The integrin α_Lβ₂ hybrid domain serves as a link for the propagation of activation signal from its stalk regions to the I-like domain

Integrin activation involves global conformational changes as demonstrated by various functional and structural analyses. The integrin β hybrid domain is proposed to be involved in the propagation of this activation signal. Our previous study showed that the integrin β₂-specific monoclonal antibody 7E4 abrogates monoclonal antibody KIM185-activated but not Mg ²⁺/EGTA-activated leukocyte function-associated antigen-1 (LFA-1; α_Lβ₂)-mediated adhesion to ICAM-1. Here we investigated the allosteric inhibitory property of 7E4. By using human/mouse chimeras and substitution mutations, the epitope of 7E4 was mapped to Val ⁴⁰⁷, located in the mid-region of the β₂ hybrid domain. Two sets of constitutively active LFA-1 variants were used to examine the effect of 7E4 on LFA-1/ICAM-1 binding. 7E4 attenuated the binding of variants that have modifications to regions membrane proximal with respect to the β₂ hybrid domain. In contrast, the inhibitory effect was minimal on variants with alterations in the α_LI- and β₂ I-like domains preceding the hybrid domain. Furthermore, 7E4 abrogated LFA-1/ICAM-1 adhesion of phorbol 12-myristate 13-acetate-treated MOLT-4 cells. Our data demonstrate that interaction between the hybrid and I-like domain is critical for the regulation of LFA-1-mediated adhesion.