Characterization of Atg6 Function in Autophagy and Growth Control during Drosophila Melanogaster Development

Title of Document: CHARACTERIZATION OF ATG6 FUNCTION IN AUTOPHAGY AND GROWTH CONTROL DURING DROSOPHILA MELANOGASTER DEVELOPMENT Jahda Hope Hill, Doctor of Philosophy, 2010 Directed By: Dr. Louisa P. Wu, Associate Professor Department of Cell Biology and Molecular Genetics University of Maryland, College Park The tumor suppressor Beclin 1 mitigates cell stress by regulating the lysosomal degradation pathway known as autophagy. This process involves formation of intracellular double-membraned vesicles, known as autophagosomes, which engulf proteins and damaged organelles and fuse with lysosomes, where the contents are degraded. It is unclear whether the function of Beclin 1 in autophagy is related to cell transformation in beclin 1 animals. Using the fruit fly, Drosophila melanogaster, I investigated the function of the Beclin 1 ortholog Atg6 in autophagy and growth control. Through transgenic experiments, I found that Atg6, like Beclin 1, induces autophagy by functioning in a complex consisting of the lipid kinase Vps34 and the serine-threonine kinase Vps15. I also generated a strong loss of function mutant, Atg6, and found that Atg6 is required for development. Atg6 mutant animals contained an excess of blood cells, which surrounded melanotic tumors prior to death. At the cellular level, Atg6 is required for autophagy and endocytosis, and cells lacking Atg6 accumulate high levels of the endoplasmic reticulum stress protein Hsc3 and the adaptor protein p62. I also showed that Atg6 mutant cells displayed mis-regulated nuclear localization of NFκB proteins, transcription factors whose downstream targets include regulators of innate immunity. Significantly, my results suggest that Atg6 may regulate growth independent of its function in autophagy, as mosaic loss of Atg6 in the eye resulted in over-representation of Atg6 mutant cells, a phenotype not shared by other autophagy gene mutant mosaics. Finally, through a collaborative effort, our lab identified a novel function for Atg6 in regulation of epithelial cell polarity. This finding is significant, as epithelial tumor cells are known to lose polarity during metastasis. Our studies have provided a significant contribution to the Beclin 1 field, by providing the first characterization of a Drosophila Atg6 mutant, and demonstrating its function in novel cellular processes. CHARACTERIZATION OF ATG6 FUNCTION IN AUTOPHAGY AND GROWTH CONTROL DURING DROSOPHILA MELANOGASTER DEVELOPMENT