The long-term objective of our studies are to determine a) if the expression of PH-20 or the Sperm Adhesion Molecule 1, SPAM1 (genetic nomenclature), reveals a mechanism for transmission ratio distortion (TRD), b) if the Spam1 gene product is retained in individual spermatids and the regulatory mechanism that controls its mRNA distribution, and c) how overexpression of SPAM1 affects sperm function. The proposal seeks to determine if the murine Spam1 gene product, a sperm membrane protein (Spam1), is shared among conjoined spermatids and whether the sperm of a heterozygote or hemizygote are phenotypically distinct with respect to this antigen. The study will make use of a Robertsonian translocation {Rb(6.16) or Rb} resource in which heterozygotes show a TRD and in which Spam1 expression is reduced, as well as hemizygotes for an overexpressed Spam1 transgene to be constructed. The effects of overexpression of Spam1 on the fertilizing competence of sperm will also be addressed by taking advantage of the hemizygotes to generate transgenic mice for functional studies. Thus the proposal addresses several important problems in reproductive biology: the biochemical equivalence of sperm in a heterozygote, a fundamental question in testicular biology; the compartmentalization of mRNA and its regulation by co-translational assembly and sequences in the 3' UTR, and the impact of overexpression of Spam1 on mammalian fertilization. The following hypotheses will thus be tested. 1) There is a bimodal distribution of the quantity of Spam1 mRNA in spermatids and protein in sperm from Rb(6.16)/+ mice, compared to a unimodal one in +/+ and Rb/Rb mice, with the lower RNA deposits in Rb-bearing cells. The latter can be rescued by transgenesis to eliminate their reduced fertilizing ability and TRD in Rb carriers; 2) Mice hemizygous for an overexpressed Spam1 transgene will reveal whether or not Spam1 mRNA and protein are equally shared among conjoined spermatids to produce biochemically and functionally equivalent or different sperm. Homozygotes will determine the effect of overexpression on the rates of investment penetration, zona binding, and fertilization, 3) Co-translational assembly, sequences mediating GPI-linkage of Spam 1, and sequences in the 3' UTR of the gene mediate its mRNA compartmentalization which leads to TRD. The work promises to impact current thinking about critical principles underlying haploid gene expression, elucidate a mechanism for meiotic drive, increase our understanding of the molecular mechanisms that underpin mammalian fertilization, and could identify a genetic basis for altered sperm-egg interactions involved in human fertility.
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