Endocytosis and shedding of the decay accelerating factor on human polymorphonuclear cells.
The decay-accelerating factor (DAF) is a cell membrane glycoprotein that functions in the control of C activation. We studied the modulation of membrane DAF on polymorphonuclear cells (PMN) by using anti-DAF antibodies. Fluorescence-activated cell sorter analysis showed that DAF expression was reduced by 43 +/- 7% on resting or stimulated cells that were held at 37 degrees C for 30 min when compared with those kept on ice. Most of this reduction occurred within the first 15 min, and was followed by a gradual further decrease in surface DAF. PMN that were held at 37 degrees C for varying periods of time before DAF measurement had a gradual decrease suggestive of release of DAF from the PMN membrane or endocytosis. To examine the latter, PMN were reacted with anti-DAF at 0 degree C, followed by 125I-Fab'2 secondary antibodies at either 0 degree C or 37 degrees C, and subsequently treated with pronase. Thirty +/- 11% of the 125I remained bound to cells kept at 37 degrees C compared to 2% in those held at 0 degrees C. Internalization was further confirmed by electron microscopy. In PMN that were not exposed to pronase, 26 +/- 2% of the surface-associated 125I was released at 37 degrees C compared with 7% at 0 degrees C. Immunoprecipitation and SDS-PAGE of surface-labeled PMN showed that the temperature-dependent released DAF had a lower m.w. than membrane DAF. Immunofluorescent studies revealed that 37 degrees C mediated the redistribution of DAF from a homogeneous pattern into caps. These results show that under the conditions studied DAF is partially internalized and partially released from the PMN membrane to the fluid phase; the latter may contribute to the presence of DAF in body fluids.
Tausk
F
F
Division of Dermatology, University of California, San Diego, School of Medicine 92103.
Fey
M
M
Gigli
I
I
D000911:Antibodies, Monoclonal / Q000494:pharmacology; D018958:CD55 Antigens; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D004705:Endocytosis; D006801:Humans; D007106:Immune Sera / Q000494:pharmacology; D007152:Immunologic Capping; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism / Q000502:physiology; D008565:Membrane Proteins / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D009504:Neutrophils / Q000378:metabolism / Q000502:physiology* / Q000648:ultrastructure
D000911:Antibodies, Monoclonal; D018958:CD55 Antigens; D003432:Cross-Linking Reagents; D007106:Immune Sera; D008562:Membrane Glycoproteins; D008565:Membrane Proteins