ANALYSIS OF RAT NONMUSCLE MYOSIN HEAVY CHAIN GENE

大鼠非肌肉肌球蛋白重链基因分析

• 批准号: 3082769
• 负责人: LUCY W ARNOLD
• 金额: $ 6.35万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-04-01 至 1995-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3082769
• 关键词: cell motility; complementary DNA; electroporation; gel electrophoresis; gene expression; genetic mapping; genetic recombination; genome; laboratory mouse; laboratory rat; microinjections; molecular cloning; myosins; natural gene amplification; nucleic acid hybridization; nucleic acid probes; nucleic acid sequence; polymerase chain reaction; protein biosynthesis; protein sequence; protein structure function; sarcomeres; site directed mutagenesis

Nonmuscle (nm) myosin is an essential part of the cytoskeleton, a complex intracellular network that orchestrates such cell functions as cytokinesis, motility, and muscle contraction. Recent work has demonstrated that dictyostelium lacking myosin heavy chain (mhc) are viable, motile, and phagocytic, although they demonstrate markedly abnormal cytokinesis. In dividing mammalian cells, nm myosin localizes to the cleavage furrow during cytokinesis; labeled myosin light chains microinjected into living cells localize in the cleavage furrow, and become reincorporated into stress fibers at the completion of cytokinesis. Nm myosin may also be necessary for sarcomere assembly, because the cytoskeleton must act as a scaffolding for synthesis of the sarcomeric contractile proteins to occur by cotranslational assembly. Exciting advances in gene targeting now make it possible to target nonselectable genes by homologous recombination. Preliminary data suggests that the nm mhc gene is encoded by one or a few genes making such gene targeting feasible. The primary goal of this project is to complete the cloning and characterization of the nm mhc gene, in order to begin gene disruption experiments by homologous recombination. By disrupting the nm mhc in mouse non and muscle cells, we should be able to elucidate the role of nm mhc in cytokinesis, cell motility and sarcomere assembly. Further studies will focus on identifying those gene domains that confer specific functions to the nm mhc protein in both muscle and nonmuscle.

非肌肉肌球蛋白是细胞骨架的重要组成部分， 细胞内网络协调细胞功能如胞质分裂， 运动和肌肉收缩。 最近的研究表明， 缺乏肌球蛋白重链（MHC）的网骨藻是有活力的、能动的， 吞噬细胞，虽然他们表现出显着异常胞质分裂。 在 在哺乳动物细胞分裂过程中，nm肌球蛋白定位于卵裂沟。 胞质分裂;显微注射到活细胞中的标记肌球蛋白轻链 定位在卵裂沟中，并重新整合到应力中 纤维在胞质分裂完成。 Nm肌球蛋白也可能是必要的 因为细胞骨架必须充当支架， 为了合成肌节收缩蛋白， 共翻译组装 令人兴奋的进展，基因靶向，现在有可能靶向 非选择性基因通过同源重组。 初步数据提示 nm MHC基因由一个或几个基因编码， 目标可行。 该项目的主要目标是完成 为了开始基因的克隆和表征， 通过同源重组进行破坏实验。 通过扰乱纳米 mhc在小鼠非和肌肉细胞中的作用，我们应该能够阐明 nmhc在胞质分裂、细胞运动和肌节组装中的作用。 进一步 研究将集中在确定那些基因域，赋予特定的 在肌肉和非肌肉中对nmmhc蛋白起作用。