FK506-binding Protein 52 Research Articles

NFAT activation by FKBP52 promotes cancer cell proliferation by suppressing p53.

FK506-binding protein 52 (FKBP52) is a member of the FKBP family of proline isomerases. FKBP52 is up-regulated in various cancers and functions as a positive regulator of steroid hormone receptors. Depletion of FKBP52 is known to inhibit cell proliferation; however, the detailed mechanism remains poorly understood. In this study, we found that FKBP52 depletion decreased MDM2 transcription, leading to stabilization of p53, and suppressed cell proliferation. We identified NFATc1 and NFATc3 as transcription factors that regulate MDM2 We also found that FKBP52 associated with NFATc3 and facilitated its nuclear translocation. In addition, calcineurin, a well-known Ca2+ phosphatase essential for activation of NFAT, plays a role in MDM2 transcription. Supporting this notion, MDM2 expression was found to be regulated by intracellular Ca2+ Taken together, these findings reveal a new role of FKBP52 in promoting cell proliferation via the NFAT-MDM2-p53 axis, and indicate that inhibition of FKBP52 could be a new therapeutic tool to activate p53 and inhibit cell proliferation.

Impact of FKBP52 on cell proliferation and hormone-dependent cancers.

FK506 binding protein 52 (FKBP52) (gene name FKBP4) is a 52 kDa protein that belongs to the FKBP family; it binds to the immunosuppressant FK506 and has proline isomerase activity. In addition to its FK domain-containing peptidylprolyl isomerase activity, FKBP52 also acts as a cochaperone through the tetratricopeptide repeat domain that mediates binding to heat shock protein 90. Previous studies have reported that FKBP52 is associated with hormone-dependent, stress-related, and neurodegenerative diseases, revealing its diverse functions. In particular, the effects of FKBP52 on cancer have attracted considerable attention. FKBP52 promotes the growth of hormone-dependent cancers by activating steroid hormone receptors. Recent studies have shown that the expression of FKBP52 is increased not only in steroid hormone-dependent cancer cells but also in colorectal, lung, and liver cancers, revealing its diverse functions that contribute to cancer growth. This review summarizes reports related to hormone-dependent cancer and cell proliferation in terms of the structure of FKBP52 and its function on interacting molecules.

FKBP52 and FKBP51 differentially regulate the stability of estrogen receptor in breast cancer

Estrogen receptor α (ERα) is a transcription factor that induces cell proliferation and exhibits increased expression in a large subset of breast cancers. The molecular mechanisms underlying the up-regulation of ERα activity, however, remain poorly understood. We identified FK506-binding protein 52 (FKBP52) as a factor associated with poor prognosis of individuals with ERα-positive breast cancer. We found that FKBP52 interacts with breast cancer susceptibility gene 1 and stabilizes ERα, and is essential for breast cancer cell proliferation. FKBP52 depletion resulted in decreased ERα expression and proliferation in breast cancer cell lines, including MCF7-derived fulvestrant resistance (MFR) cells, suggesting that inhibiting FKBP52 may provide a therapeutic effect for endocrine therapy–resistant breast cancer. In contrast, FKBP51, a closely related molecule to FKBP52, reduced the stability of ERα. Consistent with these findings, FKBP51 was more abundantly expressed in normal tissues than in cancer cells, suggesting that these FKBPs may function in the opposite direction. Collectively, our study shows that FKBP52 and FKBP51 regulate ERα stability in a reciprocal manner and reveals a regulatory mechanism by which the expression of ERα is controlled.

FKBP52 in Neuronal Signaling and Neurodegenerative Diseases: A Microtubule Story.

