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Motonori Hoshi - One of the best experts on this subject based on the ideXlab platform.
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a novel difucosylated Neutral Glycosphingolipid from the eggs of the sea urchin hemicentrotus pulcherrimus i purification and structural determination of the glycolipid
Journal of Biochemistry, 1992Co-Authors: Hideo Kubo, Fuyuhiko Inagaki, Gang Jung Jiang, Atsushi Irie, Minoru Suzuki, Motonori HoshiAbstract:A novel fucose-containing Neutral Glycosphingolipid (GL-5) was purified from the eggs of the sea urchin, Hemicentrotus pulcherrimus. The chemical structure was determined to be Fuc alpha 1-3GalNAc beta 1-4(Fuc alpha 1-3)GlcNAc beta 1-4Glc beta 1-1Cer by methylation analysis, partial acid hydrolysis, fast atom bombardment mass spectrometry, and proton nuclear magnetic resonance spectroscopy. The unique characteristics of GL-5 are that: the reducing terminal disaccharide portion is not Gal beta 1-4Glc but GlcNAc beta 1-4Glc; it includes a GalNAc beta 1-4GlcNAc sequence and a Fuc-GalNAc linkage; the defucosylated core is a novel trisaccharide chain; and the sugar structure is one of the smallest ever characterized for a difucosylated glycolipid. The major fatty acids were 22:1 and 22h:1, and about 30% of the total acids was 2-hydroxylated. All the long-chain bases were phytosphingosines, of which about 90% was n-t18:0. The similarity of the ceramide moiety to that of glucosylceramide from the same eggs [Kubo, H. et al. (1992) J. Biochem. 111, 726-731] suggests a close biosynthetic relationship between GL-5 and the glucosylceramide.
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A novel difucosylated Neutral Glycosphingolipid from the eggs of the sea urchin, Hemicentrotus pulcherrimus: II. Structural determination by two-dimensional NMR.
Journal of biochemistry, 1992Co-Authors: Fuyuhiko Inagaki, Shin-ichi Tate, Hideo Kubo, Motonori HoshiAbstract:A novel fucosylglycolipid from the eggs of the sea urchin, Hemicentrotus pulcherrimus, was determined by using two-dimensional NMR methods. Subspectra extraction by the homonuclear Hartmann-Hahn method was useful for identification of the individual sugar components. The homonuclear Hartmann-Hahn and double-quantum-filtered correlated spectra were analyzed to establish the assignments of sugar proton resonances. On the basis of the resonance assignments, the linkages of the individual sugar components were determined to be as follows. [formula: see text] This glycolipid contains a novel skeletal structure with the linkages of GalNAc beta 1-4GlcNAc beta 1-4Glc beta. We also observed that 2-hydroxylation of the fatty acids induced appreciable chemical shift changes of the proton resonances of the phytosphingosine moiety as well as the anomeric proton resonance of the reducing terminal glucose.
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Hemicentrotus pulcherrimus1
1991Co-Authors: Toshiko Matsubara, Motonori HoshiAbstract:Glucosylceramide (Glc/91-lCer) and a novel ceramide trihexoside (Gal£l-6Gal£l-6Glc£l-lCer) were purified from the eggs of the sea urchin, Hemicentrotus pulcherrimus. Their chemical structures were determined by gas-liquid chromatography, methylation analy-sis, chromic acid oxidation, enzymatic hydrolysis, enzyme-linked immunosorbent assay, fast atom bombardment mass spectrometry, and proton nuclear magnetic resonance spectroscopy. The ceramide trihexoside has a novel carbohydrate structure, and its core structure, Gal/?l-6Glc, is also novel. The ceramide moieties of these glycolipids are almost identical. Two fatty acids, 22:1 and 22h:l, constitute more than 80 % of the total acids. Long-chain bases are all phytosphingosine, approximately 90 % of which is n-tl8:0. The finding of melibiosylceramide (Galal-6Glc£l-lCer) from the eggs of another sea urchin species [Kubo, H. et al. (1988) J. Biochem. 104,765-760] and the present finding of the novel ceramide trihexoside suggest that there are a variety of unique sugar structures in sea urchin Glycosphingolipids. Eggs of the sea urchin, Anthocidaris crassispina, contain ghicosylceramide and a novel Neutral Glycosphingolipid
Anthony H Futerman - One of the best experts on this subject based on the ideXlab platform.
