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Functional maturation of steroid hormone receptors requires ordered assembly into a large multichaperone complex consisting of receptor monomer, an Hsp90 dimer, the p23 cochaperone, and an FK506-binding protein (FKBP) family member or alternate peptidylprolyl isomerase-related cochaperone. Previous cellular studies demonstrated that FKBP52 can potentiate receptor function. These results have been confirmed in fkbp4 gene knockout mice in which males are partially androgen insensitive and females display characteristics of progesterone insensitivity. Conversely, FKBP51, which has a high degree of similarity to FKBP52, antagonizes FKBP52-mediated potentiation. Both proteins consist of three domains: two FKBP12-like domains termed FK1 and FK2 and a tetratricopeptide repeat domain that targets binding to Hsp90. To help understand why the two FKBPs behave differently and to gain insight into FKBP52 potentiation activity, we have analyzed the loop structure that links FK1 and FK2. Within the FK linker of FKBP52 is the sequence TEEED, which forms a consensus casein kinase II phosphorylation site; the corresponding sequence in FKBP51 is FED. We demonstrate that the distinct FK linker sequences per se do not account for lack of potentiation activity by FKBP51. However, phosphorylation of the FK linker appears to be an important regulatory determinant of FKBP52-mediated potentiation of steroid receptor activity.

Molecular chaperones mediate multiple aspects of steroid receptor function, but the physiological importance of most receptor-associated cochaperones has not been determined. To help fill this gap, we targeted for disruption the mouse gene for the 52-kDa FK506 binding protein, FKBP52, a 90-kDa heat shock protein (Hsp90)-binding immunophilin found in steroid receptor complexes. A mouse line lacking FKBP52 (52KO) was generated and characterized. Male 52KO mice have several defects in reproductive tissues consistent with androgen insensitivity; among these defects are ambiguous external genitalia and dysgenic prostate. FKBP52 and androgen receptor (AR) are coexpressed in prostate epithelial cells of wild-type mice. However, FKBP52 and AR are similarly coexpressed in testis even though testis morphology and spermatogenesis in 52KO males are usually normal. Molecular studies confirm that FKBP52 is a component of AR complexes, and cellular studies in yeast and human cell models demonstrate that FKBP52 can enhance AR-mediated transactivation. AR enhancement requires FKBP52 peptidylprolyl isomerase activity as well as Hsp90-binding ability, and enhancement probably relates to an affect of FKBP52 on AR-folding pathways. In the presence of FKBP52, but not other cochaperones, the function of a minimally active AR point mutant can be dramatically restored. We conclude that FKBP52 is an AR folding factor that has critically important physiological roles in some male reproductive tissues.

Multiple molecular chaperones interact with steroid receptors to promote functional maturation and stability of receptor complexes. The heat shock protein (Hsp)70 cochaperone Hip has been identified in conjunction with Hsp70, Hsp90, and the Hsp70/Hsp90 cochaperone Hop/Sti1p in receptor complexes during an intermediate stage of receptor assembly, but a functional requirement for Hip in the receptor assembly process has not been established. Because the budding yeast Saccharomyces cerevisiae contains orthologs for most of the receptor-associated chaperones yet lacks an orthologous Hip gene, we exploited the well-established yeast model for steroid receptor function to ask whether Hip can alter steroid receptor function in vivo. Introducing human Hip into yeast enhances hormone-dependent activation of a reporter gene by glucocorticoid receptor (GR). Because Hip does not similarly enhance signaling by mineralocorticoid, progesterone, or estrogen receptors, a general effect on transcription can be excluded. Instead, Hip promotes functional maturation of GR without increasing steady-state levels of GR protein. Unexpectedly, Hip binding to Hsp70 is not critical for boosting GR responsiveness to hormone. In conclusion, Hip functions by a previously unrecognized mechanism to promote the efficiency of GR maturation in cells.