The FK506-binding protein 52 (FKBP52) belongs to a large family of ubiquitously expressed and highly conserved proteins (FKBPs) that share an FKBP domain and possess Peptidyl-Prolyl Isomerase (PPIase) activity. PPIase activity catalyzes the isomerization of Peptidyl-Prolyl bonds and therefore influences target protein folding and function. FKBP52 is particularly abundant in the nervous system and is partially associated with the microtubule network in different cell types suggesting its implication in microtubule function. Various studies have focused on FKBP52, highlighting its importance in several neuronal microtubule-dependent signaling pathways and its possible implication in neurodegenerative diseases such as tauopathies (i.e., Alzheimer disease) and alpha-synucleinopathies (i.e., Parkinson disease). This review summarizes our current understanding of FKBP52 actions in the microtubule environment, its implication in neuronal signaling and function, its interactions with other members of the FKBPs family and its involvement in neurodegenerative disease.

Hsp90 co-chaperones, FKBP52 and Aha1, promote tau pathogenesis in aged wild-type mice

The microtubule associated protein tau is an intrinsically disordered phosphoprotein that accumulates under pathological conditions leading to formation of neurofibrillary tangles, a hallmark of Alzheimer’s disease (AD). The mechanisms that initiate the accumulation of phospho-tau aggregates and filamentous deposits are largely unknown. In the past, our work and others’ have shown that molecular chaperones play a crucial role in maintaining protein homeostasis and that imbalance in their levels or activity can drive tau pathogenesis. We have found two co-chaperones of the 90 kDa heat shock protein (Hsp90), FK506-binding protein 52 (FKBP52) and the activator of Hsp90 ATPase homolog 1 (Aha1), promote tau aggregation in vitro and in the brains of tau transgenic mice. Based on this, we hypothesized that increased levels of these chaperones could promote tau misfolding and accumulation in the brains of aged wild-type mice. We tested this hypothesis by overexpressing Aha1, FKBP52, or mCherry (control) proteins in the hippocampus of 9-month-old wild-type mice. After 7 months of expression, mice were evaluated for cognitive and pathological changes. Our results show that FKBP52 overexpression impaired spatial reversal learning, while Aha1 overexpression impaired associative learning in aged wild-type mice. FKBP52 and Aha1 overexpression promoted phosphorylation of distinct AD-relevant tau species. Furthermore, FKBP52 activated gliosis and promoted neuronal loss leading to a reduction in hippocampal volume. Glial activation and phospho-tau accumulation were also detected in areas adjacent to the hippocampus, including the entorhinal cortex, suggesting that after initiation these pathologies can propagate through other brain regions. Overall, our findings suggest a role for chaperone imbalance in the initiation of tau accumulation in the aging brain.

The Influence Mechanism of Abnormal Immunophilin FKBP52 on the Expression Levels of PR-A and PR-B in Endometriosis Based on Endometrial Stromal Cell Model in Vitro

ABSTRACT As a chaperone protein of progesterone receptor (PR), FK-506 Binding Protein 52 (FKBP52) can enhance the activity of PR, but the mechanism of FKBP52 affecting PR expression levels is difficult to clarify. Here, we report a novel in vitro model of ectopic endometrial stromal cells (ESCM) established through the primary culture method of endometrial stromal cells, which is used to study the details of relationship between FKBP52 abnormality and PR expression level in endometriosis (Ems). At the same time, the clinical study of the relationship between FKBP52 and PR expression levels in endometriosis patients was used to verify our conclusions. The results showed that the expression levels of PR-A mRNA and protein in endometriosis are positively correlated with FKBP52 and the abnormality of FKBP52 leads to the decrease of PR-B mRNA and protein expression. When FKBP52 was deleted or reduced, the expression levels of m RNA and protein of PR-A and PR-B have decreased leading to the proliferation of ectopic endometrium cells (ESC) and the occurrence of endometriosis, which is consistent with the expression levels of clinical endometriosis patients and fully confirms our conclusions and reliability of the model, and has great guiding significance for the research of Ems disease occurrence mechanism and clinical treatment.

Tacrolimus Prevents TWEAK-Induced PLA2R Expression in Cultured Human Podocytes.