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upstream of growth and differentiation factor 1 uog1 a mammalian homolog of the yeast longevity assurance gene 1 lag1 regulates n stearoyl sphinganine c18 dihydro ceramide synthesis in a fumonisin b1 independent manner in mammalian cells
Journal of Biological Chemistry, 2002Co-Authors: Krishnan Venkataraman, Alfred H. Merrill, Cameron M Sullards, Christian Riebeling, Jacques Bodennec, Jeremy C Allegood, Howard Riezman, Anthony H FutermanAbstract:Abstract The longevity assurance gene (LAG1) and its homolog (LAC1) are required for acyl-CoA-dependent synthesis of ceramides containing very long acyl chain (e.g. C26) fatty acids in yeast, and a homolog of LAG1, ASC1, confers resistance in plants to fumonisin B1, an inhibitor of ceramide synthesis. To understand further the mechanism of regulation of ceramide synthesis, we now characterize a mammalian homolog of LAG1,upstream of growth and differentiation factor-1 (uog1). cDNA clones of uog1 were obtained from expression sequence-tagged clones and sub-cloned into a mammalian expression vector. Transient transfection of human embryonic kidney 293T cells with uog1 followed by metabolic labeling with [4,5-3H]sphinganine orl-3-[3H]serine demonstrated thatuog1 conferred fumonisin B1 resistance with respect to the ability of the cells to continue to produce ceramide. Surprisingly, this ceramide was channeled into Neutral Glycosphingolipids but not into gangliosides. Electrospray tandem mass spectrometry confirmed the elevation in sphingolipids and revealed that the ceramides and Neutral Glycosphingolipids ofuog1-transfected cells contain primarily stearic acid (C18), that this enrichment was further increased by FB1, and that the amount of stearic acid in sphingomyelin was also increased. UOG1 was localized to the endoplasmic reticulum, demonstrating that the fatty acid selectivity and the fumonisin B1 resistance are not due to a subcellular localization different from that found previously for ceramide synthase activity. Furthermore, in vitro assays ofuog1-transfected cells demonstrated elevated ceramide synthase activity when stearoyl-CoA but not palmitoyl-CoA was used as substrate. We propose a role for UOG1 in regulating C18-ceramide (N-stearoyl-sphinganine) synthesis, and we note that not only is this the first case of ceramide formation in mammalian cells with such a high degree of fatty acid specificity, but also that theN-stearoyl-sphinganine produced by UOG1 most significantly impacts Neutral Glycosphingolipid synthesis.
