Focal adhesions are sites of integrin extension.
Integrins undergo global conformational changes that specify their activation state. Current models portray the inactive receptor in a bent conformation that upon activation converts to a fully extended form in which the integrin subunit leg regions are separated to enable ligand binding and subsequent signaling. To test the applicability of this model in adherent cells, we used a fluorescent resonance energy transfer (FRET)-based approach, in combination with engineered integrin mutants and monoclonal antibody reporters, to image integrin alpha5beta1 conformation. We find that restricting leg separation causes the integrin to adopt a bent conformation that is unable to respond to agonists and mediate cell spreading. By measuring FRET between labeled alpha5beta1 and the cell membrane, we find extended receptors are enriched in focal adhesions compared with adjacent regions of the plasma membrane. These results demonstrate definitely that major quaternary rearrangements of beta1-integrin subunits occur in adherent cells and that conversion from a bent to extended form takes place at focal adhesions.
Wellcome Trust Centre for Cell-Matrix Research, Faculty of Life Sciences, University of Manchester, Manchester M13 9PT, England, UK.
Janet A
JA
Askari
Christopher J
CJ
Tynan
Stephen E D
SE
Webb
Marisa L
ML
Martin-Fernandez
Christoph
C
Ballestrem
Martin J
MJ
Humphries
D001665:Binding Sites; D031541:Fluorescence Resonance Energy Transfer; D022001:Focal Adhesions / Q000378:metabolism*; D006801:Humans; D039081:Integrin alpha5beta1 / Q000737:chemistry / Q000378:metabolism; D019012:Integrin beta1 / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation; D015202:Protein Engineering; D017434:Protein Structure, Tertiary
D039081:Integrin alpha5beta1; D019012:Integrin beta1