Primary membranous nephropathy is usually caused by antibodies against the podocyte antigen membrane M-type phospholipase A2 receptor (PLA2R). The treatment of membranous nephropathy is not fully satisfactory. The calcineurin inhibitor tacrolimus is used to treat membranous nephropathy, but recurrence upon drug withdrawal is common. TNF superfamily members are key mediators of kidney injury. We have now identified key TNF receptor superfamily members in podocytes and explored the regulation of PLA2R expression and the impact of tacrolimus. Data mining of single cell transcriptomics and glomerular transcriptomics data identified TNFRSF12a/Fn14 as the highest expressed TNF receptor superfamily gene in human membranous nephropathy, and this was confirmed by immunohistochemistry that also identified NFκB activation in membranous nephropathy podocytes. Additionally, glomerular transcriptomics identified PLA2R1 expression as being increased in membranous nephropathy in the parenteral administration of the Fn14 ligand TWEAK increased podocyte PLA2R expression in mice. Furthermore, in cultured human podocytes, TWEAK increased the expression of PLA2R as well as the expression of other genes recently identified by GWAS as linked to membranous nephropathy: NFKB1 and IRF4. Interestingly, IRF4 encodes the FK506-binding protein 52 (FKBP52), a protein associated with tacrolimus. Tacrolimus prevented the increased expression of PLA2R, NFKB1 and IRF4 induced by TWEAK in cultured podocytes. In conclusion, TWEAK upregulates the expression of PLA2R and of other genes linked to membranous nephropathy in podocytes, and this is prevented by tacrolimus. An impact of tacrolimus on the expression of PLA2R and other genes in podocytes may underlie its efficacy in treating the disease as well as the frequent recurrence of nephrotic syndrome upon tacrolimus withdrawal.

FKBP52 regulates TRPC3-dependent Ca2+ signals and the hypertrophic growth of cardiomyocyte cultures.

The transient receptor potential (TRP; C-classical, TRPC) channel TRPC3 allows a cation (Na+/Ca2+) influx that is favored by the stimulation of Gq protein-coupled receptors (GPCRs). An enhanced TRPC3 activity is related to adverse effects, including pathological hypertrophy in chronic cardiac disease states. In the present study, we identified FK506-binding protein 52 (FKBP52, also known as FKBP4) as a novel interaction partner of TRPC3 in the heart. FKBP52 was recovered from a cardiac cDNA library by a C-terminal TRPC3 fragment (amino acids 742-848) in a yeast two-hybrid screen. Downregulation of FKBP52 promoted a TRPC3-dependent hypertrophic response in neonatal rat cardiomyocytes (NRCs). A similar effect was achieved by overexpressing peptidyl-prolyl isomerase (PPIase)-deficient FKBP52 mutants. Mechanistically, expression of the FKBP52 truncation mutants elevated TRPC3-mediated currents and Ca2+ fluxes, and the activation of calcineurin and the nuclear factor of activated T-cells in NRCs. Our data demonstrate that FKBP52 associates with TRPC3 via an as-yet-undescribed binding site in the C-terminus of TRPC3 and modulates TRPC3-dependent Ca2+ signals in a PPIase-dependent manner. This functional interaction might be crucial for limiting TRPC3-dependent signaling during chronic hypertrophic stimulation.

Expression of FKBP52 in the ovaries of PCOS rats.

