APOPTOSIS IN GERM CELLS--THE ROLE OF BCL-X

生殖细胞凋亡——BCL-X 的作用

• 批准号: 6440517
• 负责人: PATRICIA L. MORRIS
• 金额: $ 17.42万
• 依托单位: POPULATION COUNCIL
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-04-01 至 2003-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6440517
• 关键词: RNase protection assay; androgens; antisense nucleic acid; apoptosis; cell growth regulation; gel mobility shift assay; gene expression; genetic regulation; genetic regulatory element; genetically modified animals; germ cells; hormone regulation /control mechanism; laboratory mouse; laboratory rat; molecular cloning; nucleic acid biosynthesis; nucleic acid sequence; oncoproteins; protein isoforms; protein structure function; spermatogenesis; testis; tissue /cell culture

The long-term goal of these studies if to determine how the survival of spermatogenic cells is regulated by the Bcl-2 family of onoproteins. Although the potential exists for multiple influences on the regulation of mitotic events in the proliferating germ cells, a significant body of data has established a correlation between the size of the spermatogenic compartment and the number of early germ ells that do not survive to the haploid stage. We recently identified bcl-x as a member of the bcl-2 family of oncogenes preferentially expressed in the rat testis. Studies in this proposal concentrate on the regulation of the expression of two protein isoforms, Bcl-xL and Bcl-xS, and their roles in determining survival versus death of germ cells. Our studies will focus on the unique features of bcl-x expression in the seminiferous tubule. First, the bcl-x gene will be cloned and potential regulatory elements identified in its proximal promoter by DNA sequence analysis. Second, the specific regulation of the isolforms of bcl-x mRNA by androgens and stem cell growth factor will be evaluated using primary culture models of highly purified spermatogonia and early spermatocytes. Third, the stage specificity of bcl-x expression in the seminiferous tubule will be determined and the effects of stem cell factor and androgens on bcl-x expression and DNA synthesis in premeiotic germ cells ascertained. Fourth, the effects of changes in the ratio of the members of the Bcl-2 family of oncoproteins on programmed cell death in spermatogonia will be examined. Specific alterations in the balance of suppression versus induction of cell death will be accomplished by specific inhibition of Bcl-xL mRNA using an antisense oligonucleotide approach and by overexpression of Bcl-2 in transfected germ cells. Characterization of germ cell development and apoptotic processes in the bcl-2 transgenic mouse will facilitate studies to delineate the role of the bcl-2 family of genes in the survival of the male germ cells. Since testicular biopsies rom infertile males often show decreased numbers of proliferating and developing germ cells, we anticipate that this research will lead to significant new data on the relevance of apoptosis and its regulation to human male fertility.

这些研究的长期目标是确定生存方式 精子生成细胞受Onopotin的Bcl-2家族调节。 尽管对监管的多种影响存在潜在的潜力 增殖生殖细胞中有丝分裂事件的重要物体 数据已经建立了大小之间的相关性 精子室和做早期细菌的数量 无法生存到单倍体阶段。 我们最近将BCL-X确定为 Bcl-2癌基因家族的成员优先表达 大鼠睾丸。 该提案中的研究集中于调节 两种蛋白质同工型的表达 它们在确定生存与死亡细胞死亡中的作用。 我们的 研究将重点介绍BCL-X表达的独特特征 生精的小管。首先，Bcl-X基因将被克隆并潜在 DNA在其近端启动子中鉴定的调节元件 序列分析。 其次，对隔板的特定法规 雄激素和干细胞生长因子的Bcl-X mRNA将是 使用高度纯化的主要培养模型进行评估 精子和早期精子细胞。第三，阶段特异性 将确定生精管中的BCl-X表达，并确定 干细胞因子和雄激素对BCL-X表达和DNA的影响 预元生殖细胞中的合成确定。 第四，效果 BCL-2家族成员的比率变化 精子中有程序性细胞死亡的癌蛋白将是 检查。 抑制平衡与 细胞死亡的诱导将通过特定的抑制来实现 BCl-XL mRNA使用反义寡核苷酸方法和 在转染的生殖细胞中Bcl-2的过表达。 表征 Bcl-2转基因的生殖细胞发育和凋亡过程 鼠标将促进研究以描绘Bcl-2家族的作用 男性生殖细胞存活中的基因。 由于睾丸 活检不育男性通常显示出数量减少 增殖和发育的生殖细胞，我们预计这是 研究将导致有关相关性的重要新数据 凋亡及其对人类男性生育能力的调节。