BASIS FOR MALE INFERTILITY: MOLECULAR MODELS

男性生育能力的一个重要方面是精子主动 通过女性生殖道的某些区域移动并穿透 鸡蛋投资正常的鞭毛不具备这些功能。 然而，我们对控制精子鞭毛装配的基因知之甚少 或者说是哺乳动物的运动能力。私家侦探已经定位了一个基因座杂交不育6号 （Hst-6），到等于或小于1厘摩（cM）的区域， 小鼠17号染色体近三分之一处。结合某些 该基因座的等位基因导致严重的雄性不育。实验室老鼠， 来自另一只小鼠的Hst-6等位基因（Hst-6S/Hst-6S）纯合 种Mus spretus（S）是不育的。它们的精子是不动的， 缺乏可识别的轴丝。杂合子实验小鼠 Hst-6S的一个拷贝和Hst-6S近端三分之一的变体形式， 17号染色体上的t单倍型（Hst-6S/t）也是不育的。他们的 精子是能动的，但这些精子的鞭毛弯曲异常，在 与携带两个t的不育雄性精子的鞭毛相同 单倍型（t/t）。因此，很明显，Hst-6具有两个遗传变异体， 等位基因，一个影响轴丝分化和其他， 活动精子鞭毛的弯曲。 因为在哺乳动物中调节这些功能的基因还没有被发现， Hst-6基因座的分离提供了一种新的方法来确定这样的基因。 因此，PI。将首先克隆所有睾丸表达的基因， Hst-6位点（Aim I）。为了确定哪些候选基因 很可能是Hst-6私家侦探将分析 睾丸mRNA和蛋白表达的基因序列和模式 每个候选基因的每个等位基因 Hst-6突变体表型（Aim II）。根据已知的功能， Hst-6（轴丝组装和鞭毛弯曲），很可能是 Hst-6蛋白特异性定位于或非常接近轴丝 附睾尾部精子私家侦探我将用以下方法来检验这一假设： 免疫细胞化学检测附睾尾部精子的特异性 候选蛋白质的亚细胞定位， 定位于发育中的精子细胞的尾部区域和/或成熟的精子细胞 睾丸精子（Aim III）。这些实验将有助于 这些信息有助于我们了解精子鞭毛的遗传控制 组装和移动。此外，他们的成功将产生一个范例， 哺乳动物精子发育和功能的研究前景 鞭毛在生化水平上的变化。

An essential aspect of male fertility is the ability of sperm to actively move through certain regions of the female genital tract and penetrate the egg investments. Indispensable to these functions is a normal flagellum. Yet little is known about the genes which control sperm flagellar assembly or motility in mammals. The P.I. has mapped a locus, Hybrid Sterility-6 (Hst-6), to a region of equal to or less than 1 centimorgan (cM) in the proximal one third of mouse chromosome 17. The combination of certain alleles of this locus causes profound male sterility. Laboratory mice who are homozygous for an allele of Hst-6 (Hst-6S/Hst-6S) from another mouse species, Mus spretus (S), are sterile. Their sperm are immotile, due to the absence of a recognizable axoneme. Laboratory mice heterozygous for one copy of Hst-6S and a variant form of the proximal one third of chromosome 17 called a t haplotype (Hst-6S/t) are also sterile. Their sperm are motile, but the flagella of these sperm curve abnormally, in the same way as the flagella of sperm from sterile males carrying two t haplotypes (t/t). Thus, it is clear that Hst-6 has two genetically variant alleles, one affecting axonemal differentiation and the other, the curvature of the flagella of motile sperm. Because genes regulating these functions in mammals have not been isolated, the Hst-6 locus offers a novel approach to identify such genes. Therefore, the P.I. will first clone all testis-expressed genes that map to the Hst-6 locus (Aim I). To determine which of the candidate genes is most likely to be Hst-6, the P.I. will analyze the differences between the gene sequences and patterns of testicular mRNA and protein expression of each allele of each candidate gene with reference to our present knowledge of the Hst-6 mutant phenotypes (Aim II). Based upon the known functions of Hst-6 (axonemal assembly and flagellar curvature), it is likely that the Hst-6 protein specifically localizes in or very close to the axoneme of cauda epididymal sperm. The P.I. will test this hypothesis by using immunocytochemistry to examine cauda epididymal sperm for the specific subcellular location of candidate proteins that have already been localized to the tail region of developing spermatids and/or mature testicular sperm (Aim III). These experiments will contribute critical information to our understanding of the genetic control of sperm flagellar assembly and movement. Moreover, their success will produce a paradigm for the future study of the development and function of the mammalian sperm flagellum at the biochemical level.