Cellular Functions of the Human Filamins

人类细丝蛋白的细胞功能

• 批准号: 6829908
• 负责人: SANDOR S SHAPIRO
• 金额: $ 39.25万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-15 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6829908
• 关键词: RNA interference; actin binding protein; actins; binding sites; cell biology; clinical research; cytoskeleton; gene expression; gene mutation; gene targeting; human subject; laboratory mouse; melanoma; microarray technology; microfilaments; neoplastic cell; phenotype; phosphatidylinositols; protein binding; protein isoforms; protein structure function; shear stress; yeast two hybrid system; zebrafish

DESCRIPTION (provided by applicant): Platelet shape change and endothelial and vascular smooth muscle migration are central to the processes of hemostasis and thrombosis, angiogenesis and wound healing. Rapid alterations in cytoskeletal structure are required for these responses. Filamins (FLNs) are important in modulating these responses. The human FLN family consists of three approximtely 280 kD paralogs -A, B and C - each containing a approximately 250 residue N-terminal actin-binding domain (ABD) followed by 24 internally homologous repeats. FLNs induce formation of three-dimensional actin networks and connect the cell membrane to the cytoskeleton by binding to the cytoplasmic tails of several integrins and other membrane proteins. FLNs also bind to a number of cellular components, including several small GTPases, and may serve as scaffolds on which intracellular signaling and protein trafficking pathways are organized; these interactions do not require the presence of an ABD. Cell lines lacking FLNA are abnormal in shape and migration and mutations in FLNA, either in the ABD or in the repeats, are associated with human disease. Very recently, our collaborators and we have found that mutations in FLNB also lead to human diseases whose phenotypes differ from those seen with FLNA mutations. We have also found that knock-down of zebrafish FLNB causes major embryologic abnormalities. Furthermore, using the yeast two-hybrid technique we have identified several new FLNB binding partners with varying FLN-binding characteristics, including Rab22B, that does not bind to FLNA, fibrillin-1, that does not bind to FLNs A or C, and a new proline-rich-domain and LIM-domain protein that we have named FBLP-1 (Filamin-Binding LIM Protein-I), that binds to all three FLNs. Thus it is almost certain that, although FLNs may share some functions, each FLN has unique functional characteristics. The aims of this proposal are: 1) To delineate the roles of the FLN ABD vs. the repeats, in FLN function in vitro; 2) To determine in vivo specificity of FLN functions by a) knockdown of FLNs in zebrafish and b) knockout of FLNB in the mouse (knockout of FLNA has been achieved by others, but not yet published); and 3) To further delineate the function of our newly identified FBLP-1.

描述（由申请方提供）：血小板形状变化以及内皮和血管平滑肌迁移是止血和血栓形成、血管生成和伤口愈合过程的核心。这些反应需要细胞骨架结构的快速改变。丝状神经元（FLN）在调节这些反应中很重要。人FLN家族由三个约280 kD的旁系同源物-A、B和C -组成，每个旁系同源物含有约250个残基的N-末端肌动蛋白结合结构域（ABD），随后是24个内部同源重复。FLN诱导三维肌动蛋白网络的形成，并通过与几种整合素和其他膜蛋白的胞质尾部结合将细胞膜连接到细胞骨架。FLN还与许多细胞组分结合，包括几种小的GTP酶，并且可以作为细胞内信号传导和蛋白质运输途径组织的支架;这些相互作用不需要ABD的存在。缺乏FLNA的细胞系在形状和迁移方面是异常的，并且FLNA中的突变，无论是在ABD中还是在重复序列中，都与人类疾病相关。最近，我们的合作者和我们发现FLNB突变也会导致人类疾病，其表型与FLNA突变不同。我们还发现，敲低斑马鱼FLNB会导致主要的胚胎学异常。此外，使用酵母双杂交技术，我们已经确定了几个新的FLNB结合伴侣与不同的FLN结合特性，包括Rab 22 B，不结合FLN的FLNA，replanin-1，不结合FLN A或C，和一个新的富含脯氨酸的结构域和LIM结构域的蛋白，我们命名为FBLP-1（Filamin-Binding LIM Protein-I），结合所有三个FLN。因此，几乎可以肯定的是，虽然FLN可能共享一些功能，但每个FLN都具有独特的功能特征。该提议的目的是：1）描述FLN ABD相对于重复序列在体外FLN功能中的作用; 2）通过a）在斑马鱼中敲除FLN和B）在小鼠中敲除FLNB来确定FLN功能的体内特异性（FLNA的敲除已由其他人实现，但尚未发表）;和3）进一步描述我们新鉴定的FBLP-1的功能。