Interaction between microRNAs and actin-associated protein Arpc5 regulates translational suppression during male germ cell differentiation.

Decoupling of transcription and translation during postmeiotic germ cell differentiation is critical for successful spermatogenesis. Here we establish that the interaction between microRNAs and actin-associated protein Arpc5 sets the stage for an elaborate translational control mechanism by facilitating the sequestration of germ cell mRNAs into translationally inert ribonucleoprotein particles until they are later translated. Our studies reveal that loss of microRNA-dependent regulation of Arpc5, which controls the distribution of germ cell mRNAs between translationally active and inactive pools, results in abnormal round spermatid differentiation and impaired fertility. Interestingly, Arpc5 functions as a broadly acting translational suppressor, as it inhibits translation initiation by blocking 80S formation and facilitates the transport of mRNAs to chromatoid/P bodies. These findings identify a unique role for actin-associated proteins in translational regulation, and suggest that mRNA-specific and general translational control mechanisms work in tandem to regulate critical germ cell differentiation events and diverse somatic cell functions.

Isolation of Sertoli, Leydig, and spermatogenic cells from the mouse testis.

A thorough understanding of the events during mammalian spermatogenesis requires studying specific molecular signatures of individual testicular cell populations as well as their interaction in co-cultures. However, most purification techniques to isolate specific testicular cell populations are time-consuming, require large numbers of animals, and/or are only able to isolate a few cell types. Here we describe a cost-effective and timesaving approach that uses a single protocol to enrich multiple testicular cell populations (Sertoli, Leydig, and several spermatogenic cell populations) from as few as one mouse. Our protocol combines rigorous enzymatic digestion of seminiferous tubules with counter-current centrifugal elutriation, yielding specific testicular cell populations with >80%-95% purity.

Role of ß-catenin in post-meiotic male germ cell differentiation.

Though roles of ß-catenin signaling during testis development have been well established, relatively little is known about its role in postnatal testicular physiology. Even less is known about its role in post-meiotic germ cell development and differentiation. Here, we report that ß-catenin is highly expressed in post-meiotic germ cells and plays an important role during spermiogenesis in mice. Spermatid-specific deletion of ß-catenin resulted in significantly reduced sperm count, increased germ cell apoptosis and impaired fertility. In addition, ultrastructural studies show that the loss of ß-catenin in post-meiotic germ cells led to acrosomal defects, anomalous release of immature spermatids and disruption of adherens junctions between Sertoli cells and elongating spermatids (apical ectoplasmic specialization; ES). These defects are likely due to altered expression of several genes reportedly involved in Sertoli cell-germ cell adhesion and germ cell differentiation, as revealed by gene expression analysis. Taken together, our results suggest that ß-catenin is an important molecular link that integrates Sertoli cell-germ cell adhesion with the signaling events essential for post-meiotic germ cell development and maturation. Since ß-catenin is also highly expressed in the Sertoli cells, we propose that binding of germ cell ß-catenin complex to ß-catenin complex on Sertoli cell at the apical ES surface triggers a signaling cascade that regulates post-meiotic germ cell differentiation.