THE CELL BIOLOGY OF ASCIDIAN SPERM ACTIVATION

海鞘精子激活的细胞生物学

• 批准号: 6240373
• 负责人: ROBERT A KOCH
• 金额: $ 4.39万
• 依托单位: CALIFORNIA STATE UNIVERSITY FULLERTON
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-02-01 至 1998-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6240373
• 关键词: Tunicata; alternatives to animals in research; biological signal transduction; cytoskeletal proteins; fertility; fertilization; gel electrophoresis; immunoelectron microscopy; immunofluorescence technique; intracellular transport; membrane transport proteins; sperm motility; western blottings

The goal of this project is to increase the number of minorities involved in research addressing the biomedical problems of fertility and fertilization, signaling across membrane, and intracellular movement of organelles via studies of activated ascidian sperm at the cellular and biochemical levels. Thirty percent of the students who have worked in my' lab over the past ten years have been minorities. Many of these students have gone into health professions, but few have continued in biomedical research. The participation of students in my lab in the MBRS program would increase their exposure to biomedical research as a viable career option. These students would be studying the early events of fertilization include sperm-egg binding, sperm activation and, subsequently, sperm penetration of egg vestments, processes which ultimately lead to fusion of sperm and egg plasma membranes. The cellular mechanisms of sperm activation and penetration, as triggered by sperm-egg binding, are the foci of this project. It is hypothesized that signaling events which cross the membrane must connect binding to activation and that rearrangement of force-producing cytoskeletal elements must connect activation to mitochondrial migration. Investigating the transmembrane signaling connection requires loading of cells with activators or blockers to indirectly test for the presence of the components of a polyphosphainositide-linked mechanism. Direct evidence will be gathered by biochemical analysis of some of the protein members of the pathway, by measuring the appearance of the products of their actions, and by studying their behavior in the presence of antibodies which either identify or interfere with them. This polyphosphainositide-dependent regulatory pathway was chosen because, in some cells, it is known to elevate intracellular pH and calcium, both of which occur as part of ascidian sperm activation. Studying mitochondrial movement requires probing the biochemical nature and cellular location of participating cytoskeletal proteins. The presence of specific cytoskeletal proteins and motility motors will be established by polyacrylamide gel electrophoresis and immunoblotting. The cellular location of these proteins will be studied by indirect immunofluorescence and immunoelectron microscopy.

该项目的目的是增加所涉及的少数民族数量 在解决生育能力和生物医学问题的研究中 受精，跨膜信号传导以及细胞内运动 细胞器通过在细胞和 生化水平。 在我的工作中有30％的学生 在过去的十年中，实验室一直是少数民族。其中许多学生 已经从事卫生职业，但很少有人继续生物医学 研究。 学生参与我的实验室参加MBRS计划 作为可行的职业，将增加他们对生物医学研究的接触 选项。这些学生将研究施肥的早期事件 包括精子 - 蛋的结合，精子激活，随后是精子 卵服的渗透，最终导致融合的过程 精子和卵质膜。 精子的细胞机制 由精子结合触发的激活和穿透是 该项目的焦点。 假设信号事件 越过膜必须连接到激活，并且 产生力的细胞骨架元素的重排必须连接 激活线粒体迁移。 研究跨膜 信号连接需要用激活器或阻滞剂加载单元格 间接测试A的存在 多磷脂固醇连接的机制。 将收集直接证据 通过对途径的某些蛋白质成员的生化分析， 测量其行动产品的外观，并研究 它们在识别或 干扰他们。这种依赖性的多磷脂酰胺依赖性调节性 之所以选择途径，是因为在某些细胞中，已知它会提升 细胞内pH和钙，这两者都作为海鞘的一部分出现 精子激活。 研究线粒体运动需要探测 参与细胞骨架的生化性质和细胞位置 蛋白质。特定细胞骨架蛋白和运动的存在 电动机将通过聚丙烯酰胺凝胶电泳和 免疫印迹。 这些蛋白质的细胞位置将研究 通过间接免疫荧光和免疫电子显微镜。