MOLECULAR DETERMINANTS OF CYTOSKELETAL ORGANIZATION

细胞骨架组织的分子决定因素

• 批准号: 3292055
• 负责人: JONATHAN G IZANT
• 金额: $ 16.38万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-07-01 至 1991-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3292055
• 关键词: Xenopus; actins; chick embryo; cytoskeleton; developmental genetics; eukaryote; gene expression; gene mutation; gene therapy; genetic library; genetic manipulation; genetic recombination; messenger RNA; microinjections; microtubule associated protein; molecular cloning; nonmammalian vertebrate embryology; plasmids; tissue /cell culture; tropomyosin

Cytoskeletal architecture is a predominant and characteristic feature of eukaryotic cell identity and function. The goal of this project is to study the activity of genes that coordinate cytoskeletal organization in cultured cells and amphibian embryos in vivo using recombinant DNA vectors including anti-sense plasmids. Anti-sense gene inhibition is a valuable molecular strategy for phenocopying gene mutations in vertebrate organisms that are not amenable to classical genetic analysis. Studies on the mechanism of DNA mediated "flipped" gene anti-sense inhibition will continue. Anti-sense chicken thymidine kinase intron and anti-sense intron-exon junction plasmids will be constructed and tested by comicroinjection with the wild type gene into cultured cell nuclei and Xenopus oocyte germinal vesicles. Gene product activity and target mRNA transcript metabolism will be carefully monitored. Related strategies for gene replacement will be explored. Anti-sense and sense plasmids containing several cytoskeleton-associated protein genes (tropomyosins, yield-actinin, and a 200 kd microtubule-associated protein) will be constructed and introduced into cultured cells and Xenopus embryos. The resultant deficiencies in cellular architecture will be studied to assess the contribution that each gene makes to cellular morphology, viral transformation, and embryonic morphogenesis. Xenopus cDNA library inserts, enriched in embryo-specific sequences, will be subcloned into anti-sense expression vectors and then introduced into Xenopus embryos to identify novel genes that direct embryonic pattern formation. The objective is to use anti-sense gene inhibition as a pseudo-genetic tool to identify previously uncharacterized factors that regulated cell organization in addition to using these molecular approaches for the analysis of cloned gene function in living cells.

细胞骨架结构是细胞的主要特征