The mechanism of inhibition of glycosylphosphatidylinositol anchor biosynthesis in Trypanosoma brucei by mannosamine

Julie E. Ralton, Kenneth Milne, Maria Lucia S. Güther, Robert A. Field, Michael A. J, Ferguson

    Research output: Contribution to journalArticle

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    Abstract

    The inhibition of glycosylphosphatidylinositol anchor biosynthesis by mannosamine has been described previously in the procyclic forms of Trypanosoma brucei and in mammalian cells (Lisanti, M. P., Field, M. C., Caras, I. W. J., Menon, A. K., and Rodriguez-Boulan, E. (1991) EMBO J. 10, 1969-1977). A recent report has suggested that mannosamine exerts these effects by becoming incorporated into glycosylphosphatidylinositol anchor intermediates (Pan, Y-T., Kamitani, T., Bhuvaneswaran, C., Hallaq, Y., Warren, C. D., Yeh, E. T. H., and Elbein, A. D. (1992) J. Biol. Chem. 267, 21250-21255). In this paper we have analyzed the effects of mannosamine on glycosylphosphatidylinositol anchor and variant surface glycoprotein biosynthesis in the blood-stream form of T. brucei. Trypanosomes were biosynthetically labeled with [3H]mannosamine, and [3H]glucosamine in the presence of mannosamine, and the structures of the labeled glycolipids which accumulated were determined. The main glycolipid metabolite of mannosamine was shown to be ManN-Man-GlcN-PI. A trypanosome cell-free system preloaded with this compound was significantly impaired in its ability to synthesize glycosylphosphatidylinositol anchor intermediates beyond Man alpha 1-6Man alpha 1-4GlcN alpha 1-6PI. This compound is therefore proposed to be an inhibitor of the Dol-P-Man:Man alpha 1-6Man alpha 1-4GlcNa alpha 1-6PI alpha 1-2-mannosyltransferase of the GPI biosynthetic pathway. In living trypanosomes, 4 mM mannosamine had no effect on protein synthesis but reduced the rate of formation of mature glycosylphosphatidylinositol anchor precursors by 80%. This reduction in anchor precursor synthesis was insufficient to prevent the attachment of glycosylphosphatidylinositol anchors to newly synthesized variant surface glycoprotein molecules. These data suggest that the rate of anchor precursor synthesis in the bloodstream form of T. brucei, in contrast to mammalian cells and the procyclic form of T. brucei, is in large excess of the cellular requirements for protein anchorage.
    Original languageEnglish
    Pages (from-to)24183-24189
    Number of pages7
    JournalJournal of Biological Chemistry
    Volume268
    Issue number32
    Publication statusPublished - 1993

    Fingerprint

    Glycosylphosphatidylinositols
    Trypanosoma brucei brucei
    Biosynthesis
    Trypanosomiasis
    Glycolipids
    Membrane Glycoproteins
    Anchors
    Mannosyltransferases
    Cells
    Cell-Free System
    Glucosamine
    Biosynthetic Pathways
    Metabolites
    mannosamine
    Proteins
    Blood
    Molecules

    Cite this

    @article{853f2da5fad94e4b9a975968f8460bd3,
    title = "The mechanism of inhibition of glycosylphosphatidylinositol anchor biosynthesis in Trypanosoma brucei by mannosamine",
    abstract = "The inhibition of glycosylphosphatidylinositol anchor biosynthesis by mannosamine has been described previously in the procyclic forms of Trypanosoma brucei and in mammalian cells (Lisanti, M. P., Field, M. C., Caras, I. W. J., Menon, A. K., and Rodriguez-Boulan, E. (1991) EMBO J. 10, 1969-1977). A recent report has suggested that mannosamine exerts these effects by becoming incorporated into glycosylphosphatidylinositol anchor intermediates (Pan, Y-T., Kamitani, T., Bhuvaneswaran, C., Hallaq, Y., Warren, C. D., Yeh, E. T. H., and Elbein, A. D. (1992) J. Biol. Chem. 267, 21250-21255). In this paper we have analyzed the effects of mannosamine on glycosylphosphatidylinositol anchor and variant surface glycoprotein biosynthesis in the blood-stream form of T. brucei. Trypanosomes were biosynthetically labeled with [3H]mannosamine, and [3H]glucosamine in the presence of mannosamine, and the structures of the labeled glycolipids which accumulated were determined. The main glycolipid metabolite of mannosamine was shown to be ManN-Man-GlcN-PI. A trypanosome cell-free system preloaded with this compound was significantly impaired in its ability to synthesize glycosylphosphatidylinositol anchor intermediates beyond Man alpha 1-6Man alpha 1-4GlcN alpha 1-6PI. This compound is therefore proposed to be an inhibitor of the Dol-P-Man:Man alpha 1-6Man alpha 1-4GlcNa alpha 1-6PI alpha 1-2-mannosyltransferase of the GPI biosynthetic pathway. In living trypanosomes, 4 mM mannosamine had no effect on protein synthesis but reduced the rate of formation of mature glycosylphosphatidylinositol anchor precursors by 80{\%}. This reduction in anchor precursor synthesis was insufficient to prevent the attachment of glycosylphosphatidylinositol anchors to newly synthesized variant surface glycoprotein molecules. These data suggest that the rate of anchor precursor synthesis in the bloodstream form of T. brucei, in contrast to mammalian cells and the procyclic form of T. brucei, is in large excess of the cellular requirements for protein anchorage.",
    author = "Ralton, {Julie E.} and Kenneth Milne and G{\"u}ther, {Maria Lucia S.} and Field, {Robert A.} and Ferguson, {Michael A. J,}",
    year = "1993",
    language = "English",
    volume = "268",
    pages = "24183--24189",
    journal = "Journal of Biological Chemistry",
    issn = "0021-9258",
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    The mechanism of inhibition of glycosylphosphatidylinositol anchor biosynthesis in Trypanosoma brucei by mannosamine. / Ralton, Julie E.; Milne, Kenneth; Güther, Maria Lucia S.; Field, Robert A.; Ferguson, Michael A. J,.