The present study aimed to examine the expression of FK-506 binding protein 52 (FKBP52) in the ovary tissues of rats with polycystic ovarian syndrome (PCOS) and its action on mediating androgen receptor (AR) through the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) pathway. PCOS model rats were established by dehydroepiandrosterone injection. Enzyme-linked immunosorbent assay (ELISA) measured serum sex hormones. Hematoxylin and eosin (H&E) staining was used to examine histological changes of the ovarian tissues. The expression levels of FKBP52 were detected by immunohistochemical (IHC) staining, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis and western blotting (WB). In addition, RT-qPCR analysis was used to detect the mRNA expression of AR, and WB was used to detect the protein expression levels of AR, ERK1/2 and phosphorylated (p-) ERK1/2. In granulosa cell (GC) experiments, primary GCs were extracted and cultured. FKBP4 is the FKBP52-encoding gene, therefore, adenovirus vectors Ad-Oe-FKBP4-EGFP and Ad-siRNA-FKBP4-EGFP were constructed to examine the association among the above factors using the RT-qPCR and WB methods. In the animal experiment, the vaginal smear, H&E staining and ELISA results showed that the PCOS model was successfully established. The IHC staining revealed that the expression of FKBP52 in the GCs of the PCOS model group was higher than the remaining groups (P<0.01). The mRNA and expression levels of FKBP52 and AR in the PCOS model rats were significantly increased, when compared with levels in the other rats (P<0.05). The expression level of p-ERK1/2 was also higher (P<0.05). In the GC experiment, following overexpression of the FKBP4 gene, the mRNA and expression levels of FKBP52 and AR were increased (P<0.05). The expression level of p-ERK1/2 was also increased (P<0.05). Following FKBP4 gene silencing, the mRNA and expression levels of FKBP52 and AR were decreased (P<0.05). The expression level of ERK1/2 was also decreased (P<0.05). However, the expression level of p-ERK1/2 was increased (P<0.05). In conclusion, the upregulation of co-chaperone FKBP52 may mediate the activation of AR through the MAPK/ERK pathway.

Molecular dynamics simulation, binding free energy calculation and unbinding pathway analysis on selectivity difference between FKBP51 and FKBP52: Insight into the molecular mechanism of isoform selectivity.

As co-chaperones of the 90-kDa heat shock protein(HSP90), FK506 binding protein 51 (FKBP51) and FK506 binding protein 52 (FKBP52) modulate the maturation of steroid hormone receptor through their specific FK1 domains (FKBP12-like domain 1). The inhibitors targeting FK1 domains are potential therapies for endocrine-related physiological disorders. However, the structural conservation of the FK1 domains between FKBP51 and FKBP52 make it difficult to obtain satisfactory selectivity in FK506-based drug design. Fortunately, a series of iFit ligands synthesized by Hausch et al exhibited excellent selectivity for FKBP51, providing new opportunity for design selective inhibitors. We performed molecular dynamics simulation, binding free energy calculation and unbinding pathway analysis to reveal selective mechanism for the inhibitor iFit4 binding with FKBP51 and FKBP52. The conformational stability evaluation of the "Phe67-in" and "Phe67-out" states implies that FKBP51 and FKBP52 have different preferences for "Phe67-in" and "Phe67-out" states, which we suggest as the determinant factor for the selectivity for FKBP51. The binding free energy calculations demonstrate that nonpolar interaction is favorable for the inhibitors binding, while the polar interaction and entropy contribution are adverse for the inhibitors binding. According to the results from binding free energy decomposition, the electrostatic difference of residue 85 causes the most significant thermodynamics effects on the binding of iFit4 to FKBP51 and FKBP52. Furthermore, the importance of substructure units on iFit4 were further evaluated by unbinding pathway analysis and residue-residue contact analysis between iFit4 and the proteins. The results will provide new clues for the design of selective inhibitors for FKBP51.

Molecular mechanisms of repeated social defeat-induced glucocorticoid resistance: Role of microRNA.

