THE MOLECULAR BIOLOGY OF ORGANELLAR TRANSLOCATION

我建议综合运用遗传学生化学和形态学 确定细胞器通过的机制的方法 细胞 理解这些机制很重要，因为 重要的生理现象，如轴突运输和酶， 激素分泌是细胞器易位形式。 我建议 研究丝状真菌细胞器移位的机制 构巢曲霉，其独特地适合于组合遗传， 这些现象的形态学和生物化学研究。 最初我 建议分离条件致死突变体并筛选这些突变体 在限制性条件下， 条件 除了随机分离的条件致死突变体， 我建议分离出条件性致死抑制因子 条件致死微管蛋白突变体，现有和 新分离的细胞器易位突变体和条件致死 细胞松弛素抗性突变体。 以往的经验表明， 细胞器易位突变体应该发生在高频率之间 这些变种人 新分离的细胞器易位突变体将是 测试了与现有细胞器易位突变体的等位性， 彼此之间，将被映射到locus。 这些变种人将会 在光学显微镜水平上进行形态学表征，以确定 如果每个突变特异性地阻断一种形式的细胞器易位， 例如核运动，或者它是否具有更普遍的影响。 这些突变体 将在电子显微镜下进行检查，以确定它们是否影响 微管组装或分布。 最后，我将尝试识别 用双向凝胶电泳和I 将尝试开发显微注射和DNA转化试验， 缺陷基因产物

I propose to use a combination of genetic, biochemical and morphological methods to determine the mechanisms by which organelles move through cells. It is important to understand these mechanisms because such physiologically important phenomena as axonal transport and enzyme and hormonal secretion are forms of organellar translocation. I propose to study the mechanisms of organellar translocation in the filamentous fungus Aspergillus nidulans which is uniquely suited for combined genetic, morphological and biochemical studies of these phenomena. Initially I propose to isolate conditionally-lethal mutants and screen these mutants for blockage of nuclear and mitochondrial migration under restrictive conditions. In addition to randomly isolated conditionally-lethal mutants, I propose to isolate conditionally-lethal suppressors of existing conditionally-lethal tubulin mutants, suppressors of existing and newly-isolated organellar translocation mutants and conditionally-lethal cytochalasin-resistant mutants. Previous experience suggests that organellar translocation mutants should occur at a high frequency among these mutants. The newly-isolated organellar translocation mutants will be tested for allelism to existing organellar translocation mutants and to each other and will be mapped to locus. These mutants will be characterized morphologically at the light-microscopic level to determine if each mutation specifically blocks one form of organellar translocation, such as nuclear movement, or if it has more general effects. These mutants will be examined in the electron microscope to determine if they affect microtubule assembly or distribution. Finally, I will attempt to identify the defective gene products by two-dimensional gel electrophoresis and I will attempt to develop microinjection and DNA transformation assays for the defective gene products.