GENE EXPRESSION IN SPERMATOGENIC CELLS

工作总结：这些研究的目的是确定 调控沙文主义基因的发育表达及其作用 它们在生精细胞中编码的蛋白质。这些蛋白质有 唯一或同源但不完全相同的序列 其他细胞中的蛋白质。同源蛋白质要么由一种 基因家族成员只在生精细胞中表达或由 生精细胞特异的替代转录本。将近50岁 沙文主义基因已经被识别出来，这些研究使用基因 基因敲除方法来研究一些据信是 在雄配子发育和功能中起重要作用。受精素b是一种精子 表面糖蛋白和ADAM的成员(一种去整合素和 金属蛋白酶结构域)家族。合作者(迈尔斯和普里马科夫) 提示它在精子-卵子相互作用中起关键作用，并与 卵子表面的A1B6整合素。小鼠为靶向杂合子 受精蛋白b基因的突变正在交配，同源雄性正在交配 预计会降低生育率。鱼精蛋白1和2是高度碱性的 取代组蛋白的核蛋白，被认为是 在精子过程中没有核小体的情况下对DNA致密是必不可少的 发展。他们的一个合作者(赫克特)已经证明他们有 精子细胞中发育和转录调控的表达。 已经产生了嵌合体小鼠，它们携带有靶向突变的 鱼精蛋白1基因，并正在交配产生杂合子后代。 鱼精蛋白2的敲除结构已经完成，将用于 在胚胎干细胞中定位这种基因。预计 一个或两个基因的破坏将导致异常的核致密 以及纯合子男性的不育症。3-磷酸甘油醛 脱氢酶是一种关键的糖酵解酶，也是一种同系物(GAPD-S) 仅在精子细胞中合成的蛋白质在调节 受精所需的三磷酸腺苷的生成。这种酶也是 男性生殖毒物(S)-3-氯甲醛可能成为攻击目标 这是工业溶剂环氧氯丙烷的代谢物。研究 用转基因小鼠确定正确所需的序列 GAPD-S基因在转录起始区230bp内的表达 地点。正在为Gapd-S准备一个击倒结构， 预测的敲除表型是看起来正常的精子 不能激活糖酵解，获得高度激活的运动，或者 让卵子受精。

Summary of Work: The aim of these studies is to determine the mechanisms that regulate developmental expression of chauvinist genes and the roles of the proteins they encode in spermatogenic cells. These proteins have sequences that are either unique or homologous but not identical to proteins in other cells. A homologous protein is either encoded by a gene family member expressed only in spermatogenic cells or by a spermatogenic cell-specific alternative transcript. Nearly fifty chauvinist genes have been identified and these studies use gene knock-out approaches to examine the role of some that are believed to be important in male gamete development and function. Fertilin b is a sperm surface glycoprotein and a member of the ADAM (a disintegrin and metalloprotease domain) family. Collaborators (Myles and Primakoff) have suggested that it has a key role in sperm-egg interaction and binds to an a1B6 integrin on the egg surface. Mice heterozygous for a targeted mutation in the fertilin b gene are being mated and homologous males are expected to have reduced fertility. Protamine 1 and 2 are highly basic nuclear proteins that replace the histones and are thought to be essential for DNA compaction in the absence of nucleosomes during sperm development. They have been shown by a collaborator (Hecht) to have developmentally and transcriptionally regulated expression in spermatids. Chimeric mice have been produced that carry a targeted mutation in the protamine 1 gene and are being mated to produce heterozygous offspring. A knock-out construct for protamine 2 has been completed and will be used to target this gene in embyronic stem cells. It is expected that disruption of one or both genes will lead to abnormal nuclear compaction and infertility in homozygous males. Glyceraldehyde 3-phosphate dehydrogenase is a key glycolytic enzyme and a homolog (GAPD-S) synthesized only in spermatids has a key role in regulating the generation of ATP required for fertilization. The enzyme is also the likely target for (S)-3-chlorolactaldehyde, a male reproductive toxicant that is a metabolite of the industrial solvent epichlorohydrin. Studies with transgenic mice determined that sequences required for correct expression of Gapd-s are within 230 bp of the transcription initiation site. A knock-out construct for Gapd-s is being prepared and the predicted phenotype of the knock-out is normal appearing sperm that are unable to activate glycolysis, achieve hyper-activated motility, or fertilize eggs.