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Chen L, Elizalde M, Alvarez-Sola G. The Role of Sulfatides in Liver Health and Disease. FRONT BIOSCI-LANDMRK 2025; 30:25077. [PMID: 39862071 DOI: 10.31083/fbl25077] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2024] [Revised: 07/31/2024] [Accepted: 08/07/2024] [Indexed: 01/27/2025]
Abstract
Sulfatides or 3-O-sulfogalactosylceramide are negatively charged sulfated glycosphingolipids abundant in the brain and kidneys and play crucial roles in nerve impulse conduction and urinary pH regulation. Sulfatides are present in the liver, specifically in the biliary tract. Sulfatides are self-lipid antigens presented by cholangiocytes to activate cluster of differentiation 1d (CD1d)-restricted type II natural killer T (NKT) cells. These cells are involved in alcohol-related liver disease (ArLD) and ischemic liver injury and exert anti-inflammatory effects by regulating the activity of pro-inflammatory type I NKT cells. Loss of sulfatides has been implicated in the chronic inflammatory disorder of the liver known as primary sclerosing cholangitis (PSC); bile ducts deficient in sulfatides increase their permeability, resulting in the spread of bile into the liver parenchyma. Previous studies have shown elevated levels of sulfatides in hepatocellular carcinoma (HCC), where sulfatides could act as adhesive molecules that contribute to cancer metastasis. We have recently demonstrated how loss of function of GAL3ST1, a limiting enzyme involved in sulfatide synthesis, reduces tumorigenic capacity in cholangiocarcinoma (CCA) cells. The biological function of sulfatides in the liver is still unclear; however, this review aims to summarize the existing findings on the topic.
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Affiliation(s)
- Lin Chen
- Department of Surgery, School of Nutrition and Translational Research in Metabolism, Maastricht University, 6200 MD Maastricht, The Netherlands
| | - Montserrat Elizalde
- Division of Gastroenterology-Hepatology, Department of Internal Medicine, Maastricht University, 6200 MD Maastricht, The Netherlands
| | - Gloria Alvarez-Sola
- Department of Surgery, School of Nutrition and Translational Research in Metabolism, Maastricht University, 6200 MD Maastricht, The Netherlands
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2
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Singh N, Singh AK. A comprehensive review on structural and therapeutical insight of Cerebroside sulfotransferase (CST) - An important target for development of substrate reduction therapy against metachromatic leukodystrophy. Int J Biol Macromol 2024; 258:128780. [PMID: 38104688 DOI: 10.1016/j.ijbiomac.2023.128780] [Citation(s) in RCA: 6] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/26/2023] [Revised: 12/09/2023] [Accepted: 12/12/2023] [Indexed: 12/19/2023]
Abstract
This review is an effort towards the development of substrate reduction therapy using cerebroside sulfotransferase (CST) as a target protein for the development of inhibitors intended to treat pathophysiological condition resulting from the accumulation of sulfatide, a product from the catalytic action of CST. Accumulation of sulfatides leads to progressive impairment and destruction of the myelin structure, disruption of normal physiological transmission of electrical impulse between nerve cells, axonal loss in the central and peripheral nervous system and cumulatively gives a clinical manifestation of metachromatic leukodystrophy. Thus, there is a need to develop specific and potent CST inhibitors to positively control sulfatide accumulation. Structural similarity and computational studies revealed that LYS85, SER172 and HIS141 are key catalytic residues that determine the catalytic action of CST through the transfer of sulfuryl group from the donor PAPS to the acceptor galactosylceramide. Computational studies revealed catalytic site of CST consists two binding site pocket including PAPS binding pocket and substrate binding pocket. Specific substrate site residues in CST can be targeted to develop specific CST inhibitors. This review also explores the challenges of CST-directed substrate reduction therapy as well as the opportunities available in natural products for inhibitor development.
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Affiliation(s)
- Nivedita Singh
- Department of Dravyaguna, Faculty of Ayurveda, Institute of Medical Sciences, Banaras Hindu University, Varanasi 221005, Uttar Pradesh, India.
| | - Anil Kumar Singh
- Department of Dravyaguna, Faculty of Ayurveda, Institute of Medical Sciences, Banaras Hindu University, Varanasi 221005, Uttar Pradesh, India
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3
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Dustin E, McQuiston AR, Honke K, Palavicini JP, Han X, Dupree JL. Adult-onset depletion of sulfatide leads to axonal degeneration with relative myelin sparing. Glia 2023; 71:2285-2303. [PMID: 37283058 PMCID: PMC11007682 DOI: 10.1002/glia.24423] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2022] [Revised: 05/10/2023] [Accepted: 05/12/2023] [Indexed: 06/08/2023]
Abstract
3-O-sulfogalactosylceramide (sulfatide) constitutes a class of sphingolipids that comprise about 4% of myelin lipids in the central nervous system. Previously, our group characterized a mouse with sulfatide's synthesizing enzyme, cerebroside sulfotransferase (CST), constitutively disrupted. Using these mice, we demonstrated that sulfatide is required for establishment and maintenance of myelin, axoglial junctions, and axonal domains and that sulfatide depletion results in structural pathologies commonly observed in Multiple Sclerosis (MS). Interestingly, sulfatide is reduced in regions of normal appearing white matter (NAWM) of MS patients. Sulfatide reduction in NAWM suggests depletion occurs early in disease development and consistent with functioning as a driving force of disease progression. To closely model MS, an adult-onset disease, our lab generated a "floxed" CST mouse and mated it against the PLP-creERT mouse, resulting in a double transgenic mouse that provides temporal and cell-type specific ablation of the Cst gene (Gal3st1). Using this mouse, we demonstrate adult-onset sulfatide depletion has limited effects on myelin structure but results in the loss of axonal integrity including deterioration of domain organization accompanied by axonal degeneration. Moreover, structurally preserved myelinated axons progressively lose the ability to function as myelinated axons, indicated by the loss of the N1 peak. Together, our findings indicate that sulfatide depletion, which occurs in the early stages of MS progression, is sufficient to drive the loss of axonal function independent of demyelination and that axonal pathology, which is responsible for the irreversible loss of neuronal function that is prevalent in MS, may occur earlier than previously recognized.
