2 years ago

2013 Scientific Report

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VARI | 2013 PI3K-mTOR and the autophagy signaling network Autophagy is a cellular recycling program essential for homeostasis and survival during cytotoxic stress. When cancer cells encounter environmental stressors such as nutrient starvation or chemotherapy, autophagy is dramatically up-regulated, resulting in cellular adaptation to the stress and subsequent survival. The autophagy process, which has an emerging role in disease etiology and treatment, is executed in four stages through the coordinated action of more than 30 proteins. An effective strategy for studying this complicated process involves the construction and analysis of computational models. When developed and refined from experimental knowledge, these models can be used to interrogate signaling pathways, formulate novel hypotheses about systems, and make predictions about cell signaling changes induced by specific interventions. In conjunction with collaborators at Los Alamos National Laboratory, we developed a computational model describing autophagic vesicle dynamics in a mammalian system. We used time-resolved live-cell microscopy to measure the synthesis and turnover of autophagic vesicles in single cells. The stochastically simulated model was consistent with data acquired during conditions of both basal and chemically induced autophagy. The model was tested by genetic modulation of the autophagic machinery and it accurately predicted the vesicle dynamics observed experimentally. Furthermore, the model generated an unforeseen prediction about vesicle size that is consistent with both published findings and our experimental observations. Thus, we have developed an accurate and useful model that can serve as the foundation for future efforts to quantitatively characterize autophagy. Ongoing efforts include building and refining a computational model of autophagy that will make reliable predictions about complex cancer cell behavior; verifying the predictions in cellular and preclinical models; and ultimately using the model to develop effective strategies for therapeutically targeting autophagy in cancer. Recent Publications Martin, Katie R., Dipak Barua, Audra L. Kauffman, Laura M. Westrate, Richard G. Posner, William S. Hlavacek, and Jeffrey P. MacKeigan. 2013. Computational model for autophagic vesicle dynamics in single cells. Autophagy 9(1): 74–92. Niemi, Natalie M., Nathan J. Lanning, Laura M. Westrate, and Jeffrey P. MacKeigan. 2013. Downregulation of the mitochondrial phosphatase PTPMT1 is sufficient to promote cancer cell death. PLoS One 8(1): e53803. Klionsky, Daniel J., Fabio C. Abdalla, Hagai Abeliovich, Robert T. Abraham, Abraham Acevedo-Arozena, Khosrow Adeli, Lotta Agholme, Maria Aganello, Patrizia Agostinis, Julio A. Aguirre-Ghiso, et al. 2012. Guidelines for the use and interpretation of assays for monitoring autophagy. Autophagy 8(4): 445–544. Looyenga, Brendan D., Danielle Hutchings, Irene Cherni, Chris Kingsley, Glen J. Weiss, and Jeffrey P. MacKeigan. 2012. STAT3 is activated by JAK2 independent of key oncogenic driver mutations in non-small cell lung carcinoma. PLoS One 7(2): e30820. Looyenga, Brendan D., and Jeffrey P. MacKeigan. 2012. Characterization of differential protein tethering at the plasma membrane in response to epidermal growth factor signaling. Journal of Proteome Research 11(6): 3101–3111. Stark, Mitchell S., Susan L. Woods, Michael G. Gartside, Vanessa F. Bonazzi, Ken Dutton-Regester, Lauren G. Aoude, Donald Chow, Chris Sereduk, Natalie M. Niemi, Nanyun Tang, et al. 2012. Frequent somatic mutations in MAP3K5 and MAP3K9 in metastatic melanoma identified by exome sequencing. Nature Genetics 44(2): 165–169. 37

Karsten Melcher, Ph.D. Laboratory of Structural Biology and Biochemistry Dr. Melcher earned his master’s in biology and his Ph.D. in biochemistry from the Eberhard Karls Universität in Tübingen, Germany, after which he was a postdoctoral fellow at the University of Texas Southwestern Medical Center in Dallas. He has been an independent investigator at the University of Ulster in Coleraine, U.K., and at Goethe University in Frankfurt. Dr. Melcher was recruited to VARI in 2007, serving as a Research Scientist within the Laboratory of Structural Sciences. In 2011, he became Assistant Professor and head of the Laboratory of Structural Biology and Biochemistry. From left: deWaal, Zhou, Li, Wang, Melcher, Kovach, Merrill, Weber Staff Amanda Kovach, B.S. Stephanie Weber, B.S. Xiaoyin (Edward) Zhou, Ph.D. Students Parker deWaal Xiaodan Li, B.S. Nate Merrill, B.S. Lili Wang, B.S. 38

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