Identification of a filamin isoform enriched at the ends of stress fibers in chicken embryo fibroblasts.
Filamin (actin-binding protein) is a cytoskeletal protein that crosslinks actin filaments in vitro. Filamin is thought to be involved in a variety of cell types in stabilizing actin networks, and in platelets it may play a role in linking actin to the membrane. In this report, we describe a monoclonal antibody (Mab 6E) that was used to immunoprecipitate an isoform of filamin from extracts of chicken fibroblasts revealed an unusual pattern: while other filamin antibodies stained the entire length of stress fibers, the Mab 6E staining was predominantly at the ends of stress fibers. In double-labeling experiments, the distribution of the Mab 6E antigen was found to be strikingly similar to that of alpha-actinin. Mab 6E staining was associated, in part, with focal adhesions, which are sites of actin-membrane attachment. Unlike other focal adhesion proteins, such as vinculin and talin, this filamin isoform is apparently not localized evenly throughout the entire area of adhesion, being absent from or greatly reduced in the distal portion of the area. The Mab 6E antigen was identified as filamin by immunological crossreactivity with a panel of antifilamin monoclonals as well as with a polyclonal anti-filamin. The Mab 6E isoform, however, was found to differ from the major form of filamin both by one-dimensional peptide analysis and slightly slower migration on SDS-containing gels. The Mab 6E antigen was also detected by immunofluorescence in the Z-lines of isolated adult myofibrils. These results suggest that chicken fibroblasts may express different isoforms of filamin that could have specialized roles within the cell.
Pavalko
F M
FM
Department of Cell Biology and Anatomy, University of North Carolina, Chapel Hill 27599.
Otey
C A
CA
Burridge
K
K
D000185:Actinin / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D000818:Animals; D000911:Antibodies, Monoclonal; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002642:Chick Embryo; D003285:Contractile Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D064448:Filamins; D005455:Fluorescent Antibody Technique; D008840:Microfilament Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*
D000199:Actins; D000911:Antibodies, Monoclonal; D003285:Contractile Proteins; D064448:Filamins; D008840:Microfilament Proteins; D000185:Actinin