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UBC Theses and Dissertations

Molecular mechanisms underlying the crosstalk between autophagy and apoptosis Hou, Ying Chen

Abstract

Macroautophagy (hereafter referred to as autophagy) is an evolutionary conserved mechanism for the degradation of long-lived proteins and organelles inside lysosomes. Autophagy functions as an adaptive survival response to starvation and other cellular stresses, but recent studies also demonstrated a role for autophagy in cell death. Functional studies indicate that a complex relationship exists between autophagy and apoptosis, the most common form of cell death, but the regulatory mechanisms underlying their interactions are largely unknown. To advance our understanding of the links between these two processes, I conducted a RNAi screen of Drosophila melanogaster cell death-related genes for their requirement in the regulation of starvation-induced autophagy. I discovered that six cell death genes, Dcp-1, hid, Bruce, Buffy, debcl, and p53 as well as Ras/Raf/MAPK signaling pathway components function in autophagy regulation in Drosophila l(2)mbn cells. I used Drosophila genetics to investigate a role for effector caspase Dcp-1 and inhibitor of apoptosis protein Bruce during autophagy in vivo. During Drosophila oogenesis, I found that autophagy is induced at two nutrient status checkpoints, the germarium and mid-oogenesis. Degenerating mid-stage egg chambers in DmAtg1 and DmAtg7 mutants exhibited reduced DNA fragmentation, suggesting autophagy may contribute to cell death during oogenesis. At the two nutrient status checkpoints in the developing ovary, Dcp-1 and Bruce function to regulate both autophagy and starvation-induced cell death, and epistasis analysis showed that Dcp-1 is downstream of Bruce, indicating that Bruce may negatively regulate Dcp-1 activity. In addition, I found that the catalytic activity of Dcp-1 is essential for autophagy regulation, suggesting that Dcp-1 dependent proteolysis may serve as a regulatory mechanism by which Dcp-1 mediates a cellular switch between autophagy and apoptosis. To identify potential substrates of Dcp-1, I employed an immuno-precipitation and mass spectrometry assay, the results of which can be used to generate a working model for how Dcp-1 controls autophagy. In summary, I developed an efficient screening method that resulted in the identification of several cell death-related regulators of autophagy. Further genetic and biochemical analyses of the effector caspase Dcp-1 in autophagy regulation provided new insights into the relationships between autophagy and apoptosis.

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Attribution-NonCommercial-NoDerivatives 4.0 International

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