The relative contribution of mannose salvage pathways to glycosylation in PMI-deficient mouse embryonic fibroblast cells

Naonobu Fujita, Ayako Tamura, Aya Higashidani, Takashi Tonozuka, Hudson H. Freeze, Atsushi Nishikawa

    研究成果: Article査読

    15 被引用数 (Scopus)

    抄録

    Mannose for mammalian glycan biosynthesis can be imported directly from the medium, derived from glucose or salvaged from endogenous or external glycans. All pathways must generate mannose 6-phosphate, the activated form of mannose. Imported or salvaged mannose is directly phosphorylated by hexokinase, whereas fructose 6-phosphate from glucose is converted to mannose 6-phosphate by phosphomannose isomerase (PMI). Normally, PMI provides the majority of mannose for glycan synthesis. To assess the contribution of PMI-independent pathways, we used PMI-null fibroblasts to study N-glycosylation of DNase I, a highly sensitive indicator protein. In PMI-null cells, imported mannose and salvaged mannose make a significant contribution to N-glycosylation. When these cells were grown in mannose-free medium along with the mannosidase inhibitor, swainsonine, to block the salvage pathways, N-glycosylation of DNase I was almost completely eliminated. Adding ∼13 μm mannose to the medium completely restored normal glycosylation. Treatment with bafilomycin A 1, an inhibitor of lysosomal acidification, also markedly reduced N-glycosylation of DNase I, but in this case only 8 μm mannose was required to restore full glycosylation, indicating that a nonlysosomal source of mannose made a significant contribution. Glycosylation levels were greatly also reduced in glycoconjugate-free medium, when endosomal membrane trafficking was blocked by expression of a mutant SKD1. From these data, we conclude that PMI-null cells can salvage mannose from both endogenous and external glycoconjugates via lysosomal and nonlysosomal degradation pathways.

    本文言語English
    ページ(範囲)788-798
    ページ数11
    ジャーナルFEBS Journal
    275
    4
    DOI
    出版ステータスPublished - 2008 2月

    ASJC Scopus subject areas

    • 生化学
    • 分子生物学
    • 細胞生物学

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