Protein quality control in the secretory pathway
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Misfolded proteins of the secretory pathway are distinguished and degraded by cellular mechanisms collectively called 'quality control'. The three parts of this thesis work all contribute to understanding the molecular mechanisms of protein quality control. The first part is about the post-ER quality control that includes two different mechanisms, a direct delivery pathway to the vacuole and turnover from the plasma Membrane. Pma1-10, a mutant of plasma membrane H+-ATPase (Pma1), is a substrate that can be delivered to the plasma membrane but undergoes rapid turnover. I found that Pma1-10 is ubiquitinated before or upon arrival at the cell surface, which is needed for Pma1-10 internalization from the plasma membrane. Moreover, the ubiquitination of Pma1-10 is reversible. If internalization is blocked, Pma1-10 stays at the plasma membrane, loses its ubiquitination, and regains extensive phosphorylation and association with lipid rafts---considered 'stability factors for long-lived wildtype Pma1. These results suggest that rapid internalization of mutant Pma1-10 may preempt stabilizing interactions.;Second. I studied the behavior of N- or C-terminal mutants of Pma1. I found that they are recognized by two different quality control mechanisms; C-terminal mutants are degraded through ER associated degradation, and N-terminal mutants are delivered to the vacuole. Both mutants have global conformational changes as detected by trypsin digestion and have the ability to form oligoners. I propose that different quality control mechanisms may recognize distinct domains of substrates instead of global conformation.;Finally, my work has revealed that quality control in the ER is regulated by heat shock response (HSR). Activated heat shock response can promote translocation, protein degradation through ERAD and protein export from the ER in the absence of UPR. Furthermore. I showed that target genes of HSR, KAR2 and ERV29, may participate in USA-mediated suppression of ER stress in UPR-deficient cells. In the absence of UPR, ER stress induced by misfolded proteins activates heat shock response. To my best knowledge, this is the first time a role for heat shock response in ER protein quality control has been described.
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