THE CELL BIOLOGY OF ASCIDIAN SPERM ACTIVATION

该项目的目标是增加少数族裔的参与人数 在解决生育和生物医学问题的研究中 受精、跨膜信号传递和细胞内运动 通过研究激活的海鞘精子在细胞和细胞器中的细胞器 生化水平。 百分之三十的学生曾在我的工作过 过去十年来实验室一直是少数。这些学生中有很多 已经进入健康行业，但很少有人继续从事生物医学 研究。 我实验室的学生参与 MBRS 计划 会增加他们对生物医学研究的接触，将其作为可行的职业 选项。这些学生将研究受精的早期事件 包括精子与卵子的结合、精子的激活以及随后的精子 渗透蛋衣，最终导致融合的过程 精子和卵子的质膜。 精子的细胞机制 由精卵结合触发的激活和渗透是 该项目的重点。 据推测，信号事件 跨膜必须将结合与激活联系起来 产生力的细胞骨架元件的重排必须连接 线粒体迁移的激活。 研究跨膜 信号连接需要给细胞加载激活剂或阻断剂 间接测试某个组件的存在 多磷脂酰肌醇连接机制。 将收集直接证据 通过对该途径的一些蛋白质成员进行生化分析， 测量其行为的产物的外观，并通过研究 他们在存在抗体的情况下的行为，这些抗体可以识别或 干扰他们。这种多磷脂酰肌醇依赖性调节 选择该途径是因为，在某些细胞中，已知它可以提高 细胞内 pH 值和钙，两者都是海鞘的一部分 精子激活。 研究线粒体运动需要探究 参与细胞骨架的生化性质和细胞位置 蛋白质。特定细胞骨架蛋白的存在和运动性 电机将通过聚丙烯酰胺凝胶电泳建立 免疫印迹。 将研究这些蛋白质的细胞位置 通过间接免疫荧光和免疫电子显微镜。

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.