Essential In Vivo Roles of the Hsp90 Molecular Chaperone

Northwestern University, Evanston IL

Investigators

Abstract

MCB 0079249 Gaber The PI has discovered that Cns1 is an essential co-chaperone that physically and functionally interacts with the Hsp90 complex of proteins. The Cns1-Hsp90 interaction is direct, being mediated via the tetratricopeptide (TPR) repeat of Cns1 and the carboxy terminus of Hsp90. The PI has isolated temperature-sensitive alleles of CNS1 that are temperature-sensitive for growth. Several of the temperature-sensitive mutations at CNS1 abolish in vitro interactions between Cns1 and Hsp90. Because both Hsp90 and Cns1 are essential, this provides circumstantial evidence that the Hsp90-CnsI physical interaction is essential. Confirmation of this would be a major new discovery in the molecular chaperone field as none of the Hsp90-cochaperone interactions investigated to date have been shown to be essential. In this project, the PI will use suppressor genetics to identify mutations that restore the ability of cns1-ts cells to grow at the restrictive temperature. One class of mutations is expected to identify key sites in Hsp90 that restore interactions with Cns1-ts proteins at the restrictive temperature. In vivo and in vitro confirmation of these restored physical interactions would provide proof that the interaction between Hsp90 and Cns1 is essential for life. In addition, the general genetic approach is expected to yield the identification of other essential proteins that interact with Hsp90 and/or Cns1, and thus enhance our understanding of essential molecular interactions involving these chaperones and co-chaperones. Proper growth control and development in all organisms depends upon the regulation of proteins that play central roles transducing signals that the cell receives from its environment. Molecular chaperones are proteins that help signal transducing proteins to achieve a three-dimensional state from which they can most efficiently transmit signals that ultimately change the profile of gene expression of the cell. Two highly investigated molecular chaperones are heat shock protein 70 (Hsp70) and heat shock protein 90 (Hsp90), so-called because of the original discovery that their abundance in the cell increases upon heat treatment. Although the role that Hsp70 plays in helping proteins fold into their proper configuration is well established, the precise role(s) played by Hsp90 in molecular chaperoning is (are) not well-known, despite the investigation of many groups over the last decade. This PI has developed a system by which the most important in vivo role of Hsp90, i.e., the role(s) it plays that is (are) essential to cell viability, can be investigated using yeast genetics. The goal of this investigation is to identify physical interactions involving Hsp90, a newly discovered protein partner of Hsp90 called Cns1, and other proteins that are required for the life of the cell. By establishing the molecular identity of these proteins and details of their physical associations, the essential role played by Hsp90 in vivo will become clearer.