Choutka, Courtney Paige - Regulation and conservation of caspase-activated autophagy...

This thesis has been submitted to the Library for purposes of graduation, but needs to be audited for technical details related to publication in order to be approved for inclusion in the Library collection.
Publication of this thesis has been postponed at the author's request until 2018-06-27.
Summer 2017
Degree type: 
Department of Molecular Biology and Biochemistry
Senior supervisor: 
Sharon Gorski
Publishing Documentation
Postponement release date: 
Wed, 2018-06-27
Thesis title: 
Regulation and conservation of caspase-activated autophagy
Given Names: 
Courtney Paige
Autophagy is an evolutionarily conserved cellular process that recycles proteins and organelles to maintain cellular homeostasis or provide an alternative source of energy in times of stress. While autophagy promotes cell survival, it can also be regulated by proteins associated traditionally with apoptosis. In an effort to better understand the complex intersections of these disparate cell fates, previous studies in Drosophila identified an apoptotic effector caspase, Dcp-1, as a positive regulator of starvation-induced autophagy. Further, the Drosophila heat-shock protein, Hsp83, was identified as a Dcp-1 interacting protein and a putative negative regulator of autophagy. The aims of my thesis were to investigate the relationship between Dcp-1 and Hsp83 in the context of autophagy, and to determine if caspase-regulated autophagy was functionally conserved in humans. In vivo analyses of Hsp83 loss-of-function mutants in fed conditions showed increases in both autophagic flux and cell death. Hsp83 mutants also had elevated levels of pro-Dcp-1, which was attributed to reduced proteasomal activity. Analyses of an Hsp83/Dcp-1 double mutant revealed that the caspase was not required for cell death in this context but was essential for the ensuing compensatory autophagy, female fertility, and organism viability. These studies not only demonstrated unappreciated roles for Hsp83 in proteasomal activity and new forms of Dcp-1 regulation, but also identified an effector caspase as a key regulatory factor for sustaining adaptation to cell stress in vivo by inducing compensatory autophagy. To address whether effector caspases also regulate starvation-induced autophagy in human cells, caspase-3 (CASP3), a human homolog of Dcp-1, was examined in several human cell lines. These studies showed that CASP3 was required for the upregulation of starvation-induced autophagy in most cell lines examined, but was not required for maintaining basal levels of autophagy. In human cells, another heat-shock family member, HSP60, was identified as a CASP3-interacting protein. HSP60 was shown to negatively regulate autophagy by controlling the subcellular localization of CASP3 in response to nutritional status. Epistasis analyses suggest that the increase in autophagy observed from loss of HSP60 was dependent on the accumulation of cleaved CASP3 in the cytosol. This work highlights a novel function for CASP3 in starvation-induced autophagy in human cells and illustrates how its response is regulated by HSP60-controlled subcellular localization. Altogether, my studies provide novel insights into stress adaptive relationships between heat-shock proteins and caspases in Drosophila and human cells.
caspase; autophagy; heat-shock proteins; regulation; proteostasis; stress
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