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up regulation of Neutral Glycosphingolipid synthesis upon long term inhibition of ceramide synthesis by fumonisin b1
Journal of Biological Chemistry, 1999Co-Authors: I Meivarlevy, Anthony H FutermanAbstract:In a previous study we observed that long term (5 days) incubation with fumonisin B1 (FB1), an inhibitor of acylation of sphingoid long chain bases to (dihydro)ceramide, resulted in morphological and biochemical changes in 3T3 fibroblasts (Meivar-Levy, I., Sabanay, H., Bershadsky, A. D., and Futerman, A. H. (1997) J. Biol. Chem. 272, 1558–1564). Among these were changes in the profile of synthesis of sphingolipids (SLs) and Glycosphingolipids (GSLs). Whereas [3H]globotriaosylceramide ([3H]Gb3) comprised 1.9% of the total [3H]SLs and [3H]GSLs synthesized in control cells, it comprised 16.5% in FB1-treated cells. We now demonstrate by in vitro analysis that inhibition of ceramide synthesis by FB1 for 5 days results in up-regulation of the activities of three enzymes in the pathway of Gb3 synthesis, namely glucosylceramide, lactosylceramide, and Gb3 synthases; up-regulation is due to an increase inV max, with no change in K mvalues toward lipid substrates. Moreover, molecular analysis (reverse transcriptase-polymerase chain reaction) of glucosylceramide synthase indicated that this enzyme is up-regulated at the transcriptional level. No changes in either the V max orK m values of sphingomyelin or of GM3synthase were detected after FB1 treatment. Analysis of SL and GSL synthesis in cultured cells using [4,5-3H]sphinganine as a metabolic precursor demonstrated that at low substrate concentrations, Gb3 synthesis is favored over GM3 synthesis and glucosylceramide synthesis is favored over sphingomyelin synthesis, whereas the opposite is true at high substrate concentrations. These data demonstrate that GSL synthesis and in particular Gb3 synthesis are tightly regulated in fibroblasts, presumably so as to maintain constant levels of Gb3 on the cell surface.
Bernhard Schermer - One of the best experts on this subject based on the ideXlab platform.
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Dysregulated Autophagy Contributes to Podocyte Damage in Fabry's Disease
PLoS ONE, 2013Co-Authors: Max C Liebau, V Bartels, Fabian Braun, Claudia Weitbrecht, Katja Höpker, Susanne Brodesser, Moin A. Saleem, Roman-ulrich Müller, Thomas Benzing, Bernhard SchermerAbstract:Fabry's disease results from an inborn error of Glycosphingolipid metabolism that is due to deficiency of the lysosomal hydrolase α-galactosidase A. This X-linked defect results in the accumulation of enzyme substrates with terminally α-glycosidically bound galactose, mainly the Neutral Glycosphingolipid Globotriaosylceramide (Gb3) in various tissues, including the kidneys. Although end-stage renal disease is one of the most common causes of death in hemizygous males with Fabry's disease, the pathophysiology leading to proteinuria, hematuria, hypertension, and kidney failure is not well understood. Histological studies suggest that the accumulation of Gb3 in podocytes plays an important role in the pathogenesis of glomerular damage. However, due to the lack of appropriate animal or cellular models, podocyte damage in Fabry's disease could not be directly studied yet. As murine models are insufficient, a human model is needed. Here, we developed a human podocyte model of Fabry's disease by combining RNA interference technology with lentiviral transduction of human podocytes. Knockdown of α-galactosidase A expression resulted in diminished enzymatic activity and slowly progressive accumulation of intracellular Gb3. Interestingly, these changes were accompanied by an increase in autophagosomes as indicated by an increased abundance of LC3-II and a loss of mTOR kinase activity, a negative regulator of the autophagic machinery. These data suggest that dysregulated autophagy in α-galactosidase A-deficient podocytes may be the result of deficient mTOR kinase activity. This finding links the lysosomal enzymatic defect in Fabry's disease to deregulated autophagy pathways and provides a promising new direction for further studies on the pathomechanism of glomerular injury in Fabry patients.
