Modulation of integrin activation and signaling by α1/α1′-helix unbending at the junction

Abstract

SummaryHow conformational signals initiated from one end of the integrin are transmitted to the other end remains elusive. At the ligand-binding bI domain, the a1/a19-helix changes from a bent to a straightened a-helical conformation upon integrin headpiece opening. We demonstrated that a conserved glycine at the a1/a19 junction is crucial for maintaining the bent conformation of the a1/a19-helix in the resting state. Mutations that facilitate a1/a19-helix unbending rendered integrin constitutively active; however, mutations that block the a1/a19-helix unbending abolished soluble ligand binding upon either outside or inside stimuli. Such mutations also blocked ligandinduced integrin extension from outside the cell, but had no effect on talin-induced integrin extension from inside the cell. In addition, integrin-mediated cell spreading, F-actin stress fiber and focal adhesion formation, and focal adhesion kinase activation were also defective in these mutant integrins, although the cells still adhered to immobilized ligands at a reduced level. Our data establish the structural role of the a1/a19 junction that allows relaxation of the a1/a19-helix in the resting state and transmission of bidirectional conformational signals by helix unbending upon integrin activation.

The dual structural roles of the membrane distal region of α integrin cytoplasmic tail in integrin inside-out activation

“…2C). Similar results were obtained with the active b 3 -G135A mutant, which facilitates formation of the active conformation of b 3 -I domain (Zhang et al, 2013). Complete deletion of the a IIb CT MD region enhanced the activation of b 3 -G135A (data not shown).…”

“…By using conformation-dependent LIBS mAbs, we examined the talin-1-head-induced conformational change of integrin ectodomains. Consistent with our previous results (Zhang et al, 2013), overexpression of the talin-1 head significantly increased the binding of the b 3 LIBS mAb 319.4, but not the a IIb LIBS mAb 370.3 to WT a IIb b 3 (Fig. 6A,B).…”

“…We have recently shown that the talin-1-head-induced integrin conformational change needs to be propagated to the ligandbinding site, probably through headpiece opening, in order to activate integrin (Zhang et al, 2013). In this study, by using the conformation-dependent mAbs that report integrin extension (319.4 for b 3 and 370.3 for a IIb ; KIM127 for b 2 ) and headpiece opening (M24 for b 2 ), we further demonstrated that the talin-1 head induced extension and headpiece opening of integrin.…”

“…DNA constructs of human a IIb b 3 , a V b 3 , a L b 2 , and the EGFP-tagged mouse talin-1 head (EGFP-TH) were as described previously (Zhu et al, 2007;Bouaouina et al, 2008;Zhang et al, 2013). Human kindlin-2 or kindlin-3 was cloned into the pmCherry-C1 vector (Clontech).…”

“…AP3 is a conformationindependent anti-b 3 mAb (Kouns et al, 1991). 319.4 is a LIBS mAb that binds to the b 3 I-EGF domains (Zhang et al, 2013). 370.3 is a LIBS mAb that binds to the a IIb calf-1 domain (Zhang et al, 2013).…”