Potential Role Of FKBP52in The Balance Between Estrogen Receptor (ER) And Androgen Receptor (AR)

[Objectives] Steroid hormones are essential to metabolism, inflammation, immune functions, and development of sexual characteristics. They can regulate gene expression by binding specific steroid receptor as the intracellular signaling transduction and transcription factor. Both estrogen receptor (ER) and androgen receptor (AR) belong to the steroid receptor superfamily. There is emerging evidence that the balance between ERa and AR signaling is a critical determinant of development and growth in the breast.FKBP52is a TPR-containing immunophilin with peptidyl-prolyl cis-trans isomerase (PPIase) activity. It is a cochaperone which can directly bind Hsp90via its respective tetratricopeptide repeat (TPR) domains, entering into steroid hormone receptor (SR) complexes at the final stage of assembly. Numerous studies uniformly point to FKBP52as a positive regulator of SR transcriptional activity, such as androgen receptor (AR) and progesterone receptor (PR). It is known that full AR activity does require the PPIase function of FKBP52. And there is less report on the balance between ER and AR effected by FKBP52, so we studied the role of FKBP52in it.[Methods] In order to study the FKBP52function in regulation of the balance between ER and AR, we analyzed the development of mammary gland in male FKBP52KO mouse.1) Phenotype analysis of mammary gland. To assess the structure of mammary gland, we prepared HE staining and wholemounts of mammary gland from10days、24days and2months in male FKBP52KO and littermate WT female and male mice.2) Estrogen. The development and growth of mammary gland in KO male mice are induced by estrogen. In order to verify the effects of estrogen on mammary gland development, we injected0.05mg estrogen daily into FKBP52KO and littermate WT female and male mice from24days and2months, respectively. After14days, we evaluated the development of mammary gland using carmine alum staining.3) RNA sequencing. RNA of nipple tissue in FKBP52KO and female WT mice and skin tissue in WT male mouse were extracted. The skin tissue at the same position in WT male mouse was taken as control. All RNA samples were sequenced b the next generation sequencing approach. From the gene expression profiling, several important genes which is critical in mammary gland development were identified in KO male mice.4) Real-time PCR. This result can show the expression of ER, AR and some related genes in mammary gland.5) Immunohistochemistry. It can further validate the location and expression of ER, AR and other genes.6) Luciferase assay. Expression plasmid containing ERE (estrogen response element) promoter and luciferase reporter gene was transfected into FK.BP52WT and KO cells. We can measure ER activity through luciferase expression.[Results]1) There is phenomenon of mammary gland development in male FKBP52KO mice.2) The structure of nipple and mammary gland is very similar in male FKBP52KO and female WT mice.3) Mammary glands in male FKBP52KO mice can initiate development normally at embryo stage, but they stop development at puberty.4) Estrogen can induce the development of mammary gland at puberty in male FKBP52KO mice.5) Some related genes will be up or down regulated lacking FKBP52.6) The location and expression of ERaare similar to FOXA1and CITED1in male FKBP52KO mice.7) There is enhancement of ERaactivity lacking FKBP52.[Conclusions]1) Development of mammary gland in FKBP52KO male mice was due to enhancement of ER activity.2) FKBP52play an important role in keeping the balance between ER and AR.