title: Targeting platelet-leukocyte interactions: identification of the integrin Mac-1 binding site for the platelet counter receptor glycoprotein Ibalpha.
abstract: The firm adhesion and transplatelet migration of leukocytes on vascular thrombus are dependent on the interaction of the leukocyte integrin Mac-1 (alphaMbeta2, CD11b/CD18) and the platelet counter receptor glycoprotein (GP) Ibalpha. Previous studies have established a central role for the I domain, a stretch of approximately 200 amino acids within the alphaM subunit, in the binding of GP Ibalpha. This study was undertaken to establish the molecular basis of GP Ibalpha recognition by alphaMbeta2. The P201-K217 sequence, which spans an exposed loop and amphipathic alpha4 helix in the three-dimensional structure of the alphaMI domain, was identified as the binding site for GP Ibalpha. Mutant cell lines in which the alphaMI domain segments P201-G207 and R208-K217 were switched to the homologous, but non-GP Ibalpha binding, alphaL domain segments failed to support adhesion to GP Ibalpha. Mutation of amino acid residues within P201-K217, H210-A212, T213-I215, and R216-K217 resulted in the loss of the binding function of the recombinant alphaMI domains to GP Ibalpha. Synthetic peptides duplicating the P201-K217, but not scrambled versions, directly bound GP Ibalpha and inhibited alphaMbeta2-dependent adhesion to GP Ibalpha and adherent platelets. Finally, grafting critical amino acids within the P201-K217 sequence onto alphaL, converted alphaLbeta2 into a GP Ibalpha binding integrin. Thus, the P201-K217 sequence within the alphaMI domain is necessary and sufficient for GP Ibalpha binding. These observations provide a molecular target for disrupting leukocyte-platelet complexes that promote vascular inflammation in thrombosis, atherosclerosis, and angioplasty-related restenosis.
authors.affiliation: Cardiovascular Division, Brigham and Women's Hospital, 75 Francis Street, Tower 3, Boston, MA 02115, USA.
authors.forename: Raila
authors.initials: R
authors.lastname: Ehlers
authors.forename: Valentin
authors.initials: V
authors.lastname: Ustinov
authors.forename: Zhiping
authors.initials: Z
authors.lastname: Chen
authors.forename: Xiaobin
authors.initials: X
authors.lastname: Zhang
authors.forename: Ravi
authors.initials: R
authors.lastname: Rao
authors.forename: F William
authors.initials: FW
authors.lastname: Luscinskas
authors.forename: Jose
authors.initials: J
authors.lastname: Lopez
authors.forename: Edward
authors.initials: E
authors.lastname: Plow
authors.forename: Daniel I
authors.initials: DI
authors.lastname: Simon
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D001792:Blood Platelets / Q000502:physiology*; D002448:Cell Adhesion; D002450:Cell Communication; D002460:Cell Line; D006801:Humans; D007962:Leukocytes / Q000502:physiology*; D016177:Macrophage-1 Antigen / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D016177:Macrophage-1 Antigen; D019038:Platelet Glycoprotein GPIb-IX Complex