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Affiliation(s)
- E Dustin
- Department of Anatomy and Neurobiology, Virginia Commonwealth University, Richmond, Virginia, USA
- Research Service, Central Virginia Veterans Affairs Health Care Systems, Richmond, Virginia, USA
| | - A R McQuiston
- Department of Anatomy and Neurobiology, Virginia Commonwealth University, Richmond, Virginia, USA
| | - K Honke
- Department of Biochemistry, Kochi University Medical School, Kochi, Japan
| | - J P Palavicini
- Department of Medicine, University of Texas Health San Antonio, San Antonio, Texas, USA
- Barshop Institute for Longevity and Aging Studies, University of Texas Health San Antonio, San Antonio, Texas, USA
| | - X Han
- Department of Medicine, University of Texas Health San Antonio, San Antonio, Texas, USA
- Barshop Institute for Longevity and Aging Studies, University of Texas Health San Antonio, San Antonio, Texas, USA
| | - J L Dupree
- Department of Anatomy and Neurobiology, Virginia Commonwealth University, Richmond, Virginia, USA
- Research Service, Central Virginia Veterans Affairs Health Care Systems, Richmond, Virginia, USA
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Soprano LL, Ferrero MR, Jacobs T, Couto AS, Duschak VG. Hallmarks of the relationship between host and Trypanosoma cruzi sulfated glycoconjugates along the course of Chagas disease. Front Cell Infect Microbiol 2023; 13:1028496. [PMID: 37256110 PMCID: PMC10225527 DOI: 10.3389/fcimb.2023.1028496] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/26/2022] [Accepted: 04/17/2023] [Indexed: 06/01/2023] Open
Abstract
American Trypanosomiasis or Chagas disease (ChD), a major problem that is still endemic in large areas of Latin America, is caused by Trypanosoma cruzi. This agent holds a major antigen, cruzipain (Cz). Its C-terminal domain (C-T) is retained in the glycoprotein mature form and bears several post-translational modifications. Glycoproteins containing sulfated N-linked oligosaccharides have been mostly implicated in numerous specific procedures of molecular recognition. The presence of sulfated oligosaccharides was demonstrated in Cz, also in a minor abundant antigen with serine-carboxypeptidase (SCP) activity, as well as in parasite sulfatides. Sulfate-bearing glycoproteins in Trypanosomatids are targets of specific immune responses. T. cruzi chronically infected subjects mount specific humoral immune responses to sulfated Cz. Unexpectedly, in the absence of infection, mice immunized with C-T, but not with sulfate-depleted C-T, showed ultrastructural heart anomalous pathological effects. Moreover, the synthetic anionic sugar conjugate GlcNAc6SO3-BSA showed to mimic the N-glycan-linked sulfated epitope (sulfotope) humoral responses that natural Cz elicits. Furthermore, it has been reported that sulfotopes participate via the binding of sialic acid Ig-like-specific lectins (Siglecs) to sulfosialylated glycoproteins in the immunomodulation by host-parasite interaction as well as in the parasite infection process. Strikingly, recent evidence involved Cz-sulfotope-specific antibodies in the immunopathogenesis and infection processes during the experimental ChD. Remarkably, sera from chronically T. cruzi-infected individuals with mild disease displayed higher levels of IgG2 antibodies specific for sulfated glycoproteins and sulfatides than those with more severe forms of the disease, evidencing that T. cruzi sulfotopes are antigenic independently of the sulfated glycoconjugate type. Ongoing assays indicate that antibodies specific for sulfotopes might be considered biomarkers of human cardiac ChD progression, playing a role as predictors of stability from the early mild stages of chronic ChD.
