Anchoring and degradation of glycolipid-anchored membrane proteins by L929 versus by LM-TK- mouse fibroblasts: implications for anchor biosynthesis.
Although many cells anchor surface proteins via moieties that are sensitive to phosphatidylinositol-specific phospholipase C (PI-PLC), the anchor moieties of surface proteins of mouse L929 cells resist PI-PLC. By constructing stable hybrids between L929 and lymphoma cells that express glycolipid-anchored proteins in a PI-PLC-sensitive form, we show that PI-PLC resistance behaves as a recessive trait. Since putative mannolipid precursors of the lipid anchors bear alkali-labile substituents which make them resist PI-PLC, these observations are most simply interpreted by postulating that L929 lacks a critical anchor deacylase. Unlike the L929 cell line, two of its descendants, the LM cell line and its thymidine kinase-negative variant (LM-TK-), do not express glycolipid-anchored proteins on their surface. Moreover, unlike L929 cells, LM-TK- cells rapidly inactivate at least one lipid-anchored enzyme in a compartment sensitive to acidotropic amines and leupeptin. By fusion of LM-TK- cells to mouse Thy-1- lymphoma mutants and monitoring of surface expression of lipid-anchored proteins, we assign LM-TK- to lymphoma mutant complementation group H. This genetic assignment is matched by analysis of mannolipids of L929, LM-TK-, wild-type, and class H lymphoma mutant cells: striking similarities are seen between the two wild-type cells by contrast to the mutants. Since the differences pertain to lipids which have properties consistent with their being anchor precursors, we suggest that LM-TK- has a lesion in the synthesis of anchor precursor mannolipids.
Singh
N
N
Institute of Pathology, Case Western Reserve University, Cleveland, Ohio 44106.
Singleton
D
D
Tartakoff
A M
AM
D000469:Alkaline Phosphatase / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002459:Cell Fusion; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D005347:Fibroblasts / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D005816:Genetic Complementation Test; D014644:Genetic Variation; D006017:Glycolipids / Q000378:metabolism*; D017261:Glycosylphosphatidylinositols; D007700:Kinetics; D007739:L Cells / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure; D008223:Lymphoma; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D010716:Phosphatidylinositols / Q000378:metabolism*; D013937:Thymidine Kinase / Q000235:genetics; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism
D006017:Glycolipids; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D010716:Phosphatidylinositols; D013937:Thymidine Kinase; D000469:Alkaline Phosphatase; D010738:Type C Phospholipases