TY - JOUR
T1 - Decreased synthesis of glycosphingolipids in cells lacking vimentin intermediate filaments
AU - Gillard, Baiba K.
AU - Clement, Rhonda
AU - Colucci-Guyon, Emma
AU - Babinet, Charles
AU - Schwarzmann, Gunter
AU - Taki, Takao
AU - Kasama, Takashi
AU - Marcus, Donald M.
N1 - Funding Information:
We acknowledge the expert technical assistance of Petra Hofmann and thank Dr. Alfred H. Merrill, Jr., for helpful discussions and Mrs. Brenda Galena for expert assistance with the manuscript. The work was supported by research Grants Al 17712 from the National Institutes of Health, BE-88B from the American Cancer Society, from the CNRS and Institut Pasteur, and SFB 284/B5 from the Deutsche Forschungsgemeinschaft.
Copyright:
Copyright 2017 Elsevier B.V., All rights reserved.
PY - 1998/8/1
Y1 - 1998/8/1
N2 - We are studying defects in glycosphingolipid synthesis in cells lacking vimentin intermediate filaments (vimentin-). Sugars can be incorporated into glycolipids whose ceramide is synthesized either de novo (pathway 1) or from sphingoid bases salvaged from hydrolysis of sphingolipids (pathway 2) and into glycolipids recycling from the endosomal pathway through the Golgi (pathway 3). Vimentin- embryonic fibroblasts, obtained from vimentin-knockout mice, incorporate less sugar into glycolipids than vimentin+ fibroblasts. Using two inhibitors of ceramide synthesis, β-chloroalanine and fumonisin B1, we found the major defect in synthesis to be in pathway 2 and not in de novo synthesis. We used two additional approaches to analyze the functions of pathways 2 and 3. First, we used exogenous glucosylthioceramide ([14C]C8- Glc-S-Cer), a synthetic, nonhydrolyzable glycosphingolipid, as a precursor for synthesis of larger glycolipids. Vimentin- SW13 cells and embryonic fibroblasts glycosylated [14C]C8-Glc-S-Cer less extensively than their vimentin+ counterparts. Second, we used chloroquine to inhibit the hydrolysis of sphingolipids in endosomes and lysosomes. Chloroquine markedly decreased the incorporation of sugars into glycolipids larger than glueosylceramide. The defect in glycolipid synthesis in vimentin- cells probably results from impaired intracellular transport of glycolipids and sphingoid bases between the endosomal/lysosomal pathway and the Golgi apparatus and endoplasmic reticulum. Intermediate filaments may accomplish this function by contributing to the organization of subcellular organelles and/or by binding proteins that participate in transport processes.
AB - We are studying defects in glycosphingolipid synthesis in cells lacking vimentin intermediate filaments (vimentin-). Sugars can be incorporated into glycolipids whose ceramide is synthesized either de novo (pathway 1) or from sphingoid bases salvaged from hydrolysis of sphingolipids (pathway 2) and into glycolipids recycling from the endosomal pathway through the Golgi (pathway 3). Vimentin- embryonic fibroblasts, obtained from vimentin-knockout mice, incorporate less sugar into glycolipids than vimentin+ fibroblasts. Using two inhibitors of ceramide synthesis, β-chloroalanine and fumonisin B1, we found the major defect in synthesis to be in pathway 2 and not in de novo synthesis. We used two additional approaches to analyze the functions of pathways 2 and 3. First, we used exogenous glucosylthioceramide ([14C]C8- Glc-S-Cer), a synthetic, nonhydrolyzable glycosphingolipid, as a precursor for synthesis of larger glycolipids. Vimentin- SW13 cells and embryonic fibroblasts glycosylated [14C]C8-Glc-S-Cer less extensively than their vimentin+ counterparts. Second, we used chloroquine to inhibit the hydrolysis of sphingolipids in endosomes and lysosomes. Chloroquine markedly decreased the incorporation of sugars into glycolipids larger than glueosylceramide. The defect in glycolipid synthesis in vimentin- cells probably results from impaired intracellular transport of glycolipids and sphingoid bases between the endosomal/lysosomal pathway and the Golgi apparatus and endoplasmic reticulum. Intermediate filaments may accomplish this function by contributing to the organization of subcellular organelles and/or by binding proteins that participate in transport processes.
KW - Endosomal
KW - Glycosphingolipid metabolism
KW - Lysosomal
KW - Vimentin intermediate filaments
KW - Vimentin knockout
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U2 - 10.1006/excr.1998.4126
DO - 10.1006/excr.1998.4126
M3 - Article
C2 - 9683542
AN - SCOPUS:0032146206
SN - 0014-4827
VL - 242
SP - 561
EP - 572
JO - Experimental Cell Research
JF - Experimental Cell Research
IS - 2
ER -