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Affiliation(s)
- Luciana L. Soprano
- Area of Protein Biochemistry and Parasite Glycobiology, Research Department National Institute of Parasitology (INP)”Dr. Mario Fatala Chaben”, National Administration of Health Institutes (ANLIS)-Malbrán, National Health Department, National Council of Scientific and Technical Research (CONICET), Buenos Aires, Argentina
| | - Maximiliano R. Ferrero
- Max-Planck Heart and Lung Laboratory, Research Institute in Biomedicine in Buenos Aires (IBioBA), Argentine-Department of Internal Medicine II, University Medical Center Giessen and Marburg, Giessen, Germany
| | - Thomas Jacobs
- Immunology Department, Bernhard Notch Institute of Tropical Medicine, Hamburg, Germany
| | - Alicia S. Couto
- Faculty in Exact and Natural Sciences (FCEN), Chemical Organic Department-National Council of Scientific and Technical Research (CONICET), Center of CarboHydrates (CHIHIDECAR), University of Buenos Aires, Buenos Aires, Argentina
| | - Vilma G. Duschak
- Area of Protein Biochemistry and Parasite Glycobiology, Research Department National Institute of Parasitology (INP)”Dr. Mario Fatala Chaben”, National Administration of Health Institutes (ANLIS)-Malbrán, National Health Department, National Council of Scientific and Technical Research (CONICET), Buenos Aires, Argentina
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Nakashima K, Hirahara Y, Koike T, Tanaka S, Gamo K, Oe S, Hayashi S, Seki-Omura R, Nakano Y, Ohe C, Yoshida T, Kataoka Y, Tsuda M, Yamashita T, Honke K, Kitada M. Sulfatide with ceramide composed of phytosphingosine (t18:0) and 2-hydroxy fatty acids in renal intercalated cells. J Lipid Res 2022; 63:100210. [PMID: 35439525 PMCID: PMC9157219 DOI: 10.1016/j.jlr.2022.100210] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2021] [Revised: 03/28/2022] [Accepted: 04/10/2022] [Indexed: 11/27/2022] Open
Abstract
Diverse molecular species of sulfatide with differences in FA lengths, unsaturation degrees, and hydroxylation statuses are expressed in the kidneys. However, the physiological functions of specific sulfatide species in the kidneys are unclear. Here, we evaluated the distribution of specific sulfatide species in the kidneys and their physiological functions. Electron microscopic analysis of kidneys of Cst-deficient mice lacking sulfatide showed vacuolar accumulation in the cytoplasm of intercalated cells in the collecting duct, whereas the proximal and distal tubules were unchanged. Immunohistochemical analysis revealed that vacuolar H+-ATPase-positive vesicles were accumulated in intercalated cells in sulfatide-deficient kidneys. Seventeen sulfatide species were detected in the murine kidney by iMScope MALDI-MS analysis. The distribution of the specific sulfatide species was classified into four patterns. Although most sulfatide species were highly expressed in the outer medullary layer, two unique sulfatide species of m/z 896.6 (predicted ceramide structure: t18:0-C22:0h) and m/z 924.6 (predicted ceramide structure: t18:0-C24:0h) were dispersed along the collecting duct, implying expression in intercalated cells. In addition, the intercalated cell-enriched fraction was purified by fluorescence-activated cell sorting using the anti-vacuolar H+-ATPase subunit 6V0A4, which predominantly contained sulfatide species (m/z 896.6 and 924.6). The Degs2 and Fa2h genes, which are responsible for ceramide hydroxylation, were expressed in the purified intercalated cells. These results suggested that sulfatide molecular species with ceramide composed of phytosphingosine (t18:0) and 2-hydroxy FAs, which were characteristically expressed in intercalated cells, were involved in the excretion of NH3 and protons into the urine.
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Affiliation(s)
| | | | | | | | | | | | | | | | | | | | - Takashi Yoshida
- Department of Urology and Andrology, Kansai Medical University, Hirakata, Osaka, Japan
| | - Yosky Kataoka
- Laboratory for Cellular Function Imaging, RIKEN Center for Biosystems Dynamics Research; Multi-Modal Microstructure Analysis Unit, RIKEN－JEOL Collaboration Center, Kobe, Hyogo, Japan
| | | | - Tatsuyuki Yamashita
- Department of Biochemistry, Kochi University Medical School, Nangoku, Kochi, Japan
| | - Koichi Honke
- Department of Biochemistry, Kochi University Medical School, Nangoku, Kochi, Japan
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Blomqvist M, Zetterberg H, Blennow K, Månsson JE. Sulfatide in health and disease. The evaluation of sulfatide in cerebrospinal fluid as a possible biomarker for neurodegeneration. Mol Cell Neurosci 2021; 116:103670. [PMID: 34562592 DOI: 10.1016/j.mcn.2021.103670] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2021] [Revised: 09/14/2021] [Accepted: 09/17/2021] [Indexed: 10/20/2022] Open
Abstract
Sulfatide (3-O-sulfogalactosylceramide, SM4) is a glycosphingolipid, highly multifunctional and particularly enriched in the myelin sheath of neurons. The role of sulfatide has been implicated in various biological fields such as the nervous system, immune system, host-pathogen recognition and infection, beta cell function and haemostasis/thrombosis. Thus, alterations in sulfatide metabolism and production are associated with several human diseases such as neurological and immunological disorders and cancers. The unique lipid-rich composition of myelin reflects the importance of lipids in this specific membrane structure. Sulfatide has been shown to be involved in the regulation of oligodendrocyte differentiation and in the maintenance of the myelin sheath by influencing membrane dynamics involving sorting and lateral assembly of myelin proteins as well as ion channels. Sulfatide is furthermore essential for proper formation of the axo-glial junctions at the paranode together with axonal glycosphingolipids. Alterations in sulfatide metabolism are suggested to contribute to myelin deterioration as well as synaptic dysfunction, neurological decline and inflammation observed in different conditions associated with myelin pathology (mouse models and human disorders). Body fluid biomarkers are of importance for clinical diagnostics as well as for patient stratification in clinical trials and treatment monitoring. Cerebrospinal fluid (CSF) is commonly used as an indirect measure of brain metabolism and analysis of CSF sulfatide might provide information regarding whether the lipid disruption observed in neurodegenerative disorders is reflected in this body fluid. In this review, we evaluate the diagnostic utility of CSF sulfatide as a biomarker for neurodegenerative disorders associated with dysmyelination/demyelination by summarising the current literature on this topic. We can conclude that neither CSF sulfatide levels nor individual sulfatide species consistently reflect the lipid disruption observed in many of the demyelinating disorders. One exception is the lysosomal storage disorder metachromatic leukodystrophy, possibly due to the genetically determined accumulation of non-metabolised sulfatide. We also discuss possible explanations as to why myelin pathology in brain tissue is poorly reflected by the CSF sulfatide concentration. The previous suggestion that CSF sulfatide is a marker of myelin damage has thereby been challenged by more recent studies using more sophisticated laboratory techniques for sulfatide analysis as well as improved sample selection criteria due to increased knowledge on disease pathology.
