title: Role of the C-terminal Extension of Formin 2 in Its Activation by Spire Protein and Processive Assembly of Actin Filaments.
abstract: Formin 2 (Fmn2), a member of the FMN family of formins, plays an important role in early development. This formin cooperates with profilin and Spire, a WASP homology domain 2 (WH2) repeat protein, to stimulate assembly of a dynamic cytoplasmic actin meshwork that facilitates translocation of the meiotic spindle in asymmetric division of mouse oocytes. The kinase-like non-catalytic domain (KIND) of Spire directly interacts with the C-terminal extension of the formin homology domain 2 (FH2) domain of Fmn2, called FSI. This direct interaction is required for the synergy between the two proteins in actin assembly. We have recently demonstrated how Spire, which caps barbed ends via its WH2 domains, activates Fmn2. Fmn2 by itself associates very poorly to filament barbed ends but is rapidly recruited to Spire-capped barbed ends via the KIND domain, and it subsequently displaces Spire from the barbed end to elicit rapid processive assembly from profilin·actin. Here, we address the mechanism by which Spire and Fmn2 compete at barbed ends and the role of FSI in orchestrating this competition as well as in the processivity of Fmn2. We have combined microcalorimetric, fluorescence, and hydrodynamic binding assays, as well as bulk solution and single filament measurements of actin assembly, to show that removal of FSI converts Fmn2 into a Capping Protein. This activity is mimicked by association of KIND to Fmn2. In addition, FSI binds actin at filament barbed ends as a weak capper and plays a role in displacing the WH2 domains of Spire from actin, thus allowing the association of actin-binding regions of FH2 to the barbed end.
authors.affiliation: From the Cytoskeleton Dynamics and Motility, Institut de Biologie Intégrative de la Cellule, CNRS, 91198 Gif-sur-Yvette, France.
authors.forename: Pierre
authors.initials: P
authors.lastname: Montaville
authors.affiliation: From the Cytoskeleton Dynamics and Motility, Institut de Biologie Intégrative de la Cellule, CNRS, 91198 Gif-sur-Yvette, France.
authors.forename: Sonja
authors.initials: S
authors.lastname: Kühn
authors.affiliation: From the Cytoskeleton Dynamics and Motility, Institut de Biologie Intégrative de la Cellule, CNRS, 91198 Gif-sur-Yvette, France.
authors.forename: Christel
authors.initials: C
authors.lastname: Compper
authors.affiliation: From the Cytoskeleton Dynamics and Motility, Institut de Biologie Intégrative de la Cellule, CNRS, 91198 Gif-sur-Yvette, France marie-france.carlier@i2bc.paris-saclay.fr.
authors.forename: Marie-France
authors.initials: MF
authors.lastname: Carlier
mesh_terms: D008841:Actin Cytoskeleton / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D001667:Binding, Competitive; D017124:Conserved Sequence; D000080865:Formins; D017353:Gene Deletion; D006801:Humans; D007700:Kinetics; D051379:Mice; D008840:Microfilament Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008970:Molecular Weight; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D054730:Protein Interaction Domains and Motifs; D055503:Protein Multimerization; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
chemical_list: D000080865:Formins; D008840:Microfilament Proteins; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C000592292:SPIRE1 protein, human; C549436:formin 2 protein, mouse
keywords: actin; fluorescence anisotropy; fluorescence correlation spectroscopy (FCS); formin; processivity; profilin