Glucocorticoid (GC) resistance is a severe problem associated with various inflammatory diseases. Previous studies have shown that repeated social stress induces GC resistance in innate immune cells, but the underlying molecular mechanisms have not been fully elucidated. Therefore, the purpose of this study was to examine potential underlying molecular mechanism(s) of repeated social defeat (RSD) stress on GC resistance in splenic macrophages. It was hypothesized that mRNA expression of receptors for GC and nuclear translocating-associated regulators in splenic macrophages would be affected by RSD, and that these changes would be associated with epigenetic modification. The data showed that the mRNA expression of GC and mineralocorticoid receptors were significantly decreased in splenic macrophages by RSD. RSD also induced a significantly decreased mRNA expression in FK506-binding protein 52 (FKBP52), consequently resulting in a significantly increased ratio of FKBP51 to FKBP52. Moreover, DNA methyltransferases 3a and 3b showed a significant decrease in their mRNA expression in the RSD group as did mRNA expression of histone deacetyltransferase 2. The RSD group also showed a significantly reduced quantity of methylated DNA in splenic macrophages. Based on microRNA (miRNA) profiling data, it was determined that RSD induced significantly increased expression of 9 different miRNAs that were predicted to interact with mRNAs of the GC receptor (6 miRNAs), mineralocorticoid receptor (3 miRNAs) and FKBP52 (2 miRNAs). Spearman correlation analysis revealed significantly strong correlations between the expression of 2 miRNAs and their target mRNA expression for GC receptors. Among these miRNAs, we verified direct effects of miRNA-29b and -340 overexpression on mRNA expression of GC receptors in L929 cells. The overexpression of miRNA-29b or -340 in L929 cells significantly reduced LPS-induced overexpression of GC receptors. In conclusion, this study provides evidence that epigenetic regulation, such as DNA methylation and miRNA expression, may play a role in the RSD-induced GC resistance that we have observed in splenic macrophages.

Expression of FK506-binding protein 52 (FKBP52) in chorionic villi with early recurrent spontaneous abortion

Objective: To investigate the mRNA and protein expression of FK506-binding protein 52 (FKBP52) in the chorionic villi of patients with recurrent spontaneous abortion (RSA) and normal women during early pregnancy.Methods: Fresh chorionic villus tissues were collected from 60 subjects. A total of 30 patients with a history of RSA were enrolled into the RSA group and 30 normal pregnant women were enrolled into the control group. The FKBP52 mRNA expression levels in chorionic villi of the RSA patients and healthy controls were measured via semiquantitative RT-PCR. The protein distribution and expression levels of FKBP52 in chorionic villi were analyzed through immunohistochemistry (IHC). The correlation between FKBP52 expression and RSA was analyzed.Results: We demonstrated that FKBP52 mRNA is expressed in chorionic villi samples of normal pregnancy and RSA. RSA patients exhibited significantly lower FKBP52 gene expression levels compared with those in normal pregnancies (p < 0.05). FKBP52 immunoreactivity in chorionic villi was mainly observed in trophoblast cell cytoplasm. The FKBP52 protein expression levels in the chorionic villi of RSA patients was significantly lower than in normal women during pregnancy (p < 0.05).Conclusions: FKBP52 protein levels were decreased in the chorionic villi of RSA patients, which indicate that the decrease in FKBP52 may be associated with RSA. The low FKBP52 mRNA expression level, which is consistent with the IHC result, may affect embryonic development and even lead to abortion. FKBP52 may be involved in the pathogenesis of RSA and new therapies that increase the FKBP52 expression may help treat RSA.

In Silico Insight into Potent of Anthocyanin Regulation of FKBP52 to Prevent Alzheimer's Disease.

Alzheimer's disease (AD) is caused by the hyperphosphorylation of Tau protein aggregation. FKBP52 (FK506 binding protein 52) has been found to inhibit Tau protein aggregation. This study found six different kinds of anthocyanins that have high binding potential. After analyzing the docking positions, hydrophobic interactions, and hydrogen bond interactions, several amino acids were identified that play important roles in protein and ligand interaction. The proteins' variation is described using eigenvectors and the distance between the amino acids during a molecular dynamics simulation (MD). This study investigates the three loops based around Glu85, Tyr113, and Lys121—all of which are important in inducing FKBP52 activation. By performing a molecular dynamic simulation process between unbound proteins and the protein complex with FK506, it was found that ligand targets that docked onto the FK1 domain will decrease the distance between Glu85/Tyr113 and Glu85/Lys121. The FKBP52 structure variation may induce FKBP52 activation and inhibit Tau protein aggregation. The results indicate that anthocyanins might change the conformation of FKBP52 during binding. In addition, the purple anthocyanins, such as cyanidin-3-glucoside and malvidin-3-glucoside, might be better than FK506 in regulating FKBP52 and treating Alzheimer's disease.

