GENETIC AND MOLECULAR ANALYSIS OF GERMLINE DETERMINANTS

这个项目的长期目标是了解生物化学的本质 以及种质的功能。果蝇的后极浆 对体细胞和生殖系组织的发育起着至关重要的作用。 这些功能依赖于母体效应基因的正常活动。 奥斯卡。来自纯合子突变奥斯卡雌性的成熟卵母细胞缺乏 在正常后极发现的形态特化 卵母细胞。从突变卵母细胞发育而来的胚胎不会形成极细胞， 生殖系前体细胞，不发育腹段。奥斯卡 RNA定位于胚胎的后极，Oskar蛋白 因此可以是极浆的一种成分。遗传证据表明 奥斯卡基因产物是定位额外基因所必需的 对生殖细胞测定至关重要的产品，它也是必需的 用于控制分段的Nanos基因产物的本地化 在腹部区域。 最初，奥斯卡基因产物的分子性质及其 将描述卵子发生和胚胎发生过程中的分布。这个 奥斯卡转录单位将被定义，基因将被克隆，并 编码区已测序。Oskar RNA和蛋白质的分布将是 非放射性标记在卵子发生和胚胎发育过程中的研究 Oskar DNA和/或反义RNA探针和抗体 奥斯卡蛋白质。随后，奥斯卡产品成为 将分析定位于后极浆的部位。的能力 在其他极质成分中突变以定位Oskar RNA和/或蛋白质 将会受到考验。Oskar RNA和蛋白质中的顺式作用序列 适当本地化的必要条件将通过对突变的分析来确定 Oskar等位基因和缺失分析。这些序列满足以下条件 将使用报告基因检测定位的Oskar RNA和/或蛋白质 融合。最后，为了理解奥斯卡在本地化方面的作用 生殖细胞和体细胞决定因素，奥斯卡突变对 后极血浆的其他成分的定位，例如 Nanos RNA和Vasa蛋白，将被研究。直接互动 在Oskar RNA和蛋白质与极质的其他成分之间 将通过使用针对人的抗体的交配试验进行研究 极浆产品。

The long term goal of this project is to understand the biochemical nature and function of the germ plasm. The posterior pole plasm of Drosophila plays a critical role for the development of somatic and germ line tissues. These functions depend on the normal activity of the maternal effect gene oskar. Mature oocytes derived from homozygous mutant oskar females lack the morphological specializations found at the posterior pole of the normal oocyte. Embryos developing from mutant oocytes do not form pole cells, the germ line precursor cells, and do not develop abdominal segments. Oskar RNA is localized at the posterior pole of the embryo and the oskar protein may thus be a component of the pole plasm. Genetic evidence indicates that the oskar gene product is required for the localization of additional gene products critical for germ cell determination and that it is also required for the localization of the nanos gene product which controls segmentation in the abdominal region. Initially, the molecular nature of the oskar gene product and its distribution during oogenesis and embryogenesis will be characterized. The oskar transcription unit will be defined, the gene will be cloned and the coding region sequenced. The distribution of oskar RNA and protein will be studied during oogenesis and embryogenesis using nonradioactively labelled oskar DNA and/or antisense RNA probes and antibodies directed against the oskar protein. Subsequently, the mechanisms by which oskar products become localized to the posterior pole plasm will be analyzed. The ability of mutants in other pole plasm components to localize oskar RNA and/or protein will be tested. Cis acting sequences in the oskar RNA and protein necessary for proper localization will be determined by analysis of mutant oskar alleles and deletion assays. The sufficiency of these sequences to localize oskar RNA and/or protein will be tested using reporter gene fusions. Finally, to understand the role of oskar for the localization of germ cell and somatic determinants, the effect of oskar mutations on the localization of other components of the posterior pole plasm, e.g. the nanos RNA and the vasa protein, will be studied. Direct interactions between the oskar RNA and protein with other components of the pole plasm will be studied by copurification assays using antibodies directed against pole plasm products.