title: Kindlin binds migfilin tandem LIM domains and regulates migfilin focal adhesion localization and recruitment dynamics.
abstract: Focal adhesions (FAs), sites of tight adhesion to the extracellular matrix, are composed of clusters of transmembrane integrin adhesion receptors and intracellular proteins that link integrins to the actin cytoskeleton and signaling pathways. Two integrin-binding proteins present in FAs, kindlin-1 and kindlin-2, are important for integrin activation, FA formation, and signaling. Migfilin, originally identified in a yeast two-hybrid screen for kindlin-2-interacting proteins, is a LIM domain-containing adaptor protein found in FAs and implicated in control of cell adhesion, spreading, and migration. By binding filamin, migfilin provides a link between kindlin and the actin cytoskeleton. Here, using a combination of kindlin knockdown, biochemical pulldown assays, fluorescence microscopy, fluorescence resonance energy transfer (FRET), and fluorescence recovery after photobleaching (FRAP), we have established that the C-terminal LIM domains of migfilin dictate its FA localization, shown that these domains mediate an interaction with kindlin in vitro and in cells, and demonstrated that kindlin is important for normal migfilin dynamics in cells. We also show that when the C-terminal LIM domain region is deleted, then the N-terminal filamin-binding region of the protein, which is capable of targeting migfilin to actin-rich stress fibers, is the predominant driver of migfilin localization. Our work details a correlation between migfilin domains that drive kindlin binding and those that drive FA localization as well as a kindlin dependence on migfilin FA recruitment and mobility. We therefore suggest that the kindlin interaction with migfilin LIM domains drives migfilin FA recruitment, localization, and mobility.
authors.affiliation: From the Departments of Cell Biology and.
authors.forename: Nina N
authors.initials: NN
authors.lastname: Brahme
authors.forename: David S
authors.initials: DS
authors.lastname: Harburger
authors.forename: Karl
authors.initials: K
authors.lastname: Kemp-O'Brien
authors.forename: Rachel
authors.initials: R
authors.lastname: Stewart
authors.forename: Srikala
authors.initials: S
authors.lastname: Raghavan
authors.forename: Maddy
authors.initials: M
authors.lastname: Parsons
authors.forename: David A
authors.initials: DA
authors.lastname: Calderwood
mesh_terms: D000818:Animals; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D002478:Cells, Cultured; D003598:Cytoskeletal Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D022001:Focal Adhesions / Q000378:metabolism*; D055786:Gene Knockout Techniques; D006801:Humans; D015603:Keratinocytes / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D009124:Muscle Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D041681:NIH 3T3 Cells; D009363:Neoplasm Proteins / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary
chemical_list: D015815:Cell Adhesion Molecules; D003598:Cytoskeletal Proteins; C475301:FERMT1 protein, human; C489422:Fblim1 protein, mouse; D008565:Membrane Proteins; D009124:Muscle Proteins; D009363:Neoplasm Proteins; C527802:kindlin-2 protein, mouse
keywords: Adhesion; Cell Adhesion; Cytoskeleton; Fluorescence Resonance Energy Transfer (FRET); Focal Adhesion; Integrin; Kindlin; LIM Domain; Migfilin