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Affiliation(s)
- Maria Blomqvist
- Department of Clinical Chemistry, Sahlgrenska University Hospital, Gothenburg, Sweden; Department of Laboratory Medicine, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
| | - Henrik Zetterberg
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, Mölndal, Sweden; Clinical Neurochemistry Laboratory, Sahlgrenska University Hospital, Mölndal, Sweden; Department of Neurodegenerative Disease, UCL Institute of Neurology, Queen Square, London, UK; UK Dementia Research Institute at UCL, London, UK
| | - Kaj Blennow
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, Mölndal, Sweden; Clinical Neurochemistry Laboratory, Sahlgrenska University Hospital, Mölndal, Sweden
| | - Jan-Eric Månsson
- Department of Clinical Chemistry, Sahlgrenska University Hospital, Gothenburg, Sweden; Department of Laboratory Medicine, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden
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Porubsky S, Nientiedt M, Kriegmair MC, Siemoneit JHH, Sandhoff R, Jennemann R, Borgmann H, Gaiser T, Weis CA, Erben P, Hielscher T, Popovic ZV. The prognostic value of galactosylceramide-sulfotransferase (Gal3ST1) in human renal cell carcinoma. Sci Rep 2021; 11:10926. [PMID: 34035403 PMCID: PMC8149814 DOI: 10.1038/s41598-021-90381-6] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2020] [Accepted: 05/11/2021] [Indexed: 12/30/2022] Open
Abstract
Renal cell carcinoma (RCC) is the deadliest primary genitourinary malignancy typically associated with asymptomatic initial presentation and poorly predictable survival. Next to established risk factors, tumor microenvironment may alter metastatic capacity and immune landscape. Due to their high concentrations, sulfoglycolipids (sulfatides) were among the first well-described antigens in RCC that are associated with worse prognosis. As sulfatide detection in routine diagnostics is not possible, we aimed to test the prognostic value of its protein counterpart, sulfatide-producing enzyme Gal3ST1. We performed retrospective long-term follow up analysis of Gal3ST1 expression as prognostic risk factor in a representative RCC patient cohort. We observed differentially regulated Gal3ST1 expression in all RCC types, being significantly more associated with clear cell RCC than to chromophobe RCC (p = 0.001). Surprisingly, in contrast to published observations from in vitro models, we could not confirm an association between Gal3ST1 expression and a malignant clinical behaviour of the RCC. In our cohort, Gal3ST1 did not significantly influence progression-free survival (Hazard Ratio (HR): 1.7 95% CI (0.6–4.9), p = 0.327). Particularly after adjusting for histology, T-stage, N-status and M-status at baseline, we observed no independent prognostic effect (HR = 1.0 95% CI (0.3–3.3), p = 0.96). The analysis of Gal3ST1 mRNA expression in a TCGA dataset supported the results of our cohort. Thus, Gal3ST1 might help to differentiate between chromophobe RCC and other frequent RCC entities but—despite previously published data from cell culture models—does not qualify as a prognostic marker for RCC. Further investigation of regulatory mechanisms of sulfatide metabolism in human RCC microenvironment is necessary to understand the role of this quantitatively prominent glycosphingolipid in RCC progression.
