Abstract
BackgroundSpermatogenesis is a complex process involving the self-renewal and differentiation of spermatogonia into mature spermatids in the seminiferous tubules. During spermatogenesis, germ cells migrate from the basement membrane to cross the blood-testis barrier (BTB) and finally reach the luminal side of the seminiferous epithelium. However, the mechanism for regulating the migration of germ cells remains unclear. In this study, we focused on the expression and function of transcriptional factor EB (TFEB), a master regulator of lysosomal biogenesis, autophagy and endocytosis, in spermatogenesis.MethodsThe expression pattern of the TFEB in mouse testes were investigated by Western blotting and immunohistochemistry analyses. Either undifferentiated spermatogonia or differentiating spermatogonia were isolated from testes using magnetic-activated cell sorting based on specific cell surface markers. Differentiation of spermatogonia was induced with 100 nM retinoic acid (RA). shRNA was used to knock down TFEB in cells. TFEB expression was detected by immunofluorescence, qRT-PCR, and Western blotting. Cell migration was determined by both transwell migration assay and wound healing assay applied to a cell line of immortalized spermatogonia, GC-1 cells.ResultsDuring testicular development, TFEB expression was rapidly increased in the testes at the period of 7 days post-partum (dpp) to 14 dpp, whereas in adult testis, it was predominantly localized in the nucleus of spermatogonia at stages VI to VIII of the seminiferous epithelial cycle. Accordingly, TFEB was observed to be mainly expressed in differentiating spermatogonia and was activated for nuclear translocation by RA treatment. Moreover, knockdown of TFEB expression by RNAi did not affect spermatogonial differentiation, but significantly reduced cell migration in GC-1 cells.ConclusionThese findings imply that regionally distinct expression and activation of TFEB was strongly associated with RA signaling, and therefore may promote cell migration across the BTB and transport along the seminiferous epithelium.
Highlights
Spermatogenesis is a complex process involving the self-renewal and differentiation of spermatogonia into mature spermatids in the seminiferous tubules
Consistent with this finding, the mRNA levels of transcriptional factor EB (TFEB), measured by qRT-PCR, in the developing testis increased during testis development, with a pronounced increase in expression during 7 dpp to 14 dpp (Fig. 1b), which corresponds to the spermatogonial differentiation and the first instances of meiosis in neonatal testes
According to the morphological analysis of the seminiferous tubules in adult mice, the immunohistochemistry staining of TFEB in spermatogonia or spermatocytes was mainly present in the seminiferous tubules at stage VI to VIII
Summary
Spermatogenesis is a complex process involving the self-renewal and differentiation of spermatogonia into mature spermatids in the seminiferous tubules. Germ cells migrate from the basement membrane to cross the blood-testis barrier (BTB) and reach the luminal side of the seminiferous epithelium. Liu et al Reproductive Biology and Endocrinology (2018) 16:105 regulated and precisely coordinated, perturbs spermatogenesis and may lead to male infertility In coordination with these cell divisions, the various types of germ cells migrate from the basement membrane to the adluminal side of the seminiferous tubules. One of the key migration steps is when the preleptotene spermatocytes (preL), differentiated from type B spermatogonia, migrate across the blood-testis barrier (BTB), at stage VIII of the seminiferous epithelial cycle [4, 5]. The capacity of RA to control spermatogonial differentiation, meiotic initiation as well as maintenance of the cycle of the seminiferous epithelium has been well studied over the last couple of years, whereas the relationship between RA and cell migration associated with meiotic initiation is less well understood
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