The Helix 1-3 Loop in the Glucocorticoid Receptor LBD Is a Regulatory Element for FKBP Cochaperones

The heat-shock protein 90 (Hsp90) cochaperone FK506-binding protein 52 (FKBP52) upregulates, whereas FKBP51 inhibits, hormone binding and nuclear targeting of the glucocorticoid receptor (GR). Decreased cortisol sensitivity in the guinea pig is attributed to changes within the helix 1 to helix 3 (H1-H3) loop of the guinea pig GR (gpGR) ligand-binding domain. It has been proposed that this loop serves as a contact point for FKBP52 and/or FKBP51 with receptor. We examined the role of the H1-H3 loop in GR activation by FKBP52 using a Saccharomyces cerevisiae model. The activity of rat GR (rGR) containing the gpGR H1-H3 loop substitutions was still potentiated by FKBP52, confirming the loop is not involved in primary FKBP52 interactions. Additional assays also excluded a role for other intervening loops between ligand-binding domain helices in direct interactions with FKBP52 associated with enhanced receptor activity. Complementary studies in FKBP51-deficient mouse embryo fibroblasts and HEK293 cells demonstrated that substitution of the gpGR H1-H3 loop residues into rGR dramatically increased receptor repression by FKBP51 without enhancing receptor-FKBP51 interaction and did not alter recruitment of endogenous Hsp90 and the p23 cochaperone to receptor complexes. FKBP51 suppression of the mutated rGR did not require FKBP51 peptidylprolyl cis-trans isomerase activity and was not disrupted by mutation of the FK1 proline-rich loop thought to mediate reciprocal FKBP influences on receptor activity. We conclude that the gpGR-specific mutations within the H1-H3 loop confer global changes within the GR-Hsp90 complex that favor FKBP51 repression over FKBP52 potentiation, thus identifying the loop as an important target for GR regulation by the FKBP cochaperones.

Immunophilins are Involved in the Altered Platelet Aggregation Observed in Patients with Type 2 Diabetes Mellitus

Platelet hyperaggregability might contribute to vascular complications associated with type 2 diabetes mellitus (DM2).Experimental evidence supports a direct link between altered Ca(2+) entry and hyperaggregability in DM2 patients. We aimed to investigate whether altered immunophilin expression and function are involved in the abnormal Ca(2+) entry observed in platelets from DM2 patients. Inhibition of immunophilins by tacrolimus (FK506) and sirolimus (rapamycin) reduced Ca(2+) entry in platelets from healthy donors and DM2 patients. Similarly, immunophilin inhibitors reduced platelet degranulation in both healthy and DM2 subjects. Nevertheless, α-granule secretion reduction was greater than that observed for dense granules in platelets from DM2 patients. However, no difference was observed in the inhibition of secretion in platelets from healthy subjects. Additionally, altered expression of FK506 binding protein-52 (FKBP52) and coupling to Ca(2+) channels were found in platelets from DM2 patients compared to healthy subjects. Finally, reduction in platelet function from healthy subjects and DM2 patients in the presence of immunophilin antagonists was observed, being this dysfunction more evident in platelets from DM2 patients. We suggest that, among others, FKBP52 expression and function are altered in platelets from DM2 patients, contributing to the altered Ca(2+) entry and hyperaggregability in these cells.

