Expression of platelet glycoprotein Ib alpha in HEL cells.
We have previously shown that platelet glycoprotein Ib is expressed in a minority of cells of the human leukemic cell line HEL (Tabilio, A., Rosa, J. P., Testa, U., Kieffer, N., Nurden, A. T., Del Canizo, M. C., Breton-Gorius, J., and Vainchenker, W. (1984) EMBO J. 3, 453-459). In this report, we have selected a stable HEL subclone with increased expression of glycoprotein (GP) Ib as assessed by 6 different monoclonal antibodies in order to investigate the biochemical characteristics of this glycoprotein. A single polypeptide chain of apparent Mr = 60,000 was precipitated under reducing and nonreducing conditions by a specific polyclonal anti-platelet glycocalicin antibody and two anti-GPIb alpha monoclonal antibodies (AN51 and AP1), both from surface-labeled and metabolically labeled HEL cells. We were unable to demonstrate the presence of a polypeptide corresponding to the beta subunit of GPIb or GPIX which is closely associated with GPIb. Competitive immunoprecipitation performed in the presence of an excess amount of cold platelet glycocalicin completely displaced the Mr = 60,000 polypeptide. Synthesis of N-linked oligosaccharide chains on this Mr = 60,000 polypeptide was inhibited by the antibiotic tunicamycin, and a shift of the apparent Mr from 60,000 to 48,000 was observed. O-Linked oligosaccharide chains identical to platelet GPIb hexasaccharides were deficient or incomplete since no peanut agglutinin binding to the Mr = 60,000 polypeptide was observed after neuraminidase treatment of HEL cells. Thus, our results provide evidence that the Mr = 60,000 polypeptide expressed on the surface membrane of HEL cells is closely related to platelet GPIb and corresponds to an incompletely or abnormally O-glycosylated GPIb alpha subunit.
Kieffer
N
N
Debili
N
N
Wicki
A
A
Titeux
M
M
Henri
A
A
Mishal
Z
Z
Breton-Gorius
J
J
Vainchenker
W
W
Clemetson
K J
KJ
D001212:Asialoglycoproteins / Q000276:immunology; D001792:Blood Platelets / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism*; D008970:Molecular Weight; D009844:Oligosaccharides / Q000032:analysis; D019038:Platelet Glycoprotein GPIb-IX Complex; D010980:Platelet Membrane Glycoproteins / Q000032:analysis / Q000096:biosynthesis* / Q000276:immunology / Q000378:metabolism; D014415:Tunicamycin / Q000494:pharmacology
D001212:Asialoglycoproteins; D009844:Oligosaccharides; D019038:Platelet Glycoprotein GPIb-IX Complex; D010980:Platelet Membrane Glycoproteins; C023592:glycocalicin; D014415:Tunicamycin