TY - JOUR
T1 - The mechanism of inhibition of glycosylphosphatidylinositol anchor biosynthesis in Trypanosoma brucei by mannosamine
AU - Ralton, Julie E.
AU - Milne, Kenneth G.
AU - Güther, Maria Lucia S.
AU - Field, Robert A.
AU - Ferguson, Michael A. J.
PY - 1993/11/15
Y1 - 1993/11/15
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 Manα1-6Manα1-4GlcNα1-6PI. This compound is therefore proposed to be an inhibitor of the Dol-P-Man:Manα1-6Manα1-4GlcNα1-6PI α1-2-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 Manα1-6Manα1-4GlcNα1-6PI. This compound is therefore proposed to be an inhibitor of the Dol-P-Man:Manα1-6Manα1-4GlcNα1-6PI α1-2-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.
UR - http://www.scopus.com/inward/record.url?scp=0027363637&partnerID=8YFLogxK
M3 - Article
C2 - 8226965
AN - SCOPUS:0027363637
VL - 268
SP - 24183
EP - 24189
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 32
ER -