Regulation of steroid hormone receptor function by the 52-kDa FK506-binding protein (FKBP52)

The large FK506-binding protein FKBP52 has been characterized as an important positive regulator of androgen, glucocorticoid and progesterone receptor signaling pathways. FKBP52 associates with receptor-Hsp90 complexes and is proposed to have roles in both receptor hormone binding and receptor subcellular localization. Data from biochemical and cellular studies have been corroborated in whole animal models as fkbp52-deficient male and female mice display characteristics of androgen, glucocorticoid and/or progesterone insensitivity. FKBP52 receptor specificity and the specific phenotypes displayed by the fkbp52-deficient mice have firmly established FKBP52 as a promising target for the treatment of a variety of hormone-dependent diseases. Recent studies demonstrated that the FKBP52 FK1 domain and the proline-rich loop within this domain are functionally important for FKBP52 regulation of receptor function. Based on these data, efforts are currently underway to target the FKBP52 FK1 domain and the proline-rich loop with small molecule inhibitors.     

FKBP51 regulation of AKT/protein kinase B phosphorylation

FK506 binding protein 51 (FKBP51, also called FKBP5) belongs to a family of immunophilins, FK506 binding proteins (FKBPs). FKBP family members are targets for drugs such as rapamycin. Although FKBP51 shares characteristics with other FKBPs, it also has unique features, especially the role in its regulation of important signaling pathways such as the AKT kinase/protein kinase B pathway. In this review, we will focus on the function of FKBP51 as a scaffolding protein in the regulation of AKT activation and, in turn, its role in tumorigenesis and response to chemotherapy.     

Steroid up-regulation of FKBP51 and its role in hormone signaling

FK506 binding protein 51 (FKBP51, FKBP5) functions as a co-chaperone for androgen, glucocorticoid, mineralocorticoid and progesterone receptors. The FKBP51 can act as an important determinant of the responses to steroids, especially to glucocorticoids in stress and mood disorders and androgens in prostate cancer, raising medical and pharmacological interests in the protein and its gene. Recent studies have revealed the molecular mechanisms by which the androgens and the glucocorticoids via their nuclear receptors elicit the robust up-regulation of the FKBP51 gene. Several polymorphisms in the FKBP51 gene have been associated with the mood disorders and differences in glucocorticoid sensitivity. The polymorphisms may contribute to the steroid up-regulation of the FKBP51 and thus influence the regulatory loops in steroid signaling.     

Gene–Stress–Epigenetic Regulation of FKBP5: Clinical and Translational Implications

Stress responses and related outcomes vary markedly across individuals. Elucidating the molecular underpinnings of this variability is of great relevance for developing individualized prevention strategies and treatments for stress-related disorders. An important modulator of stress responses is the FK506-binding protein 51 (FKBP5/FKBP51). FKBP5 acts as a co-chaperone that modulates not only glucocorticoid receptor activity in response to stressors but also a multitude of other cellular processes in both the brain and periphery. Notably, the FKBP5 gene is regulated via complex interactions among environmental stressors, FKBP5 genetic variants, and epigenetic modifications of glucocorticoid-responsive genomic sites. These interactions can result in FKBP5 disinhibition that has been shown to contribute to a number of aberrant phenotypes in both rodents and humans. Consequently, FKBP5 blockade may hold promise as treatment intervention for stress-related disorders, and recently developed selective FKBP5 blockers show encouraging results in vitro and in rodent models. Although risk for stress-related disorders is conferred by multiple environmental and genetic factors, the findings related to FKBP5 illustrate how a deeper understanding of the molecular and systemic mechanisms underlying specific gene–environment interactions may provide insights into the pathogenesis of stress-related disorders.     

