title: Integrin αIIbβ3 inside-out activation: an in situ conformational analysis reveals a new mechanism.
abstract: Integrins are a family of heterodimeric adhesion receptors that transmit signals bi-directionally across the plasma membranes. The transmembrane domain (TM) of integrin plays a critical role in mediating transition of the receptor from the default inactive to the active state on the cell surfaces. In this study, we successfully applied the substituted cysteine scanning accessibility method to determine the intracellular border of the integrin α(IIb)β(3) TM in the inactive and active states in living cells. We examined the aqueous accessibility of 75 substituted cysteines comprising the C terminus of both α(IIb) and β(3) TMs, the intracellular membrane-proximal regions, and the whole cytoplasmic tails, to the labeling of a membrane-permeable, cysteine-specific chemical biotin maleimide (BM). The active state of integrin α(IIb)β(3) heterodimer was generated by co-expression of activating partners with the cysteine-substituted constructs. Our data revealed that, in the inactive state, the intracellular lipid/aqueous border of α(IIb) TM was at Lys(994) and β(3) TM was at Phe(727) respectively; in the active state, the border of α(IIb) TM shifted to Pro(998), whereas the border of β(3) TM remained unchanged, suggesting that complex conformational changes occurred in the TMs upon α(IIb)β(3) inside-out activation. On the basis of the results, we propose a new inside-out activation mechanism for integrin α(IIb)β(3) and by inference, all of the integrins in their native cellular environment.
authors.affiliation: Department of Medicine, David Geffen School of Medicine, UCLA, Los Angeles, California 90095-1689, USA.
authors.forename: Lisa
authors.initials: L
authors.lastname: Kurtz
authors.forename: Liyo
authors.initials: L
authors.lastname: Kao
authors.forename: Debra
authors.initials: D
authors.lastname: Newman
authors.forename: Ira
authors.initials: I
authors.lastname: Kurtz
authors.forename: Quansheng
authors.initials: Q
authors.lastname: Zhu
mesh_terms: D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000378:metabolism*; D003545:Cysteine / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D008051:Lipid Bilayers / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D015398:Signal Transduction / Q000502:physiology*; D013329:Structure-Activity Relationship; D014867:Water / Q000737:chemistry
chemical_list: D008051:Lipid Bilayers; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D014867:Water; D003545:Cysteine