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Molecular and General Genetics MGG

, Volume 256, Issue 5, pp 469–480 | Cite as

A defect in GTP synthesis affects mannose outer chain elongation in Saccharomyces cerevisiae

  • Y. Shimma
  • A. Nishikawa
  • B. bin Kassim
  • A. Eto
  • Y. Jigami
ORIGINAL PAPER

Abstract

We have found that yeast mutants that are defective in mannose outer chain elongation of N-linked glycoproteins show higher cell wall porosity than normal cells, and are hypersensitive to antibiotics with a large molecular weight; such as neomycin and geneticin. Wild-type yeast cells also showed enhanced sensitivity to neomycin in the presence of tunicamycin, an inhibitor of N-glycosylation, suggesting that the extent of N-glycosylation may affect the sensitivity of yeast cells to drugs and that sensitivity to neomycin may be an effective method for screening for yeast mutants defective in N-glycosylation. Pursuing this logic, we isolated neomycin-sensitive yeast mutants and screened them for defects in N-glycosylation. The neomycin-sensitive, N-glycosylation-defective mutants fell into 15 complementation groups including alleles of the previously isolated temperature-sensitive nes mutants nes10, nes17, and nes25. Gene cloning revealed that NES10 was identical to SEC20, which is involved in ER-Golgi protein transport. NES17 was identical to ALG1, which encodes a β-1,4-mannosyltransferase present in the ER. MSN17, a multicopy suppressor of nes17/alg1, was also isolated and found to be an allele of PSA1, which is involved in GDP-mannose synthesis. NES25 was identical to GUK1, which encodes a GMP kinase. Overexpression of MSN17 increased the GDP-mannose level in a wild-type strain by about threefold, and guk1 decreased the GDP-mannose level to one-fourth, suggesting a close relationship between GTP metabolism and mannose outer chain elongation; the link is presumably provided by the process of GDP-mannose transport in the Golgi membranes.

Key words GMP kinase GDP-mannose synthesis N-glycosylation Mannose outer chain elongation Saccharomyces cerevisiae 

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Copyright information

© Springer-Verlag Berlin Heidelberg 1997

Authors and Affiliations

  • Y. Shimma
    • 1
  • A. Nishikawa
    • 2
  • B. bin Kassim
    • 1
  • A. Eto
    • 3
  • Y. Jigami
    • 1
  1. 1.Molecular Biology Department, National Institute of Bioscience and Human Technology, 1-1 Higashi, Tsukuba, Ibaraki, 305, Japan Fax: +81-298-54-6220; e-mail: yjigami@ccmail.nibh.go.jpJP
  2. 2.Institute of Biological Sciences, University of Tsukuba, 1-1 Tennodai, Tsukuba, Ibaraki, 305, JapanJP
  3. 3.Department of Agricultural Chemistry, University of Tokyo, 1-1 Yayoi, Bunkyo-ku, Tokyo, 113, JapanJP

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