9513464 Pilder The objective of this proposal is to identify and localize mutations in the mouse t complex whose products perturb sperm penetration of the investment-free oocyte, a process whose molecular nature is not well understood in mammals. The proximal 30-40 million base pairs of mouse chromosome 17 is known as the t complex. Variant forms of the t complex, called t haplotypes, can be distinguished from wild type homologs structurally, by the presence of four large, non-overlapping inversions which extend nearly over the entire region, and phenotypically, by their detrimental effects on male fertility. Male mice carrying two t haplotypes (t/t) are invariably sterile, and their sperm are unable to penetrate investment-free mouse eggs, indication that there are t complex genes intimately involved in the process of sperm-oolemma penetration (SOP). Preliminary data suggest that at least one gene affecting sperm-egg fusion is located in the largest and most distal t haplotype inversion (In4). Specific Aim I is designed to determine if or how well sperm from a mouse that is homozygous only for In4 of the t haplotype (In4/In4) can penetrate the investment-free egg, in comparison to sperm from nearly congenic +/In4 heterozygotes or +/+ homozygotes. More precise localization of a t haplotype specific sterility phenotype to an intra-inversion sub-region requires an indirect approach, since intra-inversion recombinants between t haplotypes and wild type homologs are extremely rare. Since male laboratory mice carrying the t complex from M. spretus (S), a distantly related mouse species, and a t haplotype homolog are invariably sterile, and because S and + homologs of the t complex are not inverted relative to each other in the t haplotype inversions in which sterility factors are known to reside, intra-inversion S-+ recombinants can be produced and used to more precisely map these sterility factors. Preliminary data suggest that a SOP factor that poorly complements the t haplotype SOP defect maps to the In4 region of the M. Spretus t complex. Specific Aim II is designed to use S-+ recombinants to examine the genetic basis of the S/t SOP detect for evidence of allelism between S and t haplotype SOP genes. Additionally, the phenotype of defective sperm penetration of investment-free mouse eggs will be more precisely mapped using sperm from congenic mice carrying a variety of S-+ recombinant t complexes (specific Aim III). the precise localization of t complex SOP genes will make possible their future isolation and molecular characterization. ***