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Affiliation(s)
- Stefan Porubsky
- Institute of Pathology, University Medical Center Mannheim, University of Heidelberg, Theodor-Kutzer-Ufer 1-3, 68167, Mannheim, Germany.,Institute of Pathology, University Medical Center, Johannes-Gutenberg University, Mainz, Germany
| | - Malin Nientiedt
- Department of Urology and Urosurgery, University Medical Center Mannheim, University of Heidelberg, Mannheim, Germany
| | - Maximilian C Kriegmair
- Department of Urology and Urosurgery, University Medical Center Mannheim, University of Heidelberg, Mannheim, Germany
| | - Jörn-Helge Heinrich Siemoneit
- Institute of Pathology, University Medical Center Mannheim, University of Heidelberg, Theodor-Kutzer-Ufer 1-3, 68167, Mannheim, Germany
| | - Roger Sandhoff
- Lipid Pathobiochemistry Group, German Cancer Research Center, Heidelberg, Germany
| | - Richard Jennemann
- Lipid Pathobiochemistry Group, German Cancer Research Center, Heidelberg, Germany
| | - Hendrik Borgmann
- Department of Urology, University Medical Center, Johannes-Gutenberg University, Mainz, Germany
| | - Timo Gaiser
- Institute of Pathology, University Medical Center Mannheim, University of Heidelberg, Theodor-Kutzer-Ufer 1-3, 68167, Mannheim, Germany
| | - Cleo-Aron Weis
- Institute of Pathology, University Medical Center Mannheim, University of Heidelberg, Theodor-Kutzer-Ufer 1-3, 68167, Mannheim, Germany
| | - Philipp Erben
- Department of Urology and Urosurgery, University Medical Center Mannheim, University of Heidelberg, Mannheim, Germany
| | - Thomas Hielscher
- Department of Biostatistics, German Cancer Research Center, Heidelberg, Germany
| | - Zoran V Popovic
- Institute of Pathology, University Medical Center Mannheim, University of Heidelberg, Theodor-Kutzer-Ufer 1-3, 68167, Mannheim, Germany.
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Nakayama M, Miyagawa H, Kuranami Y, Tsunooka-Ota M, Yamaguchi Y, Kojima-Aikawa K. Annexin A4 inhibits sulfatide-induced activation of coagulation factor XII. J Thromb Haemost 2020; 18:1357-1369. [PMID: 32145147 DOI: 10.1111/jth.14789] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2019] [Revised: 02/24/2020] [Accepted: 03/02/2020] [Indexed: 12/28/2022]
Abstract
BACKGROUND Factor XII (FXII) is a plasma serine protease that initiates the intrinsic pathway of blood coagulation upon contact with anionic substances, such as the sulfated glycolipid sulfatide. Annexins (ANXs) have been implicated in the regulation of the blood coagulation reaction by binding to anionic surfaces composed of phospholipids and sulfated glycoconjugates, but their physiological importance is only partially understood. OBJECTIVE To test the hypothesis that ANXs are involved in suppressing the intrinsic pathway initiated by sulfatide, we examined the effect of eight recombinant ANX proteins on the intrinsic coagulation reaction and their sulfatide binding activities. METHODS Recombinant ANXs were prepared in Escherichia coli expression systems and their anticoagulant effects on the intrinsic pathway initiated by sulfatide were examined using plasma clotting assay and chromogenic assay. ANXA4 active sites were identified by alanine scanning and fold deletion in the core domain. RESULTS AND CONCLUSIONS We found that ANXA3, ANXA4, and ANXA5 strongly inhibited sulfatide-induced plasma coagulation. Wild-type and mutated ANXA4 were used to clarify the molecular mechanism involved in inhibition. ANXA4 inhibited sulfatide-induced auto-activation of FXII to FXIIa and the conversion of its natural substrate FXI to FXIa but showed no effect on the protease activity of FXIIa or FXIa. Alanine scanning showed that substitution of the Ca2+ -binding amino acid residue in the fourth fold of the core domain of ANXA4 reduced anticoagulant activity, and deletion of the entire fourth fold of the core domain resulted in complete loss of anticoagulant activity.
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Affiliation(s)
- Moeka Nakayama
- Division of Advanced Sciences, Graduate School of Humanities and Sciences, Ochanomizu University, Tokyo, Japan
- Program for Leading Graduate Schools, Ochanomizu University, Tokyo, Japan
| | - Hitomi Miyagawa
- Division of Advanced Sciences, Graduate School of Humanities and Sciences, Ochanomizu University, Tokyo, Japan
| | - Yumiko Kuranami
- Division of Advanced Sciences, Graduate School of Humanities and Sciences, Ochanomizu University, Tokyo, Japan
| | - Miyuki Tsunooka-Ota
- Division of Advanced Sciences, Graduate School of Humanities and Sciences, Ochanomizu University, Tokyo, Japan
| | - Yoshiki Yamaguchi
- Synthetic Cellular Chemistry Laboratory, RIKEN, Saitama, Japan
- Laboratory of Pharmaceutical Physical Chemistry, Tohoku Medical and Pharmaceutical University, Miyagi, Japan
| | - Kyoko Kojima-Aikawa
- Natural Science Division, Faculty of Core Research, Ochanomizu University, Tokyo, Japan
- Institute for Human Life Innovation, Ochanomizu University, Tokyo, Japan
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9
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Tanphaichitr N, Kongmanas K, Faull KF, Whitelegge J, Compostella F, Goto-Inoue N, Linton JJ, Doyle B, Oko R, Xu H, Panza L, Saewu A. Properties, metabolism and roles of sulfogalactosylglycerolipid in male reproduction. Prog Lipid Res 2018; 72:18-41. [PMID: 30149090 PMCID: PMC6239905 DOI: 10.1016/j.plipres.2018.08.002] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2018] [Revised: 08/20/2018] [Accepted: 08/21/2018] [Indexed: 12/16/2022]
Abstract
Sulfogalactosylglycerolipid (SGG, aka seminolipid) is selectively synthesized in high amounts in mammalian testicular germ cells (TGCs). SGG is an ordered lipid and directly involved in cell adhesion. SGG is indispensable for spermatogenesis, a process that greatly depends on interaction between Sertoli cells and TGCs. Spermatogenesis is disrupted in mice null for Cgt and Cst, encoding two enzymes essential for SGG biosynthesis. Sperm surface SGG also plays roles in fertilization. All of these results indicate the significance of SGG in male reproduction. SGG homeostasis is also important in male fertility. Approximately 50% of TGCs become apoptotic and phagocytosed by Sertoli cells. SGG in apoptotic remnants needs to be degraded by Sertoli lysosomal enzymes to the lipid backbone. Failure in this event leads to a lysosomal storage disorder and sub-functionality of Sertoli cells, including their support for TGC development, and consequently subfertility. Significantly, both biosynthesis and degradation pathways of the galactosylsulfate head group of SGG are the same as those of sulfogalactosylceramide (SGC), a structurally related sulfoglycolipid important for brain functions. If subfertility in males with gene mutations in SGG/SGC metabolism pathways manifests prior to neurological disorder, sperm SGG levels might be used as a reporting/predicting index of the neurological status.
