Affected paroxysmal nocturnal hemoglobinuria T lymphocytes harbor a common defect in assembly of N-acetyl-D-glucosamine inositol phospholipid corresponding to that in class A Thy-1- murine lymphoma mutants.
Deficient expression of glycoinositol phospholipid (GPI) anchored proteins in affected paroxysmal nocturnal hemoglobinuria (PNH) cells has been traced to a defect in GPI anchor assembly. In a previous study (Schubert, J., Schmidt, R. E., and Medof, M. E. (1993) J. Biol. Chem., in press) we characterized the biosynthesis of putative Man-containing GPI anchor precursors in normal peripheral blood lymphocytes and investigated assembly of these intracellular GPI intermediates in CD48- affected and CD48+ unaffected T and natural killer cell lines of PNH patients. We found that affected T cells from five patients exhibited a uniform defect in which dolichol-phosphoryl-Man was synthesized but no GPI mannolipids were expressed. In this study, membranes of patients' affected T cells were labeled with UDP-[3H]GlcNAc to evaluate earlier steps in GPI synthesis, and intact cells were fused to Thy-1- murine lymphoma mutants harboring different defects in early GPI assembly to test for the presence of corresponding or complementary lesions. In all cases, affected cell membranes failed to assemble GlcNAc-inositol phospholipid, the initial precursor of GPI anchor structures, and the intact cells failed to complement class A mutants while complementing other classes. Affected polymorphonuclear leukocytes from three additional patients of different origin were then labeled with [3H]Man and the labeling patterns found to correspond to those obtained with the T lymphocytes. Taken together the data indicate that the genetic lesion in PNH cells resides in a DNA element which: 1) encodes a product required for the synthesis of GlcNAc-inositol phospholipid, 2) corresponds to that altered in class A Thy-1- murine lymphoma mutants, and 3) is commonly affected in different patients.
Armstrong
C
C
Institute of Pathology, Case Western Reserve University, Cleveland, Ohio 44106.
Schubert
J
J
Ueda
E
E
Knez
J J
JJ
Gelperin
D
D
Hirose
S
S
Silber
R
R
Hollan
S
S
Schmidt
R E
RE
Medof
M E
ME
D000117:Acetylglucosamine / Q000378:metabolism*; D000818:Animals; D015703:Antigens, CD / Q000235:genetics; D000071178:CD48 Antigen; D002462:Cell Membrane / Q000378:metabolism; D004287:Dolichol Monophosphate Mannose / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006457:Hemoglobinuria, Paroxysmal / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology; D008223:Lymphoma / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D008358:Mannose / Q000097:blood; D051379:Mice; D009154:Mutation; D009504:Neutrophils / Q000276:immunology / Q000378:metabolism; D014346:Trypanosoma brucei brucei / Q000378:metabolism; D014537:Uridine Diphosphate N-Acetylglucosamine / Q000378:metabolism
D015703:Antigens, CD; D000071178:CD48 Antigen; C000604065:CD48 protein, human; C000604066:Cd48 protein, mouse; D017261:Glycosylphosphatidylinositols; D014537:Uridine Diphosphate N-Acetylglucosamine; D004287:Dolichol Monophosphate Mannose; D008358:Mannose; D000117:Acetylglucosamine