Interaction of proteinase 3 with CD11b/CD18 (beta2 integrin) on the cell membrane of human neutrophils.
Proteinase 3 (PR3), the target autoantigen of antineutrophil cytoplasmic antibodies in the autoimmune vasculitis, Wegener's granulomatosis, is a serine proteinase stored in granules of human neutrophils. As previously shown, PR3 is expressed also on the plasma membrane of unactivated neutrophils, and this expression increases in primed or stimulated cells. The current study demonstrates that membrane-bound PR3 colocalizes with the adhesion molecule CD11b/CD18 (beta2 integrin). Immunoprecipitation experiments using plasma membranes of phorbol 12-myristate 13-acetate (PMA)-stimulated neutrophils revealed coimmunoprecipitation of PR3 with CD11b/CD18, indicating their location in the same complex. PR3 was also detected in TritonX-100-insoluble cytoskeleton of plasma membranes isolated from unactivated and activated neutrophils. Release of cytoskeletal PR3 by salt treatment implied electrostatic interaction with the enzyme. The serine protease inhibitor phenylmethylsulfonyl fluoride (PMSF) augmented membrane expression of PR3 in unactivated and PMA-stimulated neutrophils. PMSF significantly reduced adhesion of neutrophils to fibrinogen-coated plates and their NADPH oxidase activity. Moreover, the addition of exogenous PR3 (1-5 microg/ml) augmented the CD11b/CD18-dependent adhesion of neutrophils. Taken together, these results implicate the beta2 integrin of neutrophils in their membrane association with PR3 and suggest a role of PR3 in the modulation of cell adhesion.
David
A
A
Department Biochemistry, Tel Aviv University, Israel.
Kacher
Y
Y
Specks
U
U
Aviram
I
I
D039481:CD11b Antigen / Q000032:analysis / Q000502:physiology*; D018821:CD18 Antigens / Q000032:analysis / Q000502:physiology*; D002448:Cell Adhesion; D002462:Cell Membrane / Q000737:chemistry*; D006801:Humans; D053146:Myeloblastin; D019255:NADPH Oxidases / Q000378:metabolism; D009504:Neutrophils / Q000737:chemistry* / Q000502:physiology; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D011233:Precipitin Tests; D012697:Serine Endopeptidases / Q000032:analysis / Q000502:physiology*
D039481:CD11b Antigen; D018821:CD18 Antigens; D010664:Phenylmethylsulfonyl Fluoride; D019255:NADPH Oxidases; D012697:Serine Endopeptidases; D053146:Myeloblastin