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Affiliation(s)
- Nongnuj Tanphaichitr
- Chronic Disease Program, Ottawa Hospital Research Institute, Ottawa, Ontario, Canada; Department of Obstetrics/Gynecology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada; Department of Biochemistry, Microbiology, Immunology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada.
| | - Kessiri Kongmanas
- Chronic Disease Program, Ottawa Hospital Research Institute, Ottawa, Ontario, Canada; Department of Biochemistry, Microbiology, Immunology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada; Division of Dengue Hemorrhagic Fever Research, Department of Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand
| | - Kym F Faull
- Pasarow Mass Spectrometry Laboratory, University of California, Los Angeles, California, USA
| | - Julian Whitelegge
- Pasarow Mass Spectrometry Laboratory, University of California, Los Angeles, California, USA
| | - Federica Compostella
- Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, Via Saldini 50, 20133 Milano, Italy
| | - Naoko Goto-Inoue
- Department of Marine Science and Resources, College of Bioresource Sciences, Nihon University, Kanagawa 252-0880, Japan
| | - James-Jules Linton
- Chronic Disease Program, Ottawa Hospital Research Institute, Ottawa, Ontario, Canada
| | - Brendon Doyle
- Chronic Disease Program, Ottawa Hospital Research Institute, Ottawa, Ontario, Canada; Department of Biochemistry, Microbiology, Immunology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada
| | - Richard Oko
- Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, Canada
| | - Hongbin Xu
- Chronic Disease Program, Ottawa Hospital Research Institute, Ottawa, Ontario, Canada; Department of Biochemistry, Microbiology, Immunology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada
| | - Luigi Panza
- Department of Pharmaceutical Sciences, Università del Piemonte Orientale, Largo Donegani 2, 28100 Novara, Italy
| | - Arpornrad Saewu
- Chronic Disease Program, Ottawa Hospital Research Institute, Ottawa, Ontario, Canada
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Honke K. Biological functions of sulfoglycolipids and the EMARS method for identification of co-clustered molecules in the membrane microdomains. J Biochem 2018; 163:253-263. [PMID: 29186467 DOI: 10.1093/jb/mvx078] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2017] [Accepted: 09/03/2017] [Indexed: 01/04/2025] Open
Abstract
Two major sulfoglycolipids, sulfatide (SO3-3Gal-ceramide) and seminolipid (SO3-3Gal-alkylacylglycerol) exist in mammals. Sulfatide is abundant in the myelin sheath and seminolipid is unique to the spermatogenic cells. The carbohydrate moiety of sulfatide and seminolipid is identical and synthesized by common enzymes: ceramide galactosyltransferase (CGT) and cerebroside sulfotransferase (CST). We have purified CST homogenously, cloned the CST gene and generated CST-knockout mice. CST-null mice completely lack sulfoglycolipids all over the body. Analysis of CST-null mice has revealed that sulfatide is an essential component for the axo-glial junction at the paranode region and regulates terminal differentiation of oligodendrocytes, and that seminolipid is responsible for the formation of a functional lactate transporter assembly to take up the critical energy source for spermatocytes. We have developed a new analytical method termed EMARS to identify co-clustered molecules in the membrane microdomains in order to elucidate the functional molecules that collaborate with sulfoglycolipids.
