Mac-1 Promotes FcγRIIA-Dependent Cell Spreading and Migration on Immune Complexes

Abstract

The integrin Mac-1 plays a critical role in Fc receptor (FcR)-mediated antibody-dependent cellular cytotoxicity (ADCC). However, the mechanism by which Mac-1 facilitates the functions of FcγRIIA, a major FcR expressed on human leukocytes, is not fully understood. We report here that Mac-1 sustains cell adhesion, enhances cell spreading, and accelerates cell migration on pre-formed immune complexes (ICs) by directly interacting with FcγRIIA but not with the IC substrate. Coupling Mac-1 to FcγRIIA allows FcγRIIA to reside in the leading front of actin polymerization at the filopodial extension, and thus could potentially enhance the FcγRIIA-mediated cell spreading and migration. Direct interaction between Mac-1 and FcγRIIA is demonstrated by co-immunoprecipitation, by cell surface co-localization, and by solid-phase binding assays using recombinant α M I-domain and soluble FcγRIIA. Further mutational analysis identifies the E 253 -R 261 sequence within the α M Idomain as part of the FcγRIIA binding interface within Mac-1. Altogether, these results demonstrate that FcγRIIA recognizes Mac-1 via the α M I-domain but not the lectin domain, a distinct feature from other FcRs, and that Mac-1 binding confers FcγRIIA with the ability to prolong cell adhesion as well as to spread and migrate on the ICs, leading to effective cell killing by ADCC.Receptors recognizing the Fc fragment (FcR) 1 of the immunoglobulin (Ig) molecules are critical to host defense by mediating leukocyte migration toward ICs deposited within the site of infections and by eliciting potent cytolysis of the IC-sensitized targets, such as malignant cells and invading pathogens (1)(2)(3)(4)(5)(6)(7)(8). A number of clinical studies have demonstrated that FcγRIIA (CD32), a major FcR that recognizes the IgG subclass, is essential to the efficacy of several therapeutic antibody-based drugs, including Herceptin for HER-2/neu-positive breast cancer cells, Rituxan for non-hodgkins lymphoma, and the antibodies for melanoma (1,2,9). Yet, inappropriate engagement of FcγRIIA also causes autoimmune diseases in patients undergoing treatment using these therapeutic antibodies (10), and in transgenic mice overexpressing FcγRIIA (11).The FcγRIIA-mediated antibody-dependent cytotoxicity (ADCC) is dependent on both FcγRIIA and the integrin Mac-1 (α M β 2 , CR3, CD11b/CD18) (1,8,12,13). Despite the importance of Mac-1 in the FcγRIIA-mediated leukocyte functions, and the extensive studies conducted on Mac-1 interaction with various FcRs, including FcγRIIIB (CD16), FcαRI (CD89) and FcεRII (CD23), in addition to [14][15][16], the molecular basis underlying Mac-1 recognition of these FcRs is still less understood. Based primarily on inhibition studies using lectin-domain-specific inhibitors such as N-acetyl-D-glucosamine (NADG) (13,15,17), the binding sites within Mac-1 for most of the FcRs are mapped to its lectin-domain in the α subunit. However, as Mac-1's ability to promote FcγRIIA-mediated phagocytosis is insensitive to NADG inhibition (18), the nature of the FcγR...

HLA Class I Antibodies Trigger Increased Adherence of Monocytes to Endothelial Cells by Eliciting an Increase in Endothelial P-Selectin and, Depending on Subclass, by Engaging FcγRs

“…Moreover, despite several fold higher expression of LFA-1 compared with Mac-1 on monocytes, Mono Mac 6 and THP-1 almost exclusively utilized Mac-1 to adhere to HLA I antibody-stimulated endothelial cells. Finally, there is a well-studied signaling cross-talk between Mac-1 and FcγRs, particularly with FcγRI (72, 81, 82). For these reasons, monocytes may be more responsive to HLA antibody bound on the endothelium compared with other myeloid or lymphoid cells.…”

The many faces of Mac‐1 in autoimmune disease

“…FcγRIIA is expressed on neutrophils, eosinophils, macrophages, platelets, and DCs . Physical interaction of Mac‐1 with FcγRIIA by fluorescent resonance energy transfer and co‐immunoprecipitation has been documented . Unexpectedly, this interaction has been found to be via E 252 ‐R 261 in the ligand‐binding I‐domain of Mac‐1 , suggesting an interaction akin to a ligand‐receptor pair.…”

“…Physical interaction of Mac‐1 with FcγRIIA by fluorescent resonance energy transfer and co‐immunoprecipitation has been documented . Unexpectedly, this interaction has been found to be via E 252 ‐R 261 in the ligand‐binding I‐domain of Mac‐1 , suggesting an interaction akin to a ligand‐receptor pair. Functional interaction of FcγRIIA and Mac‐1 has been shown: Transfection of tail‐less FcγRIIA (lacking ITAMs) can phagocytose only when Mac‐1 is cotransfected , and antibody‐dependent cellular cytotoxicity (ADCC) depends on both receptors .…”

Early biochemical events in leukocyte activation through receptors for IgG

“…Also, it has been shown that the integrin Mac-1 interacts directly with FccRIIa and contributes to FccRIIa-mediated sustained cell adhesion, cell spreading, and cell migration on targets covered with immune complexes. These effects likely enhance contact between leukocyte and target, thus favoring effective target cell killing by ADCC or phagocytosis [58]. It has also been reported that FccRIIIb stimulation promotes activation of b1 integrins on human neutrophils [59].…”