LB5 antibody was selected from a monoclonal antibody (mAb) library directed against human sperm proteins. LB5 mAb detected the corresponding protein SOB3 in the neck region and the flagellum of most live ejaculated sperm while it labelled, in addition, the acrosome of about 10-20% of spermatozoa. The percentage of LB5 acrosome-stained sperm was significantly correlated with the percentages of either spontaneous or A23187-induced acrosome-reacted sperm. While SOB3 could not be detected in the testis, it appeared in spermatozoa from the corpus epididymis segment. LB5 mAb impaired neither sperm motion parameters, acrosomal reaction triggering, nor sperm binding to zona-free hamster oocytes. By contrast, LB5 Fab fragments (200 micrograms/ml) inhibited sperm binding to human zonae pellicidae by 35.7%. If sperm were induced to acrosome react with A23187 prior to LB5 treatment, the inhibitory effect shifted to 59.9%, while no significant effect was observed following A23187 incubation alone. Western blotting of human sperm and cauda epididymis extracts revealed two bands of 18 and 19 kDa. While no cross-reaction was observed with other tested organs, a similar 18-kDa band was revealed in erythocytes and one of 19 kDa in B-lymphocytes. No cross-reactivity could be evidenced in any animal sperm analyzed. SOB3 was first separated in a 17- to 20-kDa preparative electrophoresis fraction and finally purified by isoelectrofocusing according to its pl of 9.8. These results suggest that SOB3 is localized under the outer acrosomal membrane, that it participates in secondary sperm binding to the zona pellucida, and that it shares homologies with the immune system.
La fecondation met en jeu, entre les gametes mâles et femelles des mammiferes, des mecanismes d'adherence et d'activation comparables a ceux decrits dans le developpement embryonnaire et metastasique, le systeme immunitaire et l'infection virale. La premiere etape, la liaison des spermatozoides a la zone pellucide qui protege l'ovocyte, met en jeu des interactions avec les oligosaccharides portes par les proteines de la zone pellucide; c'est a ce stade qu'interviennent les barrieres d'espece qui interdisent les fecondations croisees. Cette liaison active les spermatozoides en declenchant la reaction acrosomiale qui permet la liberation des enzymes proteolytiques necessaires a la dispersion de la zone pellucide et a sa traversee. L'attachement du spermatozoide a la membrane plasmique de l'ovocyte semble dependre de la reconnaissance de sequences peptidiques et aboutit a l'activation de l'ovocyte et a la reprise de sa meiose. La reconnaissance de molecules complementaires semble engendrer a chacune de ces deux etapes la stimulation de voies de transmission responsables de l'activation de l'un et l'autre gametes.
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