Abstract
Within the endoplasmic reticulum (ER), mannoses and glucoses, donated from dolichol-phosphate-mannose and -glucose, are transferred to N-glycan and GPI-anchor precursors, and serine/threonine residues in many proteins. Glycosyltransferases that mediate these reactions are ER-resident multitransmembrane proteins with common characteristics, forming a superfamily of >10 enzymes. Here, we report an essential component of glycosylphosphatidylinositol-mannosyltransferase I (GPI-MT-I), which transfers the first of the four mannoses in the GPI-anchor precursors. We isolated a Chinese hamster ovary (CHO) cell mutant defective in GPI-MT-I but not its catalytic component PIG-M. The mutant gene, termed phosphatidylinositolglycan-class X (PIG-X), encoded a 252-amino acid ER-resident type I transmembrane protein with a large lumenal domain. PIG-X and PIG-M formed a complex, and PIG-M expression was <10% in the absence of PIG-X, indicating that PIG-X stabilizes PIG-M. We found that Saccharomyces cerevisiae Pbn1p/YCL052Cp, which was previously reported to be involved in autoprocessing of proproteinase B, is the functional homologue of PIG-X; Pbn1p is critical for Gpi14p/YJR013Wp function, the yeast homologue of PIG-M. This is the first report of an essential subcomponent of glycosyltransferases using dolichol-phosphate-monosaccharide.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Anti-Infective Agents / pharmacology
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Base Sequence
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Blotting, Western
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CD59 Antigens / biosynthesis
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CHO Cells
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Catalysis
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Cell Line
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Cell Separation
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Cell Survival
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Cloning, Molecular
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Codon, Initiator
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Cricetinae
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Dolichols / chemistry
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Endoplasmic Reticulum / metabolism
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Flow Cytometry
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Glucose / chemistry
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Glycosyltransferases / chemistry
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Humans
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Lipids / chemistry
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Mannosyltransferases / chemistry*
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Mannosyltransferases / metabolism
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Mannosyltransferases / physiology*
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Membrane Glycoproteins / physiology*
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Membrane Proteins / chemistry*
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Mice
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Molecular Sequence Data
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Monosaccharides / chemistry
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Mutation
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Oligosaccharides / chemistry
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Phenotype
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Phosphates / chemistry
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Plasmids / metabolism
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Protein Binding
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Rats
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Saccharomyces cerevisiae / metabolism
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Saccharomyces cerevisiae Proteins / physiology*
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Sequence Homology, Amino Acid
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Serine Endopeptidases / chemistry
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Transfection
Substances
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Anti-Infective Agents
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CD59 Antigens
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Codon, Initiator
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Dolichols
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Lipids
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Membrane Glycoproteins
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Membrane Proteins
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Monosaccharides
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Oligosaccharides
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PBN1 protein, S cerevisiae
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Phosphates
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Saccharomyces cerevisiae Proteins
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Glycosyltransferases
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Mannosyltransferases
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PIGM protein, human
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PIGX protein, human
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glycosylphosphatidylinositol mannosyltransferase I
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Serine Endopeptidases
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yeast proteinase B
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Glucose