Expression And Functional Analysis Of MTA2in Testicular Sertoli Cells

Sertoli cells (SCs), known as the only somatic cells possessing close structuralrelationship with germ cells inside the seminiferous tubules, have drawn much attention ofreproductive researchers because of their pivotal roles in the regulation of spermatogenicproliferation and differentiation. Accumulated evidences have demonstrated thatdeacetylation serves as a key modulator of the function of the genes predominantlyexpressed in SCs. In the current study, we demonstrated for the first time that Metastasisassociated protein2(MTA2), an important recruiter of histone deacetylase, wasexclusively expressed in testicular Sertoli cells (SCs). This expression wasdevelopmentally regulated and was stage-specific, with the maximal expression level atstage8to stage12, suggesting that this chromatin modifier may be involved in the finalrelease of mature sperm within seminiferous tubules. Adult male rats received a singleEDS injection (75mg EDS in DMSO/kg body) which resulted in the complete eliminationof adult Leydig cells (LCs) and subsequent decline of circulated testosterone (T) level atpostoperative7and14day. In line with the hormonal change, the relative expression of ratMTA2mRNA began to decrease1d after EDS treatment, with the minimal level observed at d7and14post-EDS treatment. Meanwhile, the androgen-regulated expression ofMTA2was also confirmed in cultured TM4cell line and Co-IP assay demonstrated adirect interaction between endogenous MTA2and Ar in TM4cells upon T stimulation.These results suggest that androgen signaling may serve as the upstream modulator ofMTA2expression in SCs. Moreover, in vitro GST pull-down assay performed usingGST-fusion full-length and truncated Ar proteins and His-tagged MTA2protein clearlyshowed that the interaction between Ar and MTA2upon T treatment was enhanced only inthe presence of the ligand binding domain (LBD). Treatment of SC with PP2, abroad-spectrum inhibitor of Src, or PD98059, a well-characterized inhibitor of MAPkinase activity could both abolish the elevated expression of MTA2induced by Ttreatment, suggesting that T-dependent activation of MTA2requires activated Src andMAP kinases. Taken together, the available data indicated that the androgen regulation ofMTA2expression may be achieved though the non-classical testosterone actions.Interestingly, the expression level of MTA2in SCs was also positively regulated by FSH,which is mediated via Ar signaling. The direct action of FSH on SC is transient. Therefore,continuous stimulation of SC with FSH leads to a desensitization of the cells to FSH.Desensitization of the FSH response in SC involves multiple steps in signal transduction,among which the down-regulation of the transcription of the FSHR gene is of greatimportance. We have demonstrated that MTA2represses FSH-mediated FSHRtranscription by recruiting HDAC1. Thus, down-regulation of FSHR by deacetylationthrough MTA2-HDAC1cascade operates as an indispensible self-regulating mechanismfor the continuity of FSH response in SCs. In addition, assessment of SC-related MTA2expression in human pathological testis by immunohistochemistry demonstrated aclear-cut attenuated level of this histone modifier in the SC of patients with impaired orarrested spermatogenesis, indicating that MTA2expression in SC may be associated withhuman spermatogenic failure. In summary, we propose a novel role of SC-expressingMTA2. GnRH signal-induced FSH binds to FSHR and subsequently activates a series ofdownstream signaling to facilitate normal spermatogenic differentiation. Simultaneously,FSH induces Ar activation, followed by an upregulation of MTA2expression via the non-classical androgen signaling. HDAC complexes associated with the MTA2corepressor bind to the FSHR gene and mediate its transcriptional repression in responseto FSH. Conversely, deregulated MTA2expression may result in the disruption of thehomologous down-regulation of FSHR gene and the inevitable reduction of competencyof FSHR to bind FSH, followed by an unusual elevation of serum FSH level. Overall,Ar/MTA2/HDAC1cascade may serve as an important negative feedback to modulate thetiming and magnitude of subsequent signal transduction of SC in response to FSH. Oursystematic study would be helpful to elucidate the function and mechanism ofSCs-specific expression of MTA2during spermatogenesis, which will certainly shed novellight on the underlying mechanism by which SCs modulate spermatogenesis via MTAsignaling pathway.