CONTROL OF GENE ACTIVITY AND ORGANELLE BIOGENESIS

本提案的长期目标是了解 纤毛生物发生的分子机制及其 与细胞周期的关系 低等真核生物， 以嗜热四膜虫为模型。 一个屏幕被 在纤毛中产生条件性（TS）突变 再生，其中令人惊讶的高百分比（2/5）是 细胞周期阻滞突变体。 前两个变种人 可以通过细胞质交换来拯救。 显微注射 测定将用于纯化突变基因产物。 突变基因 将被克隆并用于开发DNA介导的转化 系统，这将反过来，允许表征大量 通过互补克隆后的基因。 突变体影响 单个α-微管蛋白基因和突变的基因外回复突变体 影响纤毛生物发生和细胞周期的细胞也将被分离 和表征了 α-微管蛋白和β-微管蛋白的详细分析 将进行基因组织和表达。 通过隔离 并描述了大量正常和突变基因 在纤毛生物发生和细胞周期调节中都很重要， 应该可以阐明两种正常 细胞骨架功能和细胞周期进程， 在肿瘤细胞中经常是异常的。

The long-term goals of this proposal are to understand the molecular mechanisms underlying the biogenesis of cilia and their relationship to cell-cycle progression. The lower eukaryote, Tetrahymena thermophila is used as a model. A screen has been developed that yields conditional (ts) mutations in cilia regeneration, a surprisingly high percentage (2/5) of which are cell cycle arrest mutants. The first two of these mutants to be studied can be rescued by cytoplasmic exchange. A microinjection assay will be used to purify mutant gene products. Mutant genes will be cloned and used to develop a DNA-mediated transformation system that will, in turn, allow characterization of a large number of genes after cloning by complementation. Mutants affecting the single alpha-tubulin gene and extragenic revertants of mutations affecting cilia biogenesis and the cell cycle also will be isolated and characterized. A detailed analysis of alpha- and beta-tubulin gene organization and expression will be performed. By isolating and characterizing a large number of normal and mutant genes important in both cilia biogenesis and cell cycle regulation, it should be possible to elucidate mechanisms underlying both normal cytoskeletal function and cell cycle progression, events that frequently are abnormal in neoplastic cells.