FKBP family proteins as promising new biomarkers for cancer

FK506-binding proteins (FKBPs) belong to the immunophilin family and bind immunosuppressive drugs, such as FK506 and rapamycin. These proteins, through interactions with steroid hormone receptors, kinases, or other cellular factors, play important roles in various physiological processes and, more interestingly, in pathological processes in mammals. Accumulating evidence has implicated some FKBP members in a variety of processes, such as the cell cycle and survival and apoptotic signaling pathways, particularly in cancers. After the deregulation of their expression was observed in cancer tissues, it became increasingly clear that FKBP members played an important role in tumorigenesis and the response to chemotherapies and radiotherapies and that FKBP members could act as oncogenes or tumor suppressors depending on the tissue type. A wealth of data from in vitro and clinical studies is paving the way for novel, promising roles of FKBPs as diagnostic, prognostic or therapy-monitoring cancer biomarkers.     

Differences in FKBP51 Regulation Following Chronic Social Defeat Stress Correlate with Individual Stress Sensitivity: Influence of Paroxetine Treatment

Various clinical studies have identified FK506-binding protein 51 (FKBP51) as a target gene involved in the development of psychiatric disorders such as depression. Furthermore, FKBP51 has been shown to affect glucocorticoid receptor signaling by sensitivity modulation and it is implicated in stress reactivity as well as in molecular mechanisms of stress vulnerability and resilience. We investigated the physiological, behavioral, and neuroendocrine parameters in an established chronic stress model both directly after stress and after a recovery period of 3 weeks and also studied the efficacy of paroxetine in this model. We then examined FKBP51 mRNA levels in the dorsal and ventral part of the hippocampus and correlated the expression to behavioral and endocrine parameters. We show robust chronic stress effects in physiological, behavioral, and neuroendocrine parameters, which were only slightly affected by paroxetine treatment. On the contrary, paroxetine led to a disruption of the neuroendocrine system. FKBP51 expression was significantly increased directly after the stress period and correlated with behavioral and neuroendocrine parameters. Taken together, we were able to further elucidate the role of FKBP51 in the mechanisms of stress resilience and vulnerability, especially with respect to behavioral and neuroendocrine parameters. These findings strongly support the concept of FKBP51 as a marker for glucocorticoid receptor sensitivity and its involvement in the development of psychiatric disorders.     

The involvement of FK506-binding protein 51 (FKBP5) in the behavioral and neuroendocrine effects of chronic social defeat stress

Chronic stress is increasingly considered to be a main risk factor for the development of a variety of psychiatric diseases such as depression. This is further supported by an impaired negative feedback of the hypothalamic-pituitary-adrenal (HPA) axis, which has been observed in the majority of depressed patients. The effects of glucocorticoids, the main hormonal endpoint of the HPA axis, are mediated via the glucocorticoid receptor (GR) and the mineralocorticoid receptor. The FK506-binding protein 51 (FKBP5), a co-chaperone of the Hsp90 and component of the chaperone-receptor heterocomplex, has been shown to reduce ligand sensitivity of the GR. This study aimed to investigate the function of FKBP5 as a possible mediator of the stress response system and its potential role in the development of stress-related diseases. Therefore, we assessed whether mice lacking the gene encoding FKBP5 (51KO mice) were less vulnerable to the adverse effects of three weeks of chronic social defeat stress. Mice were subsequently analyzed with regards to physiological, neuroendocrine, behavioral and mRNA expression alterations. Our results show a less vulnerable phenotype of 51KO mice with respect to physiological and neuroendocrine parameters compared to wild-type animals. 51KO mice demonstrated lower adrenal weights and basal corticosterone levels, a diminished response to a novel acute stimulus and an enhanced recovery, as well as more active stress-coping behavior. These results suggest an enhanced negative glucocorticoid feedback within the HPA axis of 51KO mice, possibly modulated by an increased sensitivity of the GR. This article is part of a Special Issue entitled 'Anxiety and Depression'.     

Exploration of pipecolate sulfonamides as binders of the FK506-binding proteins 51 and 52

FK506-binding proteins (FKBP) 51 and 52 are cochaperones that modulate the signal transduction of steroid hormone receptors. Single nucleotide polymorphisms in the gene encoding FKBP51 have been associated with a variety of psychiatric disorders. Rapamycin and FK506 are two macrocyclic natural products, which tightly bind to most FKBP family members, including FKBP51 and FKBP52. A bioisosteric replacement of the α-ketoamide moiety of rapamycin and FK506 with a sulfonamide was envisaged with the retention of the conserved hydrogen bonds. A focused solid support-based synthesis protocol was developed, which led to ligands with submicromolar affinity for FKBP51 and FKBP52. The molecular binding mode for one sulfonamide analogue was confirmed by X-ray crystallography.     

