MEMBRANE-MICROFILAMENT CONNECTIONS

膜-微丝连接

• 批准号: 3282056
• 负责人: JOHN R GLENNEY
• 金额: $ 20.42万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-05-01 至 1991-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3282056
• 关键词: annexins; binding proteins; calcium; calmodulin; cell adhesion; cell membrane; chickens; cytoskeleton; electron microscopy; fluorescence microscopy; guinea pigs; immunoprecipitation; laboratory mouse; laboratory rabbit; membrane proteins; microfilaments; monoclonal antibody; neoplastic transformation; phosphorylation; protein structure; proteolysis; radioimmunoassay

The goal of this project is the determination of the mechanism and result of connections between the microfilament system and the plasma membrane. Previously we had analyzed the system of non-erythroid spectrins as a model for this subcortical network. We now propose to extend our research to the analysis of 36,000 Mr tyrosine kinase substrates termed calpactins. These proteins which exist as monomers or as complexes with a 10,000 Mr light chain interact with the cytoskeletal proteins actin and spectrin and are modulated by Ca++ and phospholipid. We will identify other members of this family using a technique which relies on its ability to interact with actin and phospholipid, and a detailed comparison of the structure and function of these proteins will be made. A detailed analysis of Ca++-binding, including affinity modulators will be undertaken. We will analyze the functional relatedness of the light chain with the S-100 proteins. Monoclonal and polyclonal antibodies will be raised to defined regions (peptides) of calpactin for use in analysis of its function. The tissue and subcellular distribution using Western Blotting and immunofluorescence microscopy. The in vivo target of calpactins will be determined using cross-linking reagents followed by immunoprecipitation. The calpactin heavy and light chain genes will be expressed in bacteria and protein products will be purified and tested for biological activity. We will attempt to identify the region of calpactin needed for Ca++-binding. The effect of tyrosine phosphorylation of calpactin will be tested by determining whether there is any difference in the ability of phosphorylated calpactin to interact with phospholipid, Ca++ or spectrin. These studies will further our understanding of the subcortical filament network and its relationship to cell transformation.

本项目的目标是确定该机制

项目成果

Antibody probing of western blots which have been stained with india ink.

对用印度墨水染色的蛋白质印迹进行抗体探测。

• DOI: 10.1016/0003-2697(86)90259-9
• 发表时间: 1986
• 期刊: Analytical biochemistry
• 影响因子: 2.9
• 作者: Glenney,J

Association of the S-100-related calpactin I light chain with the NH2-terminal tail of the 36-kDa heavy chain.

S-100 相关钙肽 I 轻链与 36 kDa 重链 NH2 末端尾部的关联。

• 发表时间: 1986
• 期刊: The Journal of biological chemistry
• 作者: GlenneyJr,JR;Boudreau,M;Galyean,R;Hunter,T;Tack,B
• 通讯作者: Tack,B

Novel tyrosine kinase substrates from Rous sarcoma virus-transformed cells are present in the membrane skeleton.

• DOI: 10.1083/jcb.108.6.2401
• 发表时间: 1989-06
• 期刊: JOURNAL OF CELL BIOLOGY
• 影响因子: 7.8
• 作者: Glenney, J R Jr;Zokas, L
• 通讯作者: Zokas, L

Tyrosine phosphorylation of a 22-kDa protein is correlated with transformation by Rous sarcoma virus.

• DOI: 10.1016/s0021-9258(19)47038-5
• 发表时间: 1989-12
• 期刊: The Journal of biological chemistry
• 作者: J. Glenney

Paxillin: a new vinculin-binding protein present in focal adhesions.

• DOI: 10.1083/jcb.111.3.1059
• 发表时间: 1990-09
• 期刊: JOURNAL OF CELL BIOLOGY
• 影响因子: 7.8
• 作者: Turner, C E;Glenney, J R Jr;Burridge, K
• 通讯作者: Burridge, K