    In: Journal of Biological Chemistry, Vol. 268, No. 32, 1993, p. 24183-24189.

    Research output: Contribution to journalArticle

    TY - JOUR

    T1 - The mechanism of inhibition of glycosylphosphatidylinositol anchor biosynthesis in Trypanosoma brucei by mannosamine

    AU - Ralton, Julie E.

    AU - Milne, Kenneth

    AU - Güther, Maria Lucia S.

    AU - Field, Robert A.

    AU - Ferguson, Michael A. J,

    PY - 1993

    Y1 - 1993

    N2 - The inhibition of glycosylphosphatidylinositol anchor biosynthesis by mannosamine has been described previously in the procyclic forms of Trypanosoma brucei and in mammalian cells (Lisanti, M. P., Field, M. C., Caras, I. W. J., Menon, A. K., and Rodriguez-Boulan, E. (1991) EMBO J. 10, 1969-1977). A recent report has suggested that mannosamine exerts these effects by becoming incorporated into glycosylphosphatidylinositol anchor intermediates (Pan, Y-T., Kamitani, T., Bhuvaneswaran, C., Hallaq, Y., Warren, C. D., Yeh, E. T. H., and Elbein, A. D. (1992) J. Biol. Chem. 267, 21250-21255). In this paper we have analyzed the effects of mannosamine on glycosylphosphatidylinositol anchor and variant surface glycoprotein biosynthesis in the blood-stream form of T. brucei. Trypanosomes were biosynthetically labeled with [3H]mannosamine, and [3H]glucosamine in the presence of mannosamine, and the structures of the labeled glycolipids which accumulated were determined. The main glycolipid metabolite of mannosamine was shown to be ManN-Man-GlcN-PI. A trypanosome cell-free system preloaded with this compound was significantly impaired in its ability to synthesize glycosylphosphatidylinositol anchor intermediates beyond Man alpha 1-6Man alpha 1-4GlcN alpha 1-6PI. This compound is therefore proposed to be an inhibitor of the Dol-P-Man:Man alpha 1-6Man alpha 1-4GlcNa alpha 1-6PI alpha 1-2-mannosyltransferase of the GPI biosynthetic pathway. In living trypanosomes, 4 mM mannosamine had no effect on protein synthesis but reduced the rate of formation of mature glycosylphosphatidylinositol anchor precursors by 80%. This reduction in anchor precursor synthesis was insufficient to prevent the attachment of glycosylphosphatidylinositol anchors to newly synthesized variant surface glycoprotein molecules. These data suggest that the rate of anchor precursor synthesis in the bloodstream form of T. brucei, in contrast to mammalian cells and the procyclic form of T. brucei, is in large excess of the cellular requirements for protein anchorage.

    AB - The inhibition of glycosylphosphatidylinositol anchor biosynthesis by mannosamine has been described previously in the procyclic forms of Trypanosoma brucei and in mammalian cells (Lisanti, M. P., Field, M. C., Caras, I. W. J., Menon, A. K., and Rodriguez-Boulan, E. (1991) EMBO J. 10, 1969-1977). A recent report has suggested that mannosamine exerts these effects by becoming incorporated into glycosylphosphatidylinositol anchor intermediates (Pan, Y-T., Kamitani, T., Bhuvaneswaran, C., Hallaq, Y., Warren, C. D., Yeh, E. T. H., and Elbein, A. D. (1992) J. Biol. Chem. 267, 21250-21255). In this paper we have analyzed the effects of mannosamine on glycosylphosphatidylinositol anchor and variant surface glycoprotein biosynthesis in the blood-stream form of T. brucei. Trypanosomes were biosynthetically labeled with [3H]mannosamine, and [3H]glucosamine in the presence of mannosamine, and the structures of the labeled glycolipids which accumulated were determined. The main glycolipid metabolite of mannosamine was shown to be ManN-Man-GlcN-PI. A trypanosome cell-free system preloaded with this compound was significantly impaired in its ability to synthesize glycosylphosphatidylinositol anchor intermediates beyond Man alpha 1-6Man alpha 1-4GlcN alpha 1-6PI. This compound is therefore proposed to be an inhibitor of the Dol-P-Man:Man alpha 1-6Man alpha 1-4GlcNa alpha 1-6PI alpha 1-2-mannosyltransferase of the GPI biosynthetic pathway. In living trypanosomes, 4 mM mannosamine had no effect on protein synthesis but reduced the rate of formation of mature glycosylphosphatidylinositol anchor precursors by 80%. This reduction in anchor precursor synthesis was insufficient to prevent the attachment of glycosylphosphatidylinositol anchors to newly synthesized variant surface glycoprotein molecules. These data suggest that the rate of anchor precursor synthesis in the bloodstream form of T. brucei, in contrast to mammalian cells and the procyclic form of T. brucei, is in large excess of the cellular requirements for protein anchorage.

    M3 - Article

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