Immunohistochemical analysis of FKBP51 in human cancers

FKBP51 is a FK506-binding immunophilin involved in the regulation of several fundamental biological processes. A growing body of data indicates that this protein has also a role in the abnormal cell growth of cancers, and could be considered as a promising new marker of tumor progression and response to radio/chemotherapy. However, the data concerning the expression of FKBP51 in cancer are not conclusive, and partially contradictory. They delineate a very complex scenario, in which many molecular FKBP51-related pathways are variously intersected among different tumors. This review reports the available data concerning FKBP51 expression in normal tissues and human malignancies, outlining the role of the immunohistochemical analysis as a fundamental tool for better understanding the role of this immunophilin in cancer biology.     

Evaluation of synthetic FK506 analogues as ligands for the FK506-binding proteins 51 and 52

The FK506-binding proteins (FKBP) 51 and 52 are cochaperones that modulate the signal transduction of steroid hormone receptors. Both proteins have been implicated in prostate cancer. Furthermore, single nucleotide polymorphisms in the gene encoding FKBP51 have been associated with a variety of psychiatric disorders. Rapamycin and FK506 are two macrocyclic natural products that bind to these proteins indiscriminately but with nanomolar affinity. We here report the cocrystal structure of FKBP51 with a simplified α-ketoamide analogue derived from FK506 and the first structure-activity relationship analysis for FKBP51 and FKBP52 based on this compound. In particular, the tert-pentyl group of this ligand was systematically replaced by a cyclohexyl ring system, which more closely resembles the pyranose ring in the high-affinity ligands rapamycin and FK506. The interaction with FKBPs was found to be surprisingly tolerant to the stereochemistry of the attached cyclohexyl substituents. The molecular basis for this tolerance was elucidated by X-ray cocrystallography.     

脑缺血再灌注损伤中 FK506结合蛋白51对半胱氨酸蛋白酶－3及大鼠海马 CA1区神经元坏死的影响

目的：探讨脑缺血再灌注损伤中FK506结合蛋白51（FKBP51）对半胱氨酸蛋白酶－3（Caspase－3）及大鼠海马CA1区神经元坏死的影响。方法将清洁级SD大鼠随机分成假手术组（ Sham组）、缺血再灌注组（ I／R组）、溶剂对照组（ TE组）、FKBP51反义寡核苷酸组（ FKBP51 ASODN组）和FKBP51错义寡核苷酸组（ FKBP51 MSODN组），采用四血管阻塞模型大鼠全脑缺血模型造模。应用免疫印迹法测定FKBP51蛋白表达、FKBP51 ASODN对FKBP51蛋白水平的表达和Caspase－3活性的影响；采用苏木精－伊红溶液染色法测定FKBP51 ASODN对大鼠海马CA1区神经元坏死的影响。结果（1）在Sham组和I／R组（缺血再灌注0 min、15 min、30 min、1 h、3 h、6 h、1 d、3 d）中，FKBP51蛋白均有表达，并且表达量间的差异无统计学意义（F＝0．64，P＞0．05）。（2）FKBP51 ASODN组的FKBP51蛋白表达量明显减少，与Sham组差异有统计学意义（t＝8．21，P＜0．05）。（3）Sham组活化Caspase－3的表达量明显低于其他各组，差异有统计学意义（F＝12．31，P＜0．05）；与FKBP51 MSODN组比较，FKBP51 ASODN组的表达量相对较低，差异有统计学意义（ t＝9．71，P＜0．05）。（4）Sham组海马CA1区锥体细胞数量多[（186．3±2．5）个]并且排列密集，核大且圆，核仁明显；I／R 5 d组[（15．4±2．6）个]、TE组[（18．5±2．2）个]和FKBP51 MSODN组[（17．5±1．8）个]海马CA1区锥体细胞几乎完全消失，仅残留少数细胞，大量变性锥体细胞核固缩深染，胞浆嗜伊红，胞膜破裂，胞内容物释放， Sham组与其余各组之间的差异有统计学意义（χ^2＝81．91，P＜0．05）；FKBP51 ASODN组[（92．8±2．6）个]海马CA1区锥体细胞存活数增加，与其余各组的差异有统计学意义（χ^2＝52．36，P＜0．05）。结论在脑缺血再灌注损伤中，FKBP51可以增强活化的Caspase－3的表达并抑制神经元的存?     

