The long-term goals of this research project are to continue to define both chemically and biologically a set of sperm plasma membrane molecules which are functional autoantigens and necessary for one or more steps in the fertilization process. During the current grant period, a family of rabbit sperm autoantigens (RSA) has been characterized and discovered to function in binding the sperm to the egg's zona pellucida. Moreover, mouse and human spermatozoa are immunologically cross-reactive with monoclonal anti-RSA-1. To continue this line of investigation, these RSA molecules will be further characterized with regard to their role in autoimmunity and fertilization. Therefore, the current proposal's specific aims are: 1) the biochemical identification and characterization of the site or sites on the autoantigenic sperm membrane molecules which specifically bind autoantibody and zona pellucida, 2) the isolation and characterization of the sperm zona binding proteins (ZBPs) from rabbit and mouse spermatozoa. In the rabbit, RSA-1 is one of the four major ZBPs and each of the other three will be isolated and compared to RSA-1 with regard to amino acid composition, carbohydrate composition and the ability to bind zona proteins, 3) an investigation of the specific binding of the rabbit seminal plasma protein complex (84 Kd-complex) to RSA-1 and its localization and fate during fertilization, 4) a continuation of the study of the distribution of sperm membrane autoantigens and actin in spermatogenic cells and spermatozoa.
This aim will ask: a) is filamentous actin reformed during the acrosome reaction or later during sperm penetration of the zona pellucida and b) is the actin (filamentous or non-filamentous) localized in association with either the sperm membrane autoantigens or the zona binding proteins? 5) the isolation of a cDNA clone for RSA-1 from mRNA obtained from rabbit testes. This RSA-1 cDNA clone will allow the determination of the complete amino acid sequence of RSA-1 (the N-terminal is blocked), the determination of when RSA-1 mRNA is first synthesized during development, the location and structure of the RSA-1 gene or genes, and the presence of RSA-1 genes in other species.
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