Title

The role of Hsp70B' in the cellular stress response

Date of Completion

January 2006

Keywords

Biology, Molecular|Biology, Cell

Degree

Ph.D.

Abstract

The human Hsp70 family is a highly conserved group of proteins with diverse protein chaperone, shuttling and stabilization functions within the cell. Hsp70B' is a unique member of the human Hsp70 family of chaperones about which information is scarce. Unlike the major inducible Hsp72 protein, Hsp70B' is strictly inducible having little or no basal expression levels in most cells. Hsp70B' is also interesting from an evolutionary point of view, as the Hsp70B' gene HSPA6 appears to have arisen after the divergence of rodents and humans. Experiments were performed to examine the role of the recently diverged Hsp70B' in protecting cells from stress in human colon cell lines. ^ We have obtained evidence that the expression, accumulation and subsequent degradation of Hsp70B' in comparison to Hsp72 after heat shock is very different. Hsp70B' appears transiently in response to stress whereas Hsp72 levels persist for many days. Hsp70B' can be induced under low cell number conditions, whereas Hsp72 accumulates under higher cell number. In addition, cytoprotection correlates with the cell number dependent expression in all cell lines examined. ^ A phylogenetic analysis of Hsp70B' showed that the Hsp70B' protein sequence is most closely related to Hsp72. Using siRNAs to knockdown Hsp70B' and Hsp72, we have found that the two are coordinately regulated in response to stress. We have also found that proteasome inhibition is a potent activator of Hsp70B'. In addition, knockdown of both Hsp70B' and Hsp72 sensitized cells to heat stress and increasing concentrations of proteasome inhibitor. From our work, it appears that Hsp72 is the primary Hsp70 family responder to increasing levels of proteotoxic stress, and Hsp70B' may be a secondary responder. Further, we have found that both Hsp70B' and Hsp72 are required to confer thermotolerance. These two proteins form a hetercomplex following stress and there may be optimal levels of each that are needed to reach full cytoprotective potential. ^ These results call attention to a number of important biological features of Hsp70B' and its contribution to the cellular stress response in colon cells. Implications of these findings may be important in pathological conditions in which Hsp70B' contributes to cell survival. ^