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Dysregulated Autophagy Contributes to Podocyte Damage in Fabry’s Disease
2013Co-Authors: Max C Liebau, V Bartels, Fabian Braun, Claudia Weitbrecht, Katja Höpker, Susanne Brodesser, Moin A. Saleem, Roman-ulrich Müller, Thomas Benzing, Bernhard SchermerAbstract:Fabry’s disease results from an inborn error of Glycosphingolipid metabolism that is due to deficiency of the lysosomal hydrolase α-galactosidase A. This X-linked defect results in the accumulation of enzyme substrates with terminally α-glycosidically bound galactose, mainly the Neutral Glycosphingolipid Globotriaosylceramide (Gb3) in various tissues, including the kidneys. Although end-stage renal disease is one of the most common causes of death in hemizygous males with Fabry’s disease, the pathophysiology leading to proteinuria, hematuria, hypertension, and kidney failure is not well understood. Histological studies suggest that the accumulation of Gb3 in podocytes plays an important role in the pathogenesis of glomerular damage. However, due to the lack of appropriate animal or cellular models, podocyte damage in Fabry’s disease could not be directly studied yet. As murine models are insufficient, a human model is needed. Here, we developed a human podocyte model of Fabry’s disease by combining RNA interference technology with lentiviral transduction of human podocytes. Knockdown of α-galactosidase A expression resulted in diminished enzymatic activity and slowly progressive accumulation of intracellular Gb3. Interestingly, these changes were accompanied by an increase in autophagosomes as indicated by an increased abundance of LC3-II and a loss of mTOR kinase activity, a negative regulator of the autophagic machinery. These data suggest that dysregulated autophagy in α-galactosidase A-deficient podocytes may be the result of deficient mTOR kinase activity. This finding links the lysosomal enzymatic defect in Fabry’s disease to deregulated autophagy pathways and provides a promising new direction for further studies on the pathomechanism of glomerular injury in Fabry patients.
Pamela Matto - One of the best experts on this subject based on the ideXlab platform.
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a novel mass spectrometric assay for the cerebroside sulfate activator protein saposin b and arylsulfatase a
Journal of Lipid Research, 2005Co-Authors: Andrew J Norris, Arman Yaghoubian, Julian P Whitelegge, Jeanrene Alattia, Gilbert G Prive, Tatsushi Toyokuni, Hubert Sun, Mai N Brooks, Luigi Panza, Pamela MattoAbstract:A mass spectrometric method is described for monitoring cerebrosides in the presence of excess concen- trations of alkali metal salts. This method has been adapted for use in the assay of arylsulfatase A (ASA) and the cere- broside sulfate activator protein (CSAct or saposin B). De- tection of the Neutral Glycosphingolipid cerebroside prod- uct was achieved via enhancement of ionization efficiency in the presence of lithium ions. Assay samples were ex- tracted into the chloroform phase as for the existing assays, dried, and diluted in methanol-chloroform-containing lith- ium chloride. Samples were analyzed by electrospray ioniza- tion mass spectrometry with a triple quadrupole mass spec- trometer in the multiple reaction monitoring tandem mass spectrometric mode. The assay has been used to demon- strate several previously unknown or ambiguous aspects of the coupled ASA/CSAct reaction, including an absolute in vitro preference for CSAct over the other saposins (A, C, and D) and a preference for the nonhydroxylated species of the sulfatide substrate over the corresponding hydroxylated species. The modified assay for the coupled ASA/CSAct reaction could find applicability in settings in which the as- say could not be performed previously because of the need for radiolabeled substrate, which is now not required. — Norris, A. J., J. P. Whitelegge, A. Yaghoubian, J-R. Alattia, G. G. Prive, T. Toyokuni, H. Sun, M. N. Brooks, L. Panza, P. Matto, F. Compostella, N. Remmel, R. Klingenstein, K. Sandhoff, C. Fluharty, A. Fluharty, and K. F. Faull. A novel mass spectrometric assay for the cerebroside sulfate activa- tor protein (saposin B) and arylsulfatase A. J. Lipid Res. 2005. 46: 2254-2264.
