A Transmembrane Polar Interaction Is Involved in the Functional Regulation of Integrin αLβ2

Integrins are heterodimeric transmembrane (TM) receptors formed by noncovalent associations of αandβ subunits. Each subunit contains a single α-helical TM domain. Inside-out activation of an integrin involves the separation of its cytoplasmic tails, leading to disruption of αβ TM packing. The leukocyte integrin αLβ2 is required for leukocyte adhesion, migration, proliferation, cytotoxic function, and antigen presentation. In this study, we show by mutagenesis experiments that the packing of αLβ2 TMs is consistent with that of the integrin αIIbβ3 TMs. However, molecular dynamics simulations of αLβ2 TMs in lipids predicted a polar interaction involving the side chains of αL Ser1071 and β2 Thr686 in the outer-membrane association clasp (OMC). This is supported by carbonyl vibrational shifts observed in isotope-labeled αLβ2 TM peptides that were incorporated into lipid bilayers. Molecular dynamics studies simulating the separation of αLβ2 tails showed the presence of polar interaction during the initial perturbation of the inner-membrane association clasp. When the TMs underwent further separation, the polar interaction was disrupted. OMC polar interaction is important in regulating the functions of β2 integrins because mutations that disrupt the OMC polar interaction generated constitutively activated αLβ2, αMβ2, and αxβ2 in 293T transfectants. We also show that the expression of mutant β2 Thr686Gly in β2-deficient T cells rescued cell adhesion to intercellular adhesion molecule 1, but the cells showed overt elongated morphologies in response to chemokine stromal-cell-derived factor 1α treatment as compared to wild-type β2-expressing cells. These two TM polar residues are totally conserved in other members of the β2 integrins in humans and across different species. Our results provide an example of the stabilizing effect of polar interactions within the low dielectric environment of the membrane interior and demonstrate its importance in the regulation of αLβ2 function.