RAS GENES AND REGULATION OF CELLULAR MORPHOGENESIS

本项目将研究ras蛋白在 调节真核细胞的生长和发育。 丝状 真菌，总状毛霉，被诱导经历三种特征类型 对生长条件变化的反应， 营养物质或气体环境，并提供了一个简单的模型系统， 真核细胞的发展，这是经得起生化和遗传 分析. 3种毛霉ras基因（MRAS）的累积mRNA水平 在细胞发育的几个不同阶段进行了分析。 两 基因MRAS 1和MRAS 3在细胞生长过程中差异表达， 这种真菌的发展表明这些基因在 发展过程。 例如，MRAS3基因表达几乎 仅在无性孢子形成期间，而MRAS 1在最高表达， 在"快速"分裂的菌丝细胞或转化为菌丝的细胞中的水平。 MRAS 1和MRAS 3基因具有独特的表达时间模式， 可能表明它们在生长和发育中起着不同的作用。 有机体 基于核苷酸序列分析， MRAS基因在氨基酸序列和大小上与H- ras基因的调控作用 细胞的生长和分化。 由于这种蛋白质的相似性， ras基因在细胞调控中的作用 这些简单的真核细胞的分化可能导致重要的 深入了解细胞增殖、分化和功能障碍， 发生在哺乳动物肿瘤中的过程。 本研究建议的具体目标是：1）表征 ras基因家族（MRAS）在M.总状花序; 2）改变 ras基因在M.使用MRAS表达的总状体 载体; 3）表征改变的MRAS表达对 形态发生、细胞增殖和生化变化， 伴随着这些过程在M.总状花序。 正常的ras表达和蛋白活性将通过转化 具有设计用于改变MRAS蛋白的质粒构建体的毛霉细胞 表达的结构、数量和时间模式。 直接 观察将检测形态发生反应的表型变化，或 生长率，而生化测定将用于测量的影响 ras表达改变对cAMP代谢和/或磷酸肌醇 新陈代谢.

This project will investigate the proposed role of ras proteins in regulation of growth and development of eukaryotic cells. The filamentous fungus, Mucor racemosus, is induced to undergo three characteristic types of cellular morphogenesis in response to changes in growth conditions, nutrients, or gaseous environment and provides a simple model system of eukaryotic cell development which is amenable to biochemical and genetic analysis. The accumulated mRNA levels of 3 Mucor ras genes (MRAS) have been analyzed during several different stages of cell development. Two of the genes, MRAS1 and MRAS3 are differentially expressed during cell development in this fungus suggesting that these genes play a role in the developmental processes. The MRAS3 gene for example is expressed almost exclusively during asexual sporulation, while MRAS1 is expressed at highest levels in "rapidly" dividing hyphal cells or cells converting to hyphae. The MRAS1 and MRAS3 genes have unique temporal patterns of expression which may suggest that they play different roles in growth and development of the organism. Based on nucleotide sequence analyses, the proteins encoded by the MRAS genes are highly similar in amino acid sequence and size to the H- ras gene of higher eukaryotes which has been implicated in the regulation of cell growth and differentiation. Because of this protein similarity, understanding of ras function in regard to regulation of cell differentiation in these simple eukaryotic cells may lead to important insights into cell proliferation, differentiation and malfunctions in these processes which occur in mammalian tumors. The specific aims of this research proposal are: 1) to characterize the expression of the ras gene family (MRAS) in M. racemosus; 2) to alter the normal expression patter of ras genes in M. racemosos using MRAS expression vectors; 3) to characterize the effects of altered MRAS expression on morphogenesis, cell proliferation, and the biochemical changes which accompany these processes in M. racemosus. Normal ras expression and protein activity will be altered by transforming Mucor cells with plasmid constructs designed to alter MRAS protein structure, quantity, and temporal pattern of expression. Direct observation will detect phenotypic changes in the morphogenetic response or growth rates, while biochemical assays will be used to measure the effects of altered ras expression on cAMP metabolism and/or phosphoinositid metabolism.