Fkbp52 heterozygosity alters behavioral, endocrine and neurogenetic parameters under basal and chronic stress conditions in mice

Aversive life events represent one of the main risk factors for the development of many psychiatric diseases, but the interplay between environmental factors and genetic predispositions is still poorly understood. One major finding in many depressed patients is an impaired regulation of the hypothalamic-pituitary-adrenal (HPA) axis. The negative feedback loop of the HPA axis is mediated via the glucocorticoid receptor (GR) and the mineralocorticoid receptor. The co-chaperones FK506-binding protein 51 (FKBP51) and FK506-binding protein 52 (FKBP52) are components of the heat shock protein 90-receptor-heterocomplex and are functionally divergent regulators of both receptors. Here, we characterized heterozygous Fkbp52 knockout (Fkbp52(+/-)) mice under basal or chronic social defeat stress (CSDS) conditions with regard to physiological, neuroendocrine, behavioral and mRNA expression alterations. Fkbp52(+/-) mice displayed symptoms of increased stress sensitivity in a subset of behavioral and neuroendocrine parameters. These included increased anxiety-related behavior in the elevated plus-maze and an enhanced neuroendocrine response to a forced swim test (FST), possibly mediated by reduced GR sensitivity. At the same time, Fkbp52(+/-) mice also demonstrated signs of stress resilience in other behavioral and neuroendocrine aspects, such as reduced basal corticosterone levels and more active stress-coping behavior in the FST following CSDS. These contrasting results are in line with previous reports showing that FKBP52 is not involved in all branches of GR signaling, but rather acts in a gene-specific manner to regulate GR transcriptional activation.

P.4.012 Behavioural and molecular alterations in FKBP52+/− mice under basal and chronic stress conditions

The molecular chaperone Hsp90 is abundant, ubiquitous, and catholic to biological processes in eukaryotes, controlling phosphorylation cascades, protein stability and turnover, client localization and trafficking, and ligand–receptor interactions. Not surprisingly, Hsp90 does not accomplish these activities alone. Instead, an ever-growing number of cochaperones have been identified, leading to an explosion of reports on their molecular and cellular effects on Hsp90 chaperoning of client substrates. Notable among these clients are many members of the steroid receptor family, such as glucocorticoid, androgen, estrogen and progesterone receptors. Cochaperones typically associated with the mature, hormone-competent states of these receptors include p23, the FK506-binding protein 52 (FKBP52), FKBP51, protein phosphatase 5 (PP5) and cyclophilin 40 (Cyp40). The ultimate relevance of these cochaperones to steroid receptor action depends on their physiological effects. In recent years, the first mouse genetic models of these cochaperones have been developed. This work will review the complex and intriguing phenotypes so far obtained in genetically-altered mice and compare them to the known molecular and cellular impacts of cochaperones on steroid receptors. This article is part of a Special Issue entitled: Heat Shock Protein 90 (HSP90).

Fkbp52 Regulates Androgen Receptor Transactivation Activity and Male Urethra Morphogenesis

Hypospadias is a common birth defect in humans, yet its etiology and pattern of onset are largely unknown. Recent studies have shown that male mice with targeted ablation of FK506-binding protein-52 (Fkbp52) develop hypospadias, most likely due to actions of Fkbp52 as a molecular co-chaperone of the androgen receptor (AR). Here, we further dissect the developmental and molecular mechanisms that underlie hypospadias in Fkbp52-deficient mice. Scanning electron microscopy revealed a defect in the elevation of prepucial swelling that led to the onset of the ventral penile cleft. Interestingly, expression of Fkbp52 was highest in the ventral aspect of the developing penis that undergoes fusion of the urethral epithelium. Although in situ hybridization and immunohistochemical analyses suggested that Fkbp52 mutants had a normal urethral epithelium signaling center and epithelial differentiation, a reduced apoptotic cell index at ventral epithelial cells at the site of fusion and a defect of genital mesenchymal cell migration were observed. Supplementation of gestating females with excess testosterone partially rescued the hypospadic phenotype in Fkbp52 mutant males, showing that loss of Fkbp52 desensitizes AR to hormonal activation. Direct measurement of AR activity was performed in mouse embryonic fibroblast cells treated with dihydrotestosterone or synthetic agonist R1881. Reduced AR activity at genes controlling sexual dimorphism and cell growth was found in Fkbp52-deficient mouse embryonic fibroblast cells. However, chromatin immunoprecipitation analysis revealed normal occupancy of AR at gene promoters, suggesting that Fkbp52 exerts downstream effects on the transactivation function of AR. Taken together, our data show Fkbp52 to be an important molecular regulator in the androgen-mediated pathway of urethra morphogenesis.

