title: Cell Migration and Invadopodia Formation Require a Membrane-binding Domain of CARMIL2.
abstract: CARMILs regulate capping protein (CP), a critical determinant of actin assembly and actin-based cell motility. Vertebrates have three conserved CARMIL genes with distinct functions. In migrating cells, CARMIL2 is important for cell polarity, lamellipodial assembly, ruffling, and macropinocytosis. In cells, CARMIL2 localizes with a distinctive dual pattern to vimentin intermediate filaments and to membranes at leading edges and macropinosomes. The mechanism by which CARMIL2 localizes to membranes has not been defined. Here, we report that CARMIL2 has a conserved membrane-binding domain composed of basic and hydrophobic residues, which is necessary and sufficient for membrane localization, based on expression studies in cells and on direct binding of purified protein to lipids. Most important, we find that the membrane-binding domain is necessary for CARMIL2 to function in cells, based on rescue expression with a set of biochemically defined mutants. CARMIL1 and CARMIL3 contain similar membrane-binding domains, based on sequence analysis and on experiments, but other CPI motif proteins, such as CD2AP, do not. Based on these results, we propose a model in which the membrane-binding domain of CARMIL2 tethers this multidomain protein to the membrane, where it links dynamic vimentin filaments with regulation of actin assembly via CP.
authors.affiliation: From the Department of Cell Biology and Physiology, Washington University, St. Louis, Missouri 63110.
authors.forename: M Hunter
authors.initials: MH
authors.lastname: Lanier
authors.affiliation: From the Department of Cell Biology and Physiology, Washington University, St. Louis, Missouri 63110.
authors.forename: Patrick
authors.initials: P
authors.lastname: McConnell
authors.affiliation: From the Department of Cell Biology and Physiology, Washington University, St. Louis, Missouri 63110 jacooper@wustl.edu.
authors.forename: John A
authors.initials: JA
authors.lastname: Cooper
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D002465:Cell Movement; D017124:Conserved Sequence; D003598:Cytoskeletal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D030562:Databases, Protein; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D007382:Intermediate Filaments / Q000378:metabolism*; D008081:Liposomes; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008840:Microfilament Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D010446:Peptide Fragments / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000069261:Podosomes / Q000378:metabolism*; D054730:Protein Interaction Domains and Motifs; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport; D034622:RNA Interference; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D014746:Vimentin / Q000378:metabolism*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C547525:CARMIL1 protein, human; C114545:CD2-associated protein; D003598:Cytoskeletal Proteins; D008081:Liposomes; D008164:Luminescent Proteins; D008840:Microfilament Proteins; D010446:Peptide Fragments; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; D014746:Vimentin
keywords: RLTPR; actin; capping protein; cell migration; cell polarity; invadopodia; lamellipodia; lipid binding; plasma membrane; vimentin