Physical association of complement receptor type 3 and urokinase-type plasminogen activator receptor in neutrophil membranes.
A previous study has shown that Fc gamma RIIIB (CD16), an extensively glycosylated glycosyl-phosphatidylinositol-linked neutrophil membrane protein, specifically co-caps with the iC3b R (CR3; CD11b/CD18). This study tests the possible physical interactions of another extensively glycosylated glycosyl-phosphatidylinositol-linked protein, the urokinase-type plasminogen activator receptor (uPAR), with CR3. Receptors were labeled using fluorochrome-conjugated F(ab')2 fragments of an anti-CR3 mAb. In some cases cells were capped using second step F(ab')2 fragments of an anti-mouse F(ab')2 antiserum. After 30 min at 37 degrees C, 65 +/- 4% of the cells exhibited CR3 caps whereas 61 +/- 2% demonstrated uPAR caps. When CR3-capped cells were probed with F(ab')2 fragments of anti-uPAR conjugated to a distinct fluorochrome, 45 +/- 3% of the cells co-capped uPAR. When uPAR was capped, 48 +/- 2% of the cells co-capped CR3. Similar levels of co-capping were observed using a DNP-conjugated anti-CR3 F(ab')2 and an anti-DNP second step F(ab')2 reagent for capping or using FITC-uPA as a probing reagent. Furthermore, CR3-uPAR co-capping and/or co-clustering was also observed using anti-CR3 IgM and Mn2+ as integrin aggregation stimuli. Significant co-capping of anti-CD14, anti-CD59, anti-Mo5, anti-HLA, or NBD-PE (a lipid probe) was not observed. Moreover, CR3 and uPAR co-capping was blocked by N-acetyl-D-glucosamine, but not by six other saccharides, suggesting that a lectin-like site may participate in co-capping. This suggests that CR3 may regulate adhesive events by several mechanisms, including the regulation of the spatial distribution of uPAR.
Xue
W
W
Department of Biological Sciences, Wayne State University, Detroit, MI 48202.
Kindzelskii
A L
AL
Todd
R F
RF
Petty
H R
HR
D000328:Adult; D000911:Antibodies, Monoclonal; D002241:Carbohydrates / Q000494:pharmacology; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D003432:Cross-Linking Reagents; D005455:Fluorescent Antibody Technique; D006801:Humans; D066298:In Vitro Techniques; D016177:Macrophage-1 Antigen / Q000378:metabolism*; D008345:Manganese / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D011940:Receptor Aggregation / Q000187:drug effects; D011956:Receptors, Cell Surface / Q000378:metabolism*; D055293:Receptors, Urokinase Plasminogen Activator
D000911:Antibodies, Monoclonal; D002241:Carbohydrates; D003432:Cross-Linking Reagents; D016177:Macrophage-1 Antigen; C524898:PLAUR protein, human; C524899:Plaur protein, mouse; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D008345:Manganese