Pga1 is an essential component of Glycosylphosphatidylinositol-mannosyltransferase II of Saccharomyces cerevisiae.
The Saccharomyces cerevisiae essential gene YNL158w/PGA1 encodes an endoplasmic reticulum (ER)-localized membrane protein. We constructed temperature-sensitive alleles of PGA1 by error-prone polymerase chain reaction mutagenesis to explore its biological role. Pulse-chase experiments revealed that the pga1(ts) mutants accumulated the ER-form precursor of Gas1 protein at the restrictive temperature. Transport of invertase and carboxypeptidase Y were not affected. Triton X-114 phase separation and [(3)H]inositol labeling indicated that the glycosylphosphatidylinositol (GPI)-anchoring was defective in the pga1(ts) mutants, suggesting that Pga1 is involved in GPI synthesis or its transfer to target proteins. We found GPI18, which was recently reported to encode GPI-mannosyltransferase II (GPI-MT II), as a high-copy suppressor of the temperature sensitivity of pga1(ts). Both Gpi18 and Pga1 were detected in the ER by immunofluorescence, and they were coprecipitated from the Triton X-100-solubilized membrane. The gpi18(ts) and pga1(ts) mutants accumulated the same GPI synthetic intermediate at the restrictive temperature. From these results, we concluded that Pga1 is an additional essential component of the yeast GPI-MT II.
Sato
Keisuke
K
Department of Biotechnology, University of Tokyo, Tokyo 113-8657, Japan.
Noda
Yoichi
Y
Yoda
Koji
K
D000483:Alleles; D000595:Amino Acid Sequence; D004721:Endoplasmic Reticulum / Q000201:enzymology; D004795:Enzyme Stability; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006801:Humans; D008364:Mannosyltransferases / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D050296:Microbial Viability; D008954:Models, Biological; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000302:isolation & purification / Q000378:metabolism; D010641:Phenotype; D011485:Protein Binding; D011498:Protein Precursors / Q000378:metabolism; D021122:Protein Subunits / Q000378:metabolism; D021381:Protein Transport; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D016415:Sequence Alignment; D013696:Temperature
C066709:GAS1 protein, S cerevisiae; D017261:Glycosylphosphatidylinositols; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D050505:Mutant Proteins; D011498:Protein Precursors; D021122:Protein Subunits; D029701:Saccharomyces cerevisiae Proteins; D008364:Mannosyltransferases; C524121:PGA1 protein, S cerevisiae; C033143:glycolipid 3-alpha-mannosyltransferase