In vivo analysis of the Hsp90 cochaperone Sti1 (p60)

Hsp90 interacts with Sti1 (p60) in lysates of yeast and vertebrate cells, Here we provide the first analysis of their interaction in vivo. Saccharomyces cerevisiae mutations that eliminate Sti1 or reduce intracellular concentrations of Hsp90 individually: have little or no effect on growth at normal temperatures. However,when combined, the mutations greatly reduce or eliminate growth. Furthermore; overexpression of Stil has allele-specific effects on cells carrying various hsp90(fs) point mutations. These genetic interactions provide strong evidence that Hsp90 and Stil interact in vivo and that their functions are closely allied, Indeed, deletion of ST11 reduces the in Five activity of the Hsp90 target protein, glucocorticoid receptor (GR). Mutations in GR that eliminate interaction with Hsp90 also eliminate the effects of the sti1 deletion. Examination of GR protein complexes in the sli1 deletion mutant reveals a selective increase in the concentration of GR-Ydj1 complexes, supporting previous hypotheses that Ydj1 functions at an early step in the maturation of GR and that Stil acts at an intermediate step. Deletion of STI1 also reduces the in vivo activity of another, unrelated Hsp90 target protein, v-Src, Our data indicate that Stil is a general factor in the maturation of Hsp90 target proteins and support earlier suggestions that Hsp90 matures even very different target proteins by a similar mechanism.