title: A new focal adhesion protein that interacts with integrin-linked kinase and regulates cell adhesion and spreading.
abstract: Integrin-linked kinase (ILK) is a multidomain focal adhesion (FA) protein that functions as an important regulator of integrin-mediated processes. We report here the identification and characterization of a new calponin homology (CH) domain-containing ILK-binding protein (CH-ILKBP). CH-ILKBP is widely expressed and highly conserved among different organisms from nematodes to human. CH-ILKBP interacts with ILK in vitro and in vivo, and the ILK COOH-terminal domain and the CH-ILKBP CH2 domain mediate the interaction. CH-ILKBP, ILK, and PINCH, a FA protein that binds the NH(2)-terminal domain of ILK, form a complex in cells. Using multiple approaches (epitope-tagged CH-ILKBP, monoclonal anti-CH-ILKBP antibodies, and green fluorescent protein-CH-ILKBP), we demonstrate that CH-ILKBP localizes to FAs and associates with the cytoskeleton. Deletion of the ILK-binding CH2 domain abolished the ability of CH-ILKBP to localize to FAs. Furthermore, the CH2 domain alone is sufficient for FA targeting, and a point mutation that inhibits the ILK-binding impaired the FA localization of CH-ILKBP. Thus, the CH2 domain, through its interaction with ILK, mediates the FA localization of CH-ILKBP. Finally, we show that overexpression of the ILK-binding CH2 fragment or the ILK-binding defective point mutant inhibited cell adhesion and spreading. These findings reveal a novel CH-ILKBP-ILK-PINCH complex and provide important evidence for a crucial role of this complex in the regulation of cell adhesion and cytoskeleton organization.
authors.affiliation: Department of Pathology, University of Pittsburgh, Pittsburgh, Pennsylvania 15261, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Tu
authors.forename: Y
authors.initials: Y
authors.lastname: Huang
authors.forename: Y
authors.initials: Y
authors.lastname: Zhang
authors.forename: Y
authors.initials: Y
authors.lastname: Hua
authors.forename: C
authors.initials: C
authors.lastname: Wu
mesh_terms: D000185:Actinin; D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002448:Cell Adhesion; D002451:Cell Compartmentation; D048429:Cell Size; D006224:Cricetinae; D003599:Cytoskeleton / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000378:metabolism; D022001:Focal Adhesions / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D060588:LIM Domain Proteins; D008565:Membrane Proteins; D008840:Microfilament Proteins; D008969:Molecular Sequence Data; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques; D000096985:Calponins; D000099200:Scaffold Protein ILK
chemical_list: D000185:Actinin; D048868:Adaptor Proteins, Signal Transducing; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D060588:LIM Domain Proteins; D008565:Membrane Proteins; D008840:Microfilament Proteins; D017346:Protein Serine-Threonine Kinases; D000096985:Calponins; D000099200:Scaffold Protein ILK; C088088:LIMS1 protein, human; C486590:PARVA protein, human; C486591:PARVB protein, human