Uterine FK506-binding protein 52 (FKBP52)–peroxiredoxin-6 (PRDX6) signaling protects pregnancy from overt oxidative stress

Immunophilin FK506-binding protein 52 (FKBP52) is a cochaperone that binds to the progesterone receptor (PR) to optimize progesterone (P(4))-PR signaling. We recently showed that Fkbp52-deficient (Fkbp52(-/-)) mice have reduced uterine PR responsiveness and implantation failure which is rescued by excess P(4) supplementation in a genetic background-dependent manner. This finding led us to hypothesize that FKBP52 has functions in addition to optimizing PR activity. Using proteomics analysis, we found that uterine levels of peroxiredoxin-6 (PRDX6), a unique antioxidant, are significantly lower in Fkbp52(-/-) mice than in WT and PR-null (Pgr(-/-)) mice. We also found that Fkbp52(-/-) mice with reduced uterine PRDX6 levels are susceptible to paraquat-induced oxidative stress (OS), leading to implantation failure even with P(4) supplementation. The same dose of paraquat did not interfere with implantation in WT mice. Moreover, treatment with antioxidants alpha-tocopherol and N-acetylcysteine (NAC) attenuated paraquat-induced implantation failure in P(4)-treated Fkbp52(-/-) mice. Functional analyses using mouse embryonic fibroblasts show that Fkbp52 deficiency associated with reduced PRDX6 levels promotes H(2)O(2)-induced cell death, which is reversed by the addition of NAC or by forced expression of PRDX6, suggesting that Fkbp52 deficiency diminishes the threshold against OS by reducing PRDX6 levels. These findings provide evidence that heightened uterine OS in Fkbp52(-/-) females with reduced PRDX6 levels induces implantation failure even in the presence of excess P(4). This study shows that FKBP52-PRDX6 signaling protects pregnancy from overt OS.

Susceptibility to Diet-Induced Hepatic Steatosis and Glucocorticoid Resistance in FK506-Binding Protein 52-Deficient Mice

Although FK506-binding protein 52 (FKBP52) is an established positive regulator of glucocorticoid receptor (GR) activity, an in vivo role for FKBP52 in glucocorticoid control of metabolism has not been reported. To address this question, FKBP52(+/-) mice were placed on a high-fat (HF) diet known to induce obesity, hepatic steatosis, and insulin resistance. Tissue profiling of wild-type mice showed high levels of FKBP52 in the liver but little to no expression in muscle or adipose tissue, predicting a restricted pattern of FKBP52 effects on metabolism. In response to HF, FKBP52(+/-) mice demonstrated a susceptibility to hyperglycemia and hyperinsulinemia that correlated with reduced insulin clearance and reduced expression of hepatic CEACAM1 (carcinoembryonic antigen-related cell adhesion molecule 1), a mediator of clearance. Livers of HF-fed mutant mice had high lipid content and elevated expression of lipogenic genes (peroxisome proliferator-activated receptor gamma, fatty acid synthase, and sterol regulatory element-binding protein 1c) and inflammatory markers (TNFalpha). Interestingly, mutant mice under HF showed elevated serum corticosterone, but their steatotic livers had reduced expression of gluconeogenic genes (phosphoenolpyruvate carboxy kinase, glucose 6 phosphatase, and pyruvate dehydrogenase kinase 4), whereas muscle and adipose expressed normal to elevated levels of glucocorticoid markers. These data suggest a state of glucocorticoid resistance arising from liver-specific loss of GR activity. Consistent with this hypothesis, reduced expression of gluconeogenic genes and CEACAM1 was observed in dexamethasone-treated FKBP52-deficient mouse embryonic fibroblast cells. We propose a model in which FKBP52 loss reduces GR control of gluconeogenesis, predisposing the liver to steatosis under HF-diet conditions attributable to a shunting of metabolism from glucose production to lipogenesis.