title: Identification of novel integrin binding partners for calcium and integrin binding protein 1 (CIB1): structural and thermodynamic basis of CIB1 promiscuity.
abstract: The short cytoplasmic tails of the α- and β-chains of integrin adhesion receptors regulate integrin activation and cell signaling. Significantly less is known about proteins that bind to α-integrin cytoplasmic tails (CTs) as opposed to β-CTs to regulate integrins. Calcium and integrin binding protein 1 (CIB1) was previously identified as an αIIb binding partner that inhibits agonist-induced activation of the platelet-specific integrin, αIIbβ3. A sequence alignment of all α-integrin CTs revealed that key residues in the CIB1 binding site of αIIb are well-conserved, and was used to delineate a consensus binding site (I/L-x-x-x-L/M-W/Y-K-x-G-F-F). Because the CIB1 binding site of αIIb is conserved in all α-integrins and CIB1 expression is ubiquitous, we asked if CIB1 could interact with other α-integrin CTs. We predicted that multiple α-integrin CTs were capable of binding to the same hydrophobic binding pocket on CIB1 with docking models generated by all-atom replica exchange discrete molecular dynamics. After demonstrating novel in vivo interactions between CIB1 and other whole integrin complexes with co-immunoprecipitations, we validated the modeled predictions with solid-phase competitive binding assays, which showed that other α-integrin CTs compete with the αIIb CT for binding to CIB1 in vitro. Isothermal titration calorimetry measurements indicated that this binding is driven by hydrophobic interactions and depends on residues in the CIB1 consensus binding site. These new mechanistic details of CIB1-integrin binding imply that CIB1 could bind to all integrin complexes and act as a broad regulator of integrin function.
authors.affiliation: Department of Biochemistry and Biophysics, ‡Lineberger Comprehensive Cancer Center, and §McAllister Heart Institute, School of Medicine, University of North Carolina at Chapel Hill , Chapel Hill, North Carolina 27599, United States.
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Freeman
authors.forename: Justin L
authors.initials: JL
authors.lastname: Black
authors.forename: Holly G
authors.initials: HG
authors.lastname: Bray
authors.forename: Onur
authors.initials: O
authors.lastname: Dagliyan
authors.forename: Yi I
authors.initials: YI
authors.lastname: Wu
authors.forename: Ashutosh
authors.initials: A
authors.lastname: Tripathy
authors.forename: Nikolay V
authors.initials: NV
authors.lastname: Dokholyan
authors.forename: Tina M
authors.initials: TM
authors.lastname: Leisner
authors.forename: Leslie V
authors.initials: LV
authors.lastname: Parise
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D057927:Hydrophobic and Hydrophilic Interactions; D039001:Integrin alpha Chains / Q000378:metabolism*; D056004:Molecular Dynamics Simulation; D040201:Platelet Membrane Glycoprotein IIb / Q000378:metabolism; D011485:Protein Binding; D016415:Sequence Alignment
chemical_list: C109198:CIB1 protein, human; D002135:Calcium-Binding Proteins; D039001:Integrin alpha Chains; D040201:Platelet Membrane Glycoprotein IIb