Cpr6 and Cpr7, the Saccharomyces cerevisiae homologs of cyclophilin-40 (CyP-40), were shown to form complexes with Hsp90, a protein chaperone that functions in several signal transduction pathways, Deletion of CPR7 caused severe growth defects when combined with mutations that decrease the amount of Hsp90 or Sti1, another component of the Hsp90 chaperone machinery. The activities of two heterologous Hsp90-dependent signal transducers expressed in yeast, glucocorticoid receptor and pp60(v-src) kinase, were adversely affected by cpr7 null mutations. These results suggest that CyP-40 cyclophilins play a general role in Hsp90-dependent signal transduction pathways under normal growth conditions.