Cellular Functions of the Human Filamins

点击翻译按钮获取中文摘要

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.