Cocapping of the leukoadhesin molecules complement receptor type 3 and lymphocyte function-associated antigen-1 with Fc gamma receptor III on human neutrophils. Possible role of lectin-like interactions.
We have tested the possible physical interactions between the iC3b receptor (CR3), lymphocyte function-associated Ag-1, and class III Fc gamma receptor (Fc gamma RIII) at neutrophil surfaces. Cells were labeled using fluorochrome-conjugated Fab or F(ab')2 fragments of antireceptor mAb. Labeled receptors were capped using second-step F(ab')2 fragments of goat anti-mouse Fab antiserum. After 20 min at 37 degrees C, 68% of the cells capped the anti-CR3 plus second-step complex. Capping was time, temperature, and cytochalasin B sensitive. When capped cells were probed with Fab' or F(ab')2 fragments of anti-Fc gamma RIII labeled with a distinct fluorochrome, 41% of the cells cocapped Fc gamma RIII. Indistinguishable results were obtained when potential antibody combining sites within caps were blocked with a large excess of Fab or F(ab')2 fragments. When Fc gamma RIII was capped, 49% of the cells cocapped CR3. Similarly, LFA-1 cocapped with both CR3 and Fc gamma RIII. Importantly, other membrane components including HLA class I, Mo5, CD13, CR type 1, and IL-8 receptors and N-4-nitrobenzo-2-oxa-1, 3-diszole L-alpha-dimyristoyl phosphatidylethanolamine did not cocap with CR3. However, the positive control Con A did cocap with CR3 and Fc gamma RIII. We next evaluated the effect of saccharides on CR3-Fc gamma RIII cocapping and found that 0.15 M N-acetyl-D-glucosamine (NADG), alpha-methyl-D-mannoside, and D-mannose significantly inhibited cocapping by 70, 58, and 48%, respectively. No inhibition was obtained using glucose, galactose, N-acetyl-neuraminic acid, fucose, sorbitol, fructose, or sucrose. Similarly, Fc gamma RIII-lymphocyte function-associated-1 cocapping was inhibited by NADG. However, the cocapping of CR3 with lymphocyte function-associated-1 or Con A were not affected by 0.15 M NADG, which suggests that NADG inhibition of leukoadhesin-Fc gamma RIII cocapping is not due to a general effect of NADG on capping. Inasmuch as Fc gamma RIII is a glycophospholipid-linked membrane protein, we speculate that it interacts with CR3 and/or lymphocyte function-associated-1 via lectin-like interactions.
Zhou
M
M
Department of Biological Sciences, Wayne State University, Detroit, MI 48202.
Todd
R F
RF
van de Winkel
J G
JG
Petty
H R
HR
D000328:Adult; D000906:Antibodies / Q000494:pharmacology; D003208:Concanavalin A / Q000737:chemistry; D003432:Cross-Linking Reagents; D005455:Fluorescent Antibody Technique; D006601:Hexoses / Q000494:pharmacology; D006801:Humans; D037102:Lectins / Q000737:chemistry*; D016169:Lymphocyte Function-Associated Antigen-1 / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D016177:Macrophage-1 Antigen / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry; D009504:Neutrophils / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011940:Receptor Aggregation / Q000187:drug effects / Q000276:immunology; D017452:Receptors, IgG / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D013194:Staining and Labeling
D000906:Antibodies; D003432:Cross-Linking Reagents; D006601:Hexoses; D037102:Lectins; D016169:Lymphocyte Function-Associated Antigen-1; D016177:Macrophage-1 Antigen; D008565:Membrane Proteins; D017452:Receptors, IgG; D003208:Concanavalin A