Reversal of isoproterenol-induced downregulation of phospholamban and FKBP12.6 by CPU0213-mediated antagonism of endothelin receptors

Aim： The downregulation of phospholamban （PLB） and FKBP12.6 as a result of β- receptor activation is involved in the pathway（s） of congestive heart failure. We hypothesized that the endothelin （ET）-1 system may link to downregulated PLB and FKBP12.6. Methods： Rats were subjected to ischemia/reperfusion （I/R） to cause heart failure （HF）. 1 mg/kg isoproterenol （ISO） was injected subcutaneously （sc） for 10 d to worsen HF. 30 mg/kg CPU0213 （sc）, a dual ET receptor （ETAR/ETBR） antagonist was given from d 6 to d 10. On d 11, cardiac function was assessed together with the determination of mRNA levels of ryanodine receptor 2, calstabin-2 （FKBP12.6）, PLB, and sarcoplasmic reticulum Ca^2＋-ATPase. Isolated adult rat ventricular myocytes were incubated with ISO at 1 × 10^-6 mol/L to set up an in vitro model of HF. Propranolol （PRO）, CPU0213, and darusentan （DAR, an ETAR antagonist） were incubated with cardiomyocytes at 1× 10^-5 mol/L or 1× 10^-6 mol/L in the presence of ISO （1×10^-6 mol/L）. Immunocytochemistry and Western blotting were applied for measuring the protein levels of PLB and FKBP12.6. Results： The worsened hemodynamics produced by I/R were exacerbated by ISO pretreatment. The significant downregulation of the gene expression of PLB and FKBP12.6 and worsened cardiac function by ISO were reversed by CPU0213. In vitro ISO 1× 10^-6 mol/L produced a sharp decline of PLB and FKBP12.6 proteins relative to the control. The downregulation of the protein expression was significantly reversed by the ET receptor antagonist CPU0213 or DAR, comparable to that achieved by PRO. Conclusion： This study demonstrates a role of ET in mediating the downregulation of the cardiac Ca^2＋-handling protein by ISO.     

Design and synthesis of novel FKBP inhibitors.

Small molecule FKBP inhibitors were prepared with inhibitory activity ranging from micromolar to nanomolar. The design of these inhibitors derives from a structural analysis of the substrates for FKBP and cyclophilin. As a consequence of this analysis two key observations were made, namely: (1) amino ketone moieties are suitable as FKBP recognition elements at the P1-P1' site and (2) the P3'-P4' site will accept a trans-olefin as a suitable mimetic of a peptide moiety. The preparation of these non-peptide inhibitors is readily accomplished by a protocol which includes the synthesis of chiral propargylic amines and their subsequent conversion into vinyl zirconium reagents.     

