes contributing to ATP production in each bioenergetic state. We identified several genes that drive cancer cell bioenergetics and upon which cancer cells rely for survival and proliferation. A substantial proportion of the genes we identified as novel targets were dysregulated in tumors from glioma patients, and their expression and copy number status significantly correlated with patient survival. Current experiments seek to answer questions about the cellular interactions involving these target genes and how these interactions affect the metabolic programs of normal and cancer cells. 36

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