Cns1 Is an Essential Protein Associated with the Hsp90 Chaperone Complex in Saccharomyces cerevisiae That Can Restore Cyclophilin 40-Dependent Functions in cpr7ΔCells

Abstract

Saccharomyces cerevisiae harbors two cyclophilin 40-type enzymes, Cpr6 and Cpr7, which are components of the Hsp90 molecular chaperone machinery. Cpr7 is required for normal growth and is required for maximal activity of heterologous Hsp90-dependent substrates, including glucocorticoid receptor (GR) and the oncogenic tyrosine kinase pp60 v-src . In addition, it has recently been shown that Cpr7 plays a major role in negative regulation of the S. cerevisiae heat shock transcription factor (HSF). To better understand functions associated with Cpr7, a search was undertaken for multicopy suppressors of the cpr7⌬ slow-growth phenotype. The screen identified a single gene, designated CNS1 (for cyclophilin seven suppressor), capable of suppressing the cpr7⌬ growth defect. Overexpression of CNS1 in cpr7⌬ cells also largely restored GR activity and negative regulation of HSF. In vitro protein retention experiments in which Hsp90 heterocomplexes were precipitated resulted in coprecipitation of Cns1. Interaction between Cns1 and the carboxy terminus of Hsp90 was also shown by two-hybrid analysis. The functional consequences of CNS1 overexpression and its physical association with the Hsp90 machinery indicate that Cns1 is a previously unidentified component of molecular chaperone complexes. Thus far, Cns1 is the only tetratricopeptide repeat-containing component of Hsp90 heterocomplexes found to be essential for cell viability under all conditions tested.Cells rely on molecular chaperones to facilitate the folding of nascent polypeptides, prevent protein aggregation, and enable proper inter-and intramolecular interactions. Indispensable in both Saccharomyces cerevisiae (5) and Drosophila melanogaster (13), Hsp90 is one of the most abundant and highly conserved chaperones. In vitro, Hsp90 displays general chaperone properties by preventing the aggregation of proteins such as citrate synthase (52) and casein kinase II (30) and by maintaining ␤-galactosidase (21) in a folding-competent state. In vivo, however, Hsp90 is not required for the folding of most proteins (33) but instead plays a key role in the maturation of a small subset of proteins typically involved in signal transduction (3,32,42). It is needed for the maturation of certain steroid hormone receptors (40,43,44), basic helix-loop-helix transcription factors (45

Cpr6 and Cpr7, Two Closely Related Hsp90-associated Immunophilins from Saccharomyces cerevisiae, Differ in Their Functional Properties

“…In contrast, the deletion of Cpr7 caused a growth defect (28,45). Additionally, Cpr7-depleted cells are more sensitive to geldanamycin, a specific Hsp90 inhibitor, and are defective in restoring the full glucocorticoid receptor activity (30). Our results demonstrate that purified Cpr6 and Cpr7 differ in their stability, their ability to isomerase Xaa-Pro peptide bonds, and in their activity as molecular chaperones, although they share 47% sequence homology and 38% sequence identity (28).…”

“…Cpr6 and Cpr7 are both constitutively expressed, but only Cpr6 is additionally induced by heat shock (29). Depletion of Cpr7 led to a growth defect that could not be rescued by the overexpression of Cpr6 (28,30). In vitro it was shown that Cpr6 is able to catalyze the isomerization of Xaa-Pro peptide bonds (29,31), whereas Cpr7 is assumed to be inactive (32).…”

Sti1 and Cdc37 Can Stabilize Hsp90 in Chaperone Complexes with a Protein Kinase

“…CPR6 and CPR7 have significant sequence similarity with each other but completely different functions (Figure 1). Also, defects associated with deletion of CPR7 cannot be compensated for by overexpressing CPR6 (Dolinski et al, 1998;Marsh et al, 1998). Two essential cochaperones that have no paralogs, Cdc37 and Cns1, were studied by their ability to suppress defects associated with sti1⌬ and cpr7⌬ strains, respectively, and will be discussed in more detail below.…”

“…This indicates that Cpr7 has a separate function from Cdc37, although both chaperones are clearly part of the same Hsp90 machinery and indeed have been shown to interact with each other (Abbas-Terki et al, 2002). Previous studies established that the growth phenotype of cpr7⌬ could be suppressed by CNS1 overexpression, as could a defect in GR activation (Dolinski et al, 1998;Marsh et al, 1998). On the other hand, it was recently found that temperature-sensitive cns1 mutants that also displayed a defect in GR signaling were wild-type for MAP kinase signaling (Tesic et al, 2003).…”

Chaperoning the Chaperone: A Role for the Co-chaperone Cpr7 in Modulating Hsp90 Function in Saccharomyces cerevisiae

“…Cpr6 and Cpr7 share 38% amino acid sequence identity, but overexpression of CPR6 does not rescue growth phenotypes or defects in Hsp90 client activity in cells lacking CPR7 (Dolinski et al 1998;Marsh et al 1998), indicating that they have different functions in vivo. Cpr6 and Cpr7 contain two main domains: an amino-terminal PPIase domain and a carboxy-terminal TPR domain.…”