Title

Characterization of the sperm surface protein PH-20 as an antigen for a contraceptive vaccine

Date of Completion

January 1994

Keywords

Biology, Molecular|Biology, Cell

Degree

Ph.D.

Abstract

The guinea pig sperm surface protein PH-20 has an essential function in sperm adhesion to the zona pellucida of guinea pig eggs, fully achieved by immunizing either male or female guinea pigs with purified guinea pig PH-20 (Primakoff, P., Lathrop, W. F., Woolman, L., Cowan, A. & D. G. Myles (1988) Nature 335, 543-546). The isolation of human and cynomolgus monkey PH-20 cDNAs presented in this thesis is a key step toward testing the function of primate PH-20 and the contraceptive efficacy of PH-20 immunization in primates. The deduced amino acid sequence of human PH-20 has 509 residues and is 59% identical with guinea pig PH-20, suggesting they may have a conserved function and immunogenicity. Southern blots show that there is a single PH-20 gene in the human genome and Northern blots of human testis poly(A)$\sp+$ RNA show a 2.4kb message. Northern blots of tissues other than testis are negative for PH-20 indicating that human PH-20 is testis-specific.^ A typical mammalian egg is surrounded by an outer layer of about 3000 cumulus cells embedded in an extracellular matrix rich in hyaluronic acid. A current, widely proposed model is that the fertilizing sperm, while it is acrosome intact, passes through the cumulus cell layer and binds to the egg zona pellucida. This current model lacks a well-supported explanation for how sperm penetrate the cumulus layer. My thesis prevent the evidence that the sperm protein PH-20 has a hyaluronidase activity and is present on the plasma membrane of mouse and human sperm. Brief treatment with recombinant, purified PH-20 can release all the cumulus cells surrounding mouse eggs. Acrosome intact mouse sperm incubated with anti-PH-20 antibodies can not pass through the cumulus layer and thus can not reach the zona pellucida. These results, indicating that PH-20 enables acrosome intact sperm to penetrate the cumulus barrier, reveal a mechanism for cumulus penetration and thus provide the missing element in the current model. ^