MUTATIONAL ANALYSIS OF THE HSP70-INTERACTING PROTEIN HIP

The hsp70-interacting protein Hip participates in the assembly pathway for progesterone receptor complexes, Duping assembly Nip appears at e arly assembly stages in a transient manner that parallels hsp70 intera ctions, In this study, a cDNA for human Hip was used to develop variou s mutant Hip forms in the initial mapping of functions to particular H ip structural elements, Hip regions targeted for deletion and/or trunc ation included the C-terminal region (which has some limited homology with Saccharomyces cerevisiae Stil and its vertebrate homolog p60), a glycine-glycine-methionine-proline (GGMP) tandem repeat, and a tetratr icopeptide repeat (TPR), Binding of Hip to hsp70's ATPase domain was L ost with deletions from the TPR and from an adjoining highly charged r egion; correspondingly, these Hip mutant forms were not recovered in r eceptor complexes, Truncation of Hip's Still-related C terminus result ed in Hip binding to hsp70 in a manner suggestive of a misfolded pepti de substrate; this hsp70 binding was localized to the GGMP tandem repe at, Mutants lacking either the C terminus or the GGMP tandem repeat we re still recovered in receptor complexes, Truncations from Hip's N ter minus resulted in an apparent loss of Hip homo-oligomerization, but th ese mutants retained association with hsp70 and were recovered in rece ptor complexes. This mutational analysis indicates that Rip's TPR is r equired For binding of Rip with hsp70's ATPase domain, In addition, so me data suggest that hsp70's peptide-binding domain may alternately or concomitantly bind to Hip's GGMP repeat in a manner regulated by Stil -related sequences.