Analyzing the Stress Response of Yeast
Anne Robinson
University of Delaware
Date:
Thursday, February 16, 2006
Time: 4:00 p.m.
Place: Engineering II, Room 3361
ABSTRACT
Our research program has focused on identifying critical cellular interactions during the unfolded response during protein overexpression in the yeast Saccharomyces cerevisiae. Our laboratory seeks to use a systematic approach that goes beyond the empirical observations and optimizations common to more traditional studies, in order to establish a fundamental understanding of the key interactions in these systems.
We are studying overexpression of single-chain antibody in S cerevisiae. Prior studies of many investigators show that the antibody is retained in the endoplasmic reticulum due to slow folding, and the antibody is bound by two cellular proteins, yeast BiP (Kar2) and PDI. The accumulation of unfolded protein activates the unfolded protein response and subsequently shuts down antibody expression. Using the known interactions with cellular proteins, we developed a mechanistic model that predicted that BiP overexpression would lower cell stress and improve antibody secretion. Although antibody expression was higher when BiP levels were increased experimentally, the stress response was only marginally ameliorated. Using chemical stressors, we also found that BiP overexpression had very modest effects on cell stress. These data suggest that BiP has only a minor role in the stress response pathway activation, in contrast with earlier models of BiP-regulated dimerization of the Ire1p receptor. Our data is consistent with recent studies of Kamara and colleagues, who have found that deletion of the BiP-binding site in Ire1p does not impact the stress response of cells. I will describe the results of our experiments, taking into account the other data in the literature. Finally, this new mechanistic view of endoplasmic reticulum interactions between BiP, Ire1p, and unfolded proteins will be accounted for in a new model developed in collaboration with the Doyle group.