FKBP52在重组腺相关病毒载体胞内转运中的作用研究

目的探讨细胞质中FKBP52对重组腺相关病毒(AAV)介导的基因转导效率的作用。方法测定AAV在FKBP52-野生型(WT)、FKBP52-杂合型(HE)及FKBP52-基因敲除(KO)鼠胚纤维母细胞(MEFs)中的基因转导效率,观察羟基脲(HU)对不同基因型MEFs中AAV转导效率的影响。采用流式细胞术、EMSA、Southernblot、免疫沉淀、West-ernblot等方法分析FKBP52对AAV在细胞内转运的作用。结果传统的单链AAV不能有效地转导FKBP52-WTMEFs,在FKBP52-HE及FKBP52-KOMEFs中的转导效率也未见明显增加。在这些细胞中,AAV不能有效地转运至细胞核。HU处理后能使AAV在WTMEFs中的转导效率提高25倍,但是在KOMEFs中AAV转导效率仅提高4倍。为避免AAV第二链合成的影响,进一步采用自身互补的AAV(scAAV)载体进行研究,结果发现,HU能使WTMEFsscAAV基因转导效率增强23倍,但在KOMEFs中转导效率仅增加4倍。提示,HU处理后KOMEFs中转导效率未增加的原因并非由于AAV第二链的合成障碍所致。HU处理后,59%的AAV基因组DNA出现在WTMEFs细胞核,KOMEFs细胞核中为28%,提示HU介导的AAV转运通路在KOMEFs中存在缺陷。转染FKBP52表达质粒的KOMEFs经HU处理后,AAV所介导的基因转导效率恢复至WTMEFs的水平,且直接与其转运至细胞核的转运功能改善相吻合。结论作为一种细胞分子伴侣蛋白,FKBP52能促进AAV在细胞内转运,有益于重组AAV载体在人类基因治疗中的应用。     

FKBP5 Genotype and Structural Integrity of the Posterior Cingulum

Alterations in the microarchitecture of the posterior cingulum (PC), a white matter tract proximal to the hippocampus that facilitates communication between the entorhinal and cingulate cortices, have been observed in individuals with psychiatric disorders, such as depression and post-traumatic stress disorder (PTSD). PC decrements may be a heritable source of vulnerability for the development of affective disorders; however, genetic substrates for these white matter abnormalities have not been identified. The FKBP5 gene product modulates glucocorticoid receptor function and has been previously associated with differential hippocampal structure, function, and affect disorder risk. Thus, FKBP5 is an attractive genetic target for investigations of PC integrity. We examined associations between PC integrity, measured through diffusion tensor imaging (DTI) and fractional anisotropy (FA; an index of white matter integrity), and polymorphisms in the FKBP5 SNP rs1360780 in a sample of 82 traumatized female civilians. Findings indicated that, compared with individuals without this allele, individuals who carried two ‘risk'' alleles for this FKBP5 SNP (T allele; previously associated with mood and anxiety disorder risk) demonstrated significantly lower FA in the left PC, even after statistically controlling for variance associated with age, trauma exposure, and PTSD symptoms. These data suggest that specific allelic variants for an FKBP5 polymorphism are associated with decrements in the left PC microarchitecture. These white matter abnormalities may be a heritable biological marker that indicates increased vulnerability for the development of psychiatric disorders, such as PTSD.     

FKBP5 Genotype-Dependent DNA Methylation and mRNA Regulation After Psychosocial Stress in Remitted Depression and Healthy Controls

Background:

Polymorphisms in the FK506 binding protein 5 (FKBP5) gene have been shown to influence glucocorticoid receptor sensitivity, stress response regulation, and depression risk in traumatized subjects, with most consistent findings reported for the functional variant rs1360780. In the present study, we investigated whether the FKBP5 polymorphism rs1360780 and lifetime history of major depression are associated with DNA methylation and FKBP5 gene expression after psychosocial stress.

Methods:

A total of 116 individuals with a positive (n = 61) and negative (n = 55) lifetime history of major depression participated in the Trier Social Stress Test. We assessed plasma cortisol concentrations, FKBP5 mRNA expression, and CpG methylation of FKBP5 intron 7 in peripheral blood cells.

Results:

Genotype-dependent plasma cortisol response to psychosocial stress exposure was observed in healthy controls, with the highest and longest-lasting cortisol increase in subjects with the TT genotype of the FKBP5 polymorphism rs1360780, and healthy controls carrying the T risk allele responded with a blunted FKBP5 mRNA expression after psychosocial stress. No genotype effects could be found in remitted depression.

Conclusions:

The FKBP5 rs1360780 polymorphism is associated with plasma cortisol and FKBP5 mRNA expression after psychosocial stress in healthy controls but not in remitted depression. Preliminary results of the DNA methylation analysis suggest that epigenetic modifications could be involved.