Sti1 Is a Novel Activator of the Ssa Proteins

Abstract

The molecular chaperones Hsp70 and Hsp90 are involved in the folding and maturation of key regulatory proteins in eukaryotes. Of specific importance in this context is a ternary multichaperone complex in which Hsp70 and Hsp90 are connected by Hop. In Saccharomyces cerevisiae two components of the complex, yeast Hsp90 (yHsp90) and Sti1, the yeast homologue of Hop, had already been identified, but it remained to be shown which of the 14 different yeast Hsp70s are part of the Sti1 complex and what were the functional consequences resulting from this interaction. With a two-hybrid approach and co-immunoprecipitations, we show here that Sti1 specifically interacts with the Ssa group of the cytosolic yeast Hsp70 proteins. Using purified components, we reconstituted the dimeric Ssa1-Sti1 complex and the ternary Ssa1-Sti1-yHsp90 complex in vitro. The dissociation constant between Sti1 and Ssa1 was determined to be 2 orders of magnitude weaker than the affinity of Sti1 for yHsp90. Surprisingly, binding of Sti1 activates the ATPase of Ssa1 by a factor of about 200, which is in contrast to the behavior of Hop in the mammalian Hsp70 system. Analysis of the underlying activation mechanism revealed that ATP hydrolysis is rate-limiting in the Ssa1 ATPase cycle and that this step is accelerated by Sti1. Thus, Sti1 is a potent novel effector for the Hsp70 ATPase.

Saccharomyces cerevisiae KNU5377 Stress Response during High-Temperature Ethanol Fermentation

“…Hsp90 depends on its association with a variety of cochaperones [Hsp40 (or Hsp42) and cyclophilins], and cofactors (Mayr et al, 2000;Zhao et al, 2005). Hsp90 complex formation is mediated by an adaptor protein, Sti, and an essential ER chaperone, Grp (Grp94) (Chu et al, 2006), which functions as an interaction domain for Hsp90 (Wegele et al, 2003). Zpr1 is a zinc finger (ZnF) protein that translocates to the nucleus in response to growth stimuli and is a lethal partner with Cpr1p for PPIase activity.…”

Cns1 Is an Activator of the Ssa1 ATPase Activity

“…Cns1 Binds to Ssa1 and Activates Its ATPase-For Sti1, it had been shown recently that, in addition to binding to Hsp90, it also interacts with the Ssa proteins (16). Therefore, we tested whether Cns1 is also able to bind to Ssa1.…”

“…Another partner protein, Sti1 (stress-inducible protein 1), inhibits the ATPase of yeast Hsp90 as a noncompetitive inhibitor (14,15). Furthermore, Sti1 was recently identified as a potent activator of the Ssa members of the Hsp70 family (6,16). Hop (Hsp70/Hsp90 organizing protein), the mammalian homologue of Sti1 (17,18), seems to be of central importance for regulating the Hsp90 chaperone cycle, as it binds to the C termini of both Hsp90 and Hsp70 (19), thus providing a physical link between the Hsp70 and Hsp90 chaperone machineries.…”

Evidence for an Unfolding/Threading Mechanism for Protein Disaggregation by Saccharomyces cerevisiae Hsp104

“…Even more recently it has been reported that Sti1 also stimulates the ATPase activity of Ssa1 (yeast cytosolic Hsp70) by about 200-fold (40). The significance of these interactions with respect to the function of Hsp104 has yet to be established.…”