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Affiliation(s)
- Koichi Honke
- Department of Biochemistry, Kochi University Medical School, Kohasu, Oko-cho, Nankoku, Kochi 783-8505, Japan
- Center for Innovative and Translational Medicine, Kochi University Medical School, Kohasu, Oko-cho, Nankoku, Kochi 783-8505, Japan
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11
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Hunter M, Demarais NJ, Faull RLM, Grey AC, Curtis MA. Layer-specific lipid signatures in the human subventricular zone demonstrated by imaging mass spectrometry. Sci Rep 2018; 8:2551. [PMID: 29416059 PMCID: PMC5803191 DOI: 10.1038/s41598-018-20793-4] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/24/2017] [Accepted: 01/19/2018] [Indexed: 02/01/2023] Open
Abstract
The subventricular zone is a key site of adult neurogenesis and is also implicated in neurodegenerative diseases and brain cancers. In the subventricular zone, cell proliferation, migration and differentiation of nascent stem cells and neuroblasts are regulated at least in part by lipids. The human subventricular zone is distinctly layered and each layer contains discrete cell types that support the processes of neuroblast migration and neurogenesis. We set out to determine the lipid signatures of each subventricular layer in the adult human brain (n = 4). We utilised matrix-assisted laser desorption/ionisation (MALDI) imaging mass spectrometry and liquid chromatography-mass spectrometry to characterise the lipidome of the subventricular zone, with histology and microscopy used for identifying anatomical landmarks. Our findings showed that the subventricular zone was rich in sphingomyelins and phosphatidylserines but deficient in phosphatidylethanolamines. The ependymal layer had an abundance of phosphatidylinositols, whereas the myelin layer was rich in sulfatides and triglycerides. The hypocellular layer showed enrichment of sphingomyelins. No discrete lipid signature was seen in the astrocytic ribbon. The biochemical functions of these lipid classes are consistent with the localisation we observed within the SVZ. Our study may, therefore, shed new light on the role of lipids in the regulation of adult neurogenesis.
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Affiliation(s)
- Mandana Hunter
- Centre for Brain Research, University of Auckland, Auckland, 1023, New Zealand.,Department of Pharmacology and Clinical Pharmacology, Faculty of Medical and Health Sciences, University of Auckland, Auckland, 1023, New Zealand
| | - Nicholas J Demarais
- School of Biological Sciences, Faculty of Science, University of Auckland, Auckland, 1010, New Zealand
| | - Richard L M Faull
- Centre for Brain Research, University of Auckland, Auckland, 1023, New Zealand.,Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, University of Auckland, Auckland, 1023, New Zealand
| | - Angus C Grey
- Centre for Brain Research, University of Auckland, Auckland, 1023, New Zealand.,Department of Physiology, Faculty of Medical and Health Sciences, University of Auckland, Auckland, 1023, New Zealand
| | - Maurice A Curtis
- Centre for Brain Research, University of Auckland, Auckland, 1023, New Zealand. .,Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, University of Auckland, Auckland, 1023, New Zealand.
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Zhou Z, Thiagarajan P, Udden M, López J, Guchhait P. Erythrocyte membrane sulfatide plays a crucial role in the adhesion of sickle erythrocytes to endothelium. Thromb Haemost 2017; 105:1046-52. [DOI: 10.1160/th10-11-0716] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2010] [Accepted: 02/03/2011] [Indexed: 11/05/2022]
Abstract
SummaryEnhanced adhesion of sickle erythrocytes to the vascular endothelium and subendothelial matrix is fundamental to the development of vascular occlusion in sickle cell disease. Erythrocyte membrane sulfatide is implicated in the pathogenesis of vasoocclusive crises in sickle cell disease (SCD) patients. Because previous evidence linking sulfatide to cell adhesion has largely been circumstantial due to a lack of reagents that specifically target sulfatide, we used two sulfatide-specific strategies to address the role of erythrocyte membrane sulfatide in sickle cell adhesion to the vascular endothelium: a single-chain fragment variable chain (scFv) antibody against sulfatide as well as cerebroside sulfotransferase-deficient mice incapable of synthesising sulfatide. The sickle erythrocytes from mice and humans adhered at a greater extent and at higher shear stresses to activated endothelium than normal erythrocytes, and approximately 60% of the adhesion was prevented by the anti-sulfatide scFv. Similarly, the extent of adhesion of sulfatide-deficient erythrocytes was lower than normal erythrocytes. These findings suggest an important role for membrane sulfatide in sickle cell disease pathophysiology.
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Identification of a new liver-specific c-type mRNA transcriptional variant for mouse ST3GAL5 (GM3/GM4 synthase). Glycoconj J 2017; 34:651-659. [DOI: 10.1007/s10719-017-9788-1] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/22/2017] [Accepted: 07/26/2017] [Indexed: 10/19/2022]
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Abstract
Sulfatide is a 3-O-sulfated galactosylceramide that is abundantly expressed in the gastrointestinal tract, kidney, trachea, and particularly the central nervous system. Cellular sulfatide is mainly localized in the Golgi apparatus, cellular membrane, and lysosomes in cytosol. Since our earlier report showed that the influenza A virus specifically binds to sulfatide, we have investigated the roles of sulfatide in the influenza A virus lifecycle. The viral binding is independent of sialic acids, which function as virus receptors in virus attachment to the host cell surface. Sulfatide is recognized by the ectodomain of the viral envelope glycoprotein hemagglutinin (HA). Nascent HA is transported on the surface membrane of infected cells. The binding of HA with sulfatide on the cell surface induces apoptosis through potential loss of the mitochondrial membrane and nuclear translocation of apoptosis-inducing factor in mitochondria, where PB1-F2 peptide from the viral gene is accumulated. In the nucleus of infected cells, viral ribonucleoprotein (vRNP) complexes are formed from viral RNA genomes, viral nucleoprotein, and viral RNA polymerase subunits, and these complexes are selectively exported into cytosol through the nuclear membrane. The apoptosis significantly enhances the nuclear export of vRNP complexes, resulting in efficient formation of progeny viruses and facilitation of virus replication. At that time, activation of the Raf/mitogen-activated protein extracellular kinase (MEK)/extracellular signal-regulated kinase (ERK) pathway through sulfatide is associated with virus replication. Our studies have demonstrated that sulfatide is not a viral receptor for virus infection, and that the binding of HA with sulfatide functions as an initiation switch for the formation of progeny viruses.
