title: Disruption of the integrin alphaLbeta2 transmembrane domain interface by beta2 Thr-686 mutation activates alphaLbeta2 and promotes micro-clustering of the alphaL subunits.
abstract: Integrins are type I heterodimeric cell adhesion molecules that mediate a wide array of biological processes. Integrin bidirectional signaling allows communication between the cell interior with its microenvironment. The integrin transmembrane domains (TMs) are the transducers of activation signal that is relayed from the cytoplasmic domains to the distal ligand binding site located in the ectodomain of the integrin and vice versa. In this study, we showed that the disruption of the alphaLbeta2 TMs by mutation of a key interface residue Thr-686 in the beta2 TM promoted alphaLbeta2 activation with ICAMs binding properties that are reminiscent of an intermediate affinity receptor. The activated alphaLbeta2 TM mutants, however, showed minimal reactivity with the reporter mAb KIM127 that recognizes a highly extended alphaLbeta2. Two models of alphaLbeta2 TM interaction were proposed previously. One with GXXXG-type interaction, and another that is based on TM cysteine-scanning analyses. Our data are consistent with a GXXXG-type interaction of the alphaLbeta2 TMs. Finally, we observed by FRET analyses that perturbation of the alphaLbeta2 TMs by beta2 Thr-686 mutation facilitated alphaL micro-cluster formation. This was diminished by linking the alphaLbeta2 TMs with a disulfide bond, which served to clasp the TMs. These data suggest that disruption of the TM interface changes alphaLbeta2 ligand binding affinity, and it may contribute to alphaL micro-cluster formation.
authors.affiliation: School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore.
authors.forename: Ardcharaporn
authors.initials: A
authors.lastname: Vararattanavech
authors.affiliation: School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore.
authors.forename: Xin
authors.initials: X
authors.lastname: Lin
authors.affiliation: School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore.
authors.forename: Jaume
authors.initials: J
authors.lastname: Torres
authors.affiliation: School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore. Electronic address: smtan@ntu.edu.sg.
authors.forename: Suet-Mien
authors.initials: SM
authors.lastname: Tan
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D004220:Disulfides / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D047468:Immunoprecipitation; D016169:Lymphocyte Function-Associated Antigen-1 / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids; D017386:Sequence Homology, Amino Acid; D013912:Threonine / Q000235:genetics*
chemical_list: D004220:Disulfides; D016169:Lymphocyte Function-Associated Antigen-1; D013912:Threonine