MORPHOGENESIS AND GENE EXPRESSION

形态发生和基因表达

• 批准号: 3274288
• 负责人: SUMIKO INOUYE
• 金额: $ 21.99万
• 依托单位: UNIV OF MED/DENT NJ-R W JOHNSON MED SCH
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-04-01 至 1991-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3274288
• 关键词: Escherichia coli; Myxococcus; bacterial DNA; bacterial genetics; cell free system; developmental genetics; gene expression; genetic manipulation; genetic regulation; growth /development; membrane proteins; membrane structure; molecular biology; nucleic acid hybridization; plasmids; protein S; protein biosynthesis; protein engineering; protein sequence; protein structure

Myxococcus xanthus is an excellent prokaryotic system for studying the regulation of development. M. xanthus show a developmental cycle which is similar to the eukaryotic slime molds such as Dictyostelium discoideum. However, there are several distinct advantages to studying development in M. xanthus, since it is a simple gram-negative bacterium. We propose to study morphogenesis and the molecular mechanisms of control of gene expression during development using M xanthus as a model system. We will continue to work on the on-going projects such as protein S, a development-specific protein, and its gene. We will also attempt to explore several new aspects of M. xanthus, including characterization of multicopy single-stranded DNA (msDNA), a unique DNA found in myxobacteria, the use of anti-sense RNA (micRNA) for characterization of developmental genes, and identification and characterization of specific organelle(s) required for gliding motility. The following are specific aims which we will pursue in this proposal: (1) Development-specific Genes. Two tandemly repeated, homologous genes, ops (protein S-1) and tps (protein S) are differentially expressed during development. Using oligonucleotide-directed site-specific mutagenesis, we will elucidate the exact nucleotide sequences in their promoters which are responsible for the regulation. We will develop a micRNA cloning vector for study of the regulatory mechanisms and the functions of developmental genes isolated. We will also explore cloning the genes for sigma factors for developmental genes. (2) Functions of Protein S and Protein S-1. Protein S-1 will be purified and crystallized, and its biochemical properties will be compared with protein S. Roles of protein S will be further investigated by creating mutations at putative Ca++ binding sites. (3) msDNA. We will determine the RNA primer structures of both msDNAs, from M. xanthus and Sigmatella aurantiaca, and attempt to elucidate the molecular mechanisms of the biosynthesis of msDNA and its role in myxobacteria. (4) Organelles for Gliding Motility. We have isolated microtubule-like structures from M. xanthus. We will characterize these components from biochemical as well as functional aspects. (5) Integration-type Plasmids. We will construct plasmids which facilitate P1 specialized transduction to integrate any genes (or DNA fragments) of interest into the specific site of the M. xanthus chromosome.

粘粒叶thus是一个出色的核系统，用于研究 调节发展。 M. Xanthus显示出一个发育周期 类似于真核粘液胶模具，例如迪斯特尔迪斯特尔。 但是，研究发展的发展有几个不同的优势 M. Xanthus，因为它是一种简单的革兰氏阴性细菌。 我们建议 研究形态发生和基因控制的分子机制 在开发过程中，使用M Xanthus作为模型系统。 我们将 继续从事正在进行的项目，例如蛋白质S，a 发育特异性蛋白质及其基因。 我们还将尝试 探索M. Xanthus的几个新方面，包括表征 多拷贝单链DNA（MSDNA），一种在粘菌病中发现的独特DNA， 使用抗敏感RNA（micro）来表征发育 基因以及特定细胞器的鉴定和表征 滑行运动所需。 以下是我们的特定目标 将在此提案中追求：（1）特定于发展的基因。 二 串联重复，同源基因，OPS（蛋白质S-1）和TPS（蛋白质S） 在开发过程中差异表达。 使用 寡核苷酸定向位点特异性诱变，我们将阐明 其启动子中的确切核苷酸序列负责 规定。 我们将开发一个微NA克隆向量，以研究 调节机制和分离的发育基因的功能。 我们还将探索用于发育的Sigma因子的克隆基因 基因。 （2）蛋白质S和蛋白质S-1的功能。 蛋白质S-1将是 纯化和结晶，将比较其生化特性 蛋白质S.蛋白质的作用将通过 在推定的CA ++结合位点创建突变。 （3）MSDNA。 我们将 确定来自M. Xanthus和M. Xanthus和 Sigmatella aurantiaca，并试图阐明 MSDNA的生物合成及其在粘细菌中的作用。 （4）细胞器 滑行运动。 我们有来自M的分离的微管样结构。 Xanthus。 我们将从生化和 功能方面。 （5）整合型质粒。 我们将构建 促进P1专用转导的质粒以整合任何 感兴趣的基因（或DNA片段）进入M的特定位点 黄质染色体。