EXPRESSION AND LOCALIZATION OF CRISP PROTEINS IN EPIDIDYMAL EPITHELIUM AND EPISIDIDYMOSOMES

REBAGLIATI CID A; CARVAJAL G; VALERIA SULZYK; BATTISTONE A; BRETON S; WEIGEL MUÑOZ M; CUASNICÚ PS

Congreso; Reunión Anual de Sociedades de Biociencias; 2023

Mammalian sperm acquire their fertilizing ability during a maturationprocess that occurs in the epididymis, mainly as a consequenceof the association of epididymal proteins with the sperm surface.Results from our laboratory showed that double knockout mice fortwo such proteins, epididymal CRISP1 and CRISP4, were subfertileand exhibit clear defects in epididymal epithelium differentiationand luminal acidification critical for sperm maturation. In view of this,in the present work, we studied cell expression and localization ofCRISP1 and CRISP4 along the epididymis. Analysis of protein expressionby Western Blot and indirect immunofluorescence showedthat whereas CRISP1 was expressed in the three regions of theorgan (i.e. caput, corpus and cauda), CRISP4 was mostly detectedin the caput and proximal corpus, being almost absent in thecauda. The use of specific markers for each epididymal epithelialcell type revealed that whereas CRISP1 and CRISP4 were not detectedin basal cells, both were expressed not only in principal cells,as previously reported, but also in clear cells known to be involvedin proton secretion, supporting the involvement of these proteins inluminal pH regulation. Considering the important role that extracellularvesicles (i.e epididymosomes) play in protein association withsperm during maturation, the presence of CRISP1 and CRISP4 inepididymosomes from different regions of the organ was examined.After confirmation of successful isolation of the epididymosomes byelectron microscopy, the presence of CRISP1 and CRISP4 in thesevesicles was analyzed by Western blot. Results showed that whereasCRISP1 was present in those vesicles obtained from either thecaput or the cauda, CRISP4 was only detected in caput epididymosomes.Together, these results support the relevance of epididymalCRISP proteins for sperm maturation, contributing to a better understandingof the mechanisms underlying epididymal physiology andmale fertility.