Mammalian homolog of the yeast cyclase associated protein, CAP/Srv2p, regulates actin filament assembly

Control of cell shape and motility requires rearrangements of the actin cyt oskeleton. One cytoskeletal protein that may regulate actin dynamics is CAP (cyclase associated protein; CAP/Srv2p; ASP-56). CAP was first isolated fr om yeast as an adenylyl cyclase associated protein required for RAS regulat ion of cAMP signaling. In addition, CAP also regulates the actin cytoskelet on primarily through an actin monomer binding activity. CAP homologs are fo und in many eukaryotes, including mammals where they also bind actin, but l ittle is known about their biological function. We, therefore, designed exp eriments to address CAP1 regulation of the actin cytoskeleton. CAP1 localiz ed to membrane ruffles and actin stress fibers in fixed cells of various ty pes. To address localization in living cells, we constructed GFP-CAP1 fusio n proteins and found that fusion proteins lacking the actin-binding region localized like the wild type protein. We also performed microinjection stud ies with affinity-purified anti-CAP1 antibodies in Swiss 3T3 fibroblasts an d found that the antibodies attenuated serum stimulation of stress fibers. Finally, CAP1 purified from platelets through a monoclonal antibody affinit y purification step stimulated the formation of stress fiber-like filaments when it was microinjected into serum-starved Swiss 3T3 cells. Taken togeth er, these data suggest that CAP1 promotes assembly of the actin cytoskeleto n.Cell Motil. (C) 2000 Wiley-Liss, Inc.