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a novel mass spectrometric assay for the cerebroside sulfate activator protein saposin b and arylsulfatase a
Journal of Lipid Research, 2005Co-Authors: Andrew J Norris, Arman Yaghoubian, Julian P Whitelegge, Jeanrene Alattia, Gilbert G Prive, Tatsushi Toyokuni, Hubert Sun, Mai N Brooks, Luigi Panza, Pamela MattoAbstract:A mass spectrometric method is described for monitoring cerebrosides in the presence of excess concentrations of alkali metal salts. This method has been adapted for use in the assay of arylsulfatase A (ASA) and the cerebroside sulfate activator protein (CSAct or saposin B). Detection of the Neutral Glycosphingolipid cerebroside product was achieved via enhancement of ionization efficiency in the presence of lithium ions. Assay samples were extracted into the chloroform phase as for the existing assays, dried, and diluted in methanol-chloroform-containing lithium chloride. Samples were analyzed by electrospray ionization mass spectrometry with a triple quadrupole mass spectrometer in the multiple reaction monitoring tandem mass spectrometric mode. The assay has been used to demonstrate several previously unknown or ambiguous aspects of the coupled ASA/CSAct reaction, including an absolute in vitro preference for CSAct over the other saposins (A, C, and D) and a preference for the nonhydroxylated species of the sulfatide substrate over the corresponding hydroxylated species. The modified assay for the coupled ASA/CSAct reaction could find applicability in settings in which the assay could not be performed previously because of the need for radiolabeled substrate, which is now not required.
John Gordon - One of the best experts on this subject based on the ideXlab platform.
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dynamic interplay between the Neutral Glycosphingolipid cd77 gb3 and the therapeutic antibody target cd20 within the lipid bilayer of model b lymphoma cells
Biochemical and Biophysical Research Communications, 2007Co-Authors: Rosemary M Jarvis, Michael J Lord, Michelle J Holder, Anita Chamba, Anita Challa, Daniel C Smith, Matthew N Hodgkin, John GordonAbstract:The centroblast-specific differentiation marker CD77 (Gb(3)), is the receptor for Shiga-like toxin (SLT). The dynamic relationship between Gb(3)/CD77 and key B-cell membrane proteins was studied in Burkitt's lymphoma cells with a focus on CD20. Engagement of Gb3/CD77 with SLT-B reduced the amount of CD20 and CXCR4 available, but levels of BCR, MHC Class 11, CD21, CD27 and CD54 remained unchanged. Cholesterol depletion promoted a decrease in the number of sites accessed by CD20, CXCR4 and Gb(3)/ CD77 antibodies. Constitutive localisation of Gb3/CD77 to lipid rafts was unperturbed by either SLT-B binding or cholesterol depletion, whereas the opposite was true for CD20. The effects were specific to SLT-B, highlighted by the inability of cholera toxin B-subunit to alter CD20 availability. Thus, the binding of Gb3/CD77 by its cognate ligand transmits information within the lipid bilayer of model lymphoma cells to impact the behaviour of selective proteins, most notably CD20, via a mechanism influenced by the level of cholesterol within the membrane. (c) 2007 Elsevier Inc. All rights reserved.
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cd40 ligand bcl 2 and bcl xl spare group i burkitt lymphoma cells from cd77 directed killing via verotoxin 1 b chain but fail to protect against the holotoxin
Cell Death & Differentiation, 2000Co-Authors: John Gordon, Anita Challa, Jacqueline Levens, Christopher D Gregory, Julie Williams, Richard J Armitage, J P Cook, Lynne M Roberts, Janet M LordAbstract:Owing to its lineage and differentiation stage-restricted expression, CD77 has been mooted as a therapeutic target in Burkitt lymphoma (BL). The recognition that the globotriaosyl moiety of this Neutral Glycosphingolipid is a receptor for Escherichia coli-derived Verotoxin-1 (Shiga-Like Toxin-1) offers a potential delivery system for the attack. Here we show that CD77-expressing Group I BL cells which are normally susceptible to activation-induced death on binding Verotoxin-1 B chain are protected in the presence of CD40 ligand. Ectopic expression of either bcl-2 or bcl-xL also afforded resistance to the actions of the B chain. In total contrast, neither of the survival genes nor a CD40 signal - even when acting in concert - protected against killing mediated by the holotoxin. These findings indicate that while therapeutic modalities for CD77-expressing B cell tumors (which include follicular lymphoma) based on the use of Verotoxin-1 B chain might be compromised by the activation of endogenous or exogenous survival pathways, those exploiting the holotoxin should be left unscathed.