Human cyclophilin 40 is a heat shock protein that exhibits altered intracellular localization following heat shock

The unactivated steroid receptors are chaperoned into a conformation that i s optimal for binding hormone by a number of heat shock proteins, including Hsp90, Hsp70, Hsp40, and the immunophilin, FKBP52 (Hsp56). Together with i ts partner cochaperones, cyclophilin 40 (CyP40) and FKBP51, FKBP52 belongs to a distinct group of structurally related immunophilins that modulate ste roid receptor function through their association with Hsp90. Due to the str uctural similarity between the component immunophilins, FKBP52 and cyclophi lin 40, we decided to investigate whether CyP40 is also a heat shock protei n. Exposure of MCF-7 breast cancer cells to elevated temperatures (42 degre esC for 3 hours) resulted in a 75-fold increase in CyP40 mRNA levels, but n o corresponding increase in CyP40 protein expression, even after 7 hours of heat stress. The use of cycloheximide to inhibit protein synthesis reveale d that in comparison to MCF-7 cells cultured at 37 degreesC, those exposed to heat stress (42 degreesC for 3 hours) displayed an elevated rate of degr adation of both CyP40 and FKBP52 proteins. Concomitantly, the half-life of the CyP40 protein was reduced from more than 24 hours to just over 8 hours following heat shock. As no alteration in CyP40 protein levels occurred in cells exposed to heat shock, an elevated rate of degradation would imply th at CyP40 protein was synthesized at an increased rate. hence the designatio n of human CyP40 as a heat shock protein. Application of heat stress elicit ed a marked redistribution of CyP40 protein in MCF-7 cells from a predomina ntly nucleolar localization, with some nuclear and cytoplasmic staining, to a pattern characterized by a pronounced nuclear accumulation of CyP40, wit h no distinguishable nucleolar staining. This increase in nuclear CyP40 pos sibly resulted from a redistribution of cytoplasmic and nucleolar CyP40, as no net increase in CyP40 expression levels occurred in response to stress. Exposure of MCF-7 cells to actinomycin D for 4 hours resulted in the trans location of the nucleolar marker protein, B23, from the nucleolus, with onl y a small reduction in nucleolar CyP40 levels. Under normal growth conditio ns, MCF-7 cells exhibited an apparent colocalization of CyP40 and FKBP52 wi thin the nucleolus.