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Affiliation(s)
- Tadanobu Takahashi
- Department of Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka
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15
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Functional analysis of glyco-molecules that bind with influenza virus. Uirusu 2016; 66:101-116. [PMID: 28484173 DOI: 10.2222/jsv.66.101] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/19/2022]
Abstract
Influenza A virus (IAV) recognizes terminal sialic acid of sialoglyco-conjugates on host cells through the viral envelope glycoprotein hemagglutinin (HA), followed by initiation of entry into the cells. Molecular species of sialic acid are largely divided into two moieties: N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc). A receptor for IAV infection generally means Neu5Ac. Almost all equine IAVs and some human, swine, and duck IAVs bind not only to Neu5Ac but also to Neu5Gc. In nonhuman animals, Neu5Gc has been detected in swine and equine tracheas and the duck colon, which are the main replication sites of mammalian and avian IAVs. Therefore, Neu5Gc in these sites has been suggested to be a functional receptor for IAV infection. Humans cannot synthesize Neu5Gc due to a genetic defect of the Neu5Gc-synthesizing enzyme. We evaluated the receptor function of Neu5Gc in IAV infection in human cells. Our results indicated that Neu5Gc expression on the surface of human cells is not a functional receptor for IAV infection and that it has a negative effect on infectivity of IAV possessing Neu5Gc binding ability. IAV also binds to non-sialo 3-O-sulfated galactosylceramide (sulfatide). Sulfatide has been suggested to be a functional receptor for IAV infection. However, we have shown that sulfatide is not a functional receptor for IAV infection and that the binding of HA with sulfatide enhances progeny virus production. It is expected that functions of these glyco-molecules can be used in prevention and development of new drugs against IAV.
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Allende ML, Proia RL. Simplifying complexity: genetically resculpting glycosphingolipid synthesis pathways in mice to reveal function. Glycoconj J 2014; 31:613-22. [PMID: 25351657 PMCID: PMC4245496 DOI: 10.1007/s10719-014-9563-5] [Citation(s) in RCA: 43] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/03/2014] [Accepted: 10/03/2014] [Indexed: 11/30/2022]
Abstract
Glycosphingolipids (GSLs) are a group of plasma-membrane lipids notable for their extremely diverse glycan head groups. The metabolic pathways for GSLs, including the identity of the biosynthetic enzymes needed for synthesis of their glycans, are now well understood. Many of their cellular functions, which include plasma-membrane organization, regulation of cell signaling, endocytosis, and serving as binding sites for pathogens and endogenous receptors, have also been established. However, an understanding of their functions in vivo had been lagging. Studies employing genetic manipulations of the GSL synthesis pathways in mice have been used to systematically reduce the large numbers and complexity of GSL glycan structures, allowing the in vivo functions of GSLs to be revealed from analysis of the resulting phenotypes. Findings from these studies have produced a clearer picture of the role of GSLs in mammalian physiology, which is the topic of this review.
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Affiliation(s)
- Maria Laura Allende
- Genetics of Development and Disease Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Building 10, Room 9D-06; 10 Center DR MSC 1821, Bethesda, MD, 20892-1821, USA
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Honke K. Galactose-3-O-Sulfotransferase 1-4 (GAL3ST1-4). HANDBOOK OF GLYCOSYLTRANSFERASES AND RELATED GENES 2014:1123-1134. [DOI: 10.1007/978-4-431-54240-7_56] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/04/2025]
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18
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Nakajima T, Kamijo Y, Yuzhe H, Kimura T, Tanaka N, Sugiyama E, Nakamura K, Kyogashima M, Hara A, Aoyama T. Peroxisome proliferator-activated receptor α mediates enhancement of gene expression of cerebroside sulfotransferase in several murine organs. Glycoconj J 2013; 30:553-560. [PMID: 23065187 DOI: 10.1007/s10719-012-9454-6] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2012] [Revised: 09/26/2012] [Accepted: 10/01/2012] [Indexed: 12/28/2022]
Abstract
Sulfatides, 3-O-sulfogalactosylceramides, are known to have multifunctional properties. These molecules are distributed in various tissues of mammals, where they are synthesized from galactosylceramides by sulfation at C3 of the galactosyl residue. Although this reaction is specifically catalyzed by cerebroside sulfotransferase (CST), the mechanisms underlying the transcriptional regulation of this enzyme are not understood. With respect to this issue, we previously found potential sequences of peroxisome proliferator-activated receptor (PPAR) response element on upstream regions of the mouse CST gene and presumed the possible regