USP8/UBPy-regulated sorting and the development of sperm acrosome: the recruitment of MET

Abstract

The acrosome is a peculiar vacuole that at fertilization undergoes the acrosome reaction (AR), an event unique in the sperm life. Contents released promote sperm penetration through oocyte's investments; membranous components are involved in sperm-egg interaction/fusion. Therefore, both constituents play a role in fertilization. The biogenesis of this vacuole, however, has not been clarified yet; recently, it has been proposed as a novel lysosome-related organelle (LRO). Our research focuses on the involvement of the endosomal pathway in acrosomogenesis starting from the early phases. The trafficking sorted by USP8/UBPy, an endosomal regulator recently described as a compelling candidate for male fertility gene, was investigated in comparison to that of SP56, a marker of the biosynthetic pathway. Mouse spermatids were double/triple immunolabeled and examined by confocal microscopy. The contribution of the vesicular traffic assisted by the cortical microtubule array was also evaluated in nocodazole-treated spermatids. USP8/UBPy-sorted cargo contributes early to acrosomogenesis and its trafficking is microtubule mediated. It was identified, through co-immunoprecipitation/co-immunolocalization assays, that the membrane receptor MET, described herein for the first time in spermatids, as an USP8/UBPy-target substrate is delivered to the acrosome. MET and USP8/UBPy still colocalize in epididymal spermatozoa. Following the AR, MET and USP8/UBPy show a distinct fate. MET, in particular, translocates at the PAS, the post acrosomal segment known to harbor sperm-borne factors involved in oocyte activation. Overall, our results support the concept of the acrosome as a LRO and provide evidence for the identification of MET as a tyrosine kinase receptor that may play a role in fertilization.

PTBP2-dependent alternative splicing regulates protein transport and mitochondria morphology in post-meiotic germ cells.

“…To do so, we performed an ultrastructural analysis of WT and Ptbp2 cKO testes using transmission electron microscopy (TEM), with an emphasis on round spermatids (the most advanced germ cell stage present in Ptbp2 cKO mice). In particular, we focused on acrosome formation, which is heavily dependent on the trafficking and fusion of vesicles derived from the Golgi apparatus and the endosomal-sorting pathway (Berruti and Paiardi, 2015;Berruti, 2016). Indeed, our TEM analysis indicates these processes are perturbed in the absence of PTBP2.…”

Mouse spermatogenesis‐associated protein 1 (SPATA1), an IFT20 binding partner, is an acrosomal protein

“…However, we do not have evidence showing that SPATA1 is localized in the Golgi bodies in the present study, suggesting that the Golgi bodies may not be the major settlement of SPATA1. SPATA1 may represent other resources instead of Golgi bodies for acrosome biogenesis …”

“…The cartoon was drawn based on a model for acrosome biogenesis published previously . Two major systems, Golgi body vesicles and endosomal vesicles, contribute to acrosome formation . IFT20 is present in the Golgi bodies, acrosome area, and manchette.…”

“…SPATA1 may represent other resources instead of Golgi bodies for acrosome biogenesis. 27,28 In CHO cells, both SPATA1 and IFT20 were present as cytoplasmic vesicles; but when co-expressed, two proteins co-localized as large cytoplasmic clusters. Moreover, co-immunoprecipitation assay showed an interaction between SPATA1 and IFT20 in vitro and in vivo.…”

“…30 Two major systems, Golgi body vesicles and endosomal vesicles, contribute to acrosome formation. 27 IFT20 is present in the Golgi bodies, acrosome area, and manchette. SPATA1 is detected in the acrosome, where it provides a docking site for IFT20 AH109 and used a positive control.…”