Mechano-redox control of Mac-1 de-adhesion from ICAM-1 by protein disulfide isomerase promotes directional movement of neutrophils under flow

Abstract

Macrophage-1 antigen or Mac-1 (CD11b/CD18, αMβ2) is a leukocyte integrin essential for firm 20 adhesion of neutrophils, lymphocytes and monocytes against flow when recruited to the 21 endothelium. To migrate to the site of inflammation, leukocytes require coordinated adhesion 22 and de-adhesion for directional movement. The vascular thiol isomerase, protein disulfide 23 isomerase (PDI), was found by fluorescence microscopy to colocalize with high affinity Mac-1 at 24 the trailing edge of stimulated neutrophils when adhered to ICAM-1 under fluid shear. From 25 differential cysteine alkylation and mass spectrometry studies, PDI cleaves two allosteric 26 disulfide bonds, C169-C176 and C224-C264, in the βI domain of the β2 subunit, and in 27 mutagenesis and cell transfection studies, cleavage of the C224-C264 disulfide bond was 28 shown to selectively control Mac-1 dis-engagement from ICAM-1 under fluid shear. Molecular 29 dynamics simulations and binding of conformation-specific antibodies reveal that cleavage of 30 the C224-C264 bond induces conformational change and mechanical stress in the βI domain 31 that allosterically alters exposure of an αI domain epitope and shifts Mac-1 to a lower affinity 32 state.