GENETICS OF CAENORHABDITIS ELEGANS SPERM MORPHOGENESIS

秀丽隐杆线虫精子形态发生的遗传学

基本信息

批准号：
2608882
负责人：
STEVEN W. L'HERNAULT
金额：
$ 17.33万
依托单位：
EMORY UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1988
资助国家：
美国
起止时间：
1988-08-01 至 1999-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2608882
关键词：
Caenorhabditis elegans alleles cell differentiation cytogenetics developmental genetics electrophysiology gene expression gene mutation genetic mapping genetic recombination genetically modified animals histogenesis immunocytochemistry molecular cloning site directed mutagenesis spermatogenesis suppressor mutations voltage /patch clamp

项目摘要

Asymmetric partitioning of cytoplasm and generation of cellular polarity both play important roles during differentiation. We propose to further characterize genes in a system where asymmetric partitioning of organelles and cytoplasm is both obvious and amenable to genetic analyses: the amoeboid sperm of the nematode Caenorhabditis elegans. Many stages in the differentiation of this cell can be genetically dissected, its will differentiate in vitro and biochemistry on purified cells is possible. Five genes (spe-4, spe-5, spe-17, spe-16, e1947 dominant spec) with specific cytoplasmic partitioning defects during spermatogenesis will be analyzed by genetic, cytological and molecular analysis. These genes were chosen because each is associated with a mutant phenotype that affects asymmetric cytoplasmic partitioning and all have defects only in sperm. After all affect an unusual Golgi derived organelle, the fibrous body- membranous organelle (FB-MO) complexes. These organelles play a central role in partitioning of cytoplasm during the asymmetric cytokinesis that accompanies meiosis II. Depending on the mutant, disruption of FB-MO structure can either abolish the second cytokinesis or grossly affect both the cytology and function of resulting spermatids. Consequently, understanding the morphogenesis and function of the FB-MO complexes is essential in order to determine the roles that govern cellular polarity in this experimental system. The specific aims are: 1) development of a germline expression vector to allow optimized expression of reporter gene expression in the testes; 2) positional cloning of spe-16 and localizing of its encoded protein; 3) positional cloning of spe-5 and localization of its encoded protein; 4) knockout mutagenesis for a potential FB-MO resident ion channel, named ssp-20, for which we have a specific; 5) genetic analysis of dominant spe mutants have global defects in cellular polarity such that aspects of FB-MO differentiation proceed in a cell that cannot divide; 6) recovery of suppressors to begin constructing developmental pathways operative during spermatogenesis. These analysis will show how molecules involved in the sperm morphogenesis play their essential role in asymmetric cytoplasmic partitioning during sperm differentiation. This is a generally important problem during both normal cytodifferentiation and in the pathologic changes such as cancer and Alzheimer's disease. Additionally. C. elegans spermatogenesis will contribute to understanding this process in the medically and agriculturally important nematode parasites that have amoeboid sperm, but are difficult to study in the laboratory.

细胞质的不对称分区和细胞极性产生两者在差异化过程中都起着重要的作用。我们建议进一步在细胞器不对称分区的系统中表征基因细胞质既明显又可以接受遗传分析：线虫秀丽隐杆线虫的变形虫精子。许多阶段该细胞的分化可以在遗传上解剖，其意愿在纯化细胞上可以分化体外和生物化学。具有五个基因（SPE-4，SPE-5，SPE-17，SPE-16，E1947主要规格）精子发生过程中特定的细胞质分配缺陷将是通过遗传，细胞学和分子分析分析。这些基因是之所以选择，是因为每个都与影响的突变表型相关联不对称的细胞质分配，并且仅在精子中存在缺陷。毕竟影响了异常的高尔基体衍生的细胞器，纤维的身体膜细胞器（FB-MO）复合物。这些细胞器的中央在不对称细胞因子过程中细胞质分割中的作用伴随减数分裂II。根据突变体，FB-MO的破坏结构可以废除第二个细胞因子或严重影响两者产生的精子的细胞学和功能。最后，了解FB-MO复合物的形态发生和功能是为了确定控制细胞极性的作用至关重要这个实验系统。具体目的是：1）开发种系表达载体允许优化报告基因的表达在睾丸中的表达； 2）SPE-16的位置克隆和本地化其编码蛋白； 3）SPE-5的位置克隆和定位它的编码蛋白； 4）潜在FB-MO的敲除诱变居民离子频道，名为SSP-20，我们有一个特定的； 5）主导SPE突变体的遗传分析在细胞中具有全球缺陷极性使得FB-MO分化的各个方面在细胞中进行无法分裂； 6）恢复抑制器以开始构建精子发生过程中的发育途径。这些分析将展示与精子形态发生有关的分子如何发挥他们的作用精子期间的不对称细胞质分配中的重要作用分化。这是正常的两个通常重要的问题细胞分化以及在病理变化中，例如癌症和阿尔茨海默氏病。此外。秀丽隐杆线虫的精子发生将在医学上理解这一过程以及具有变性精子的农业重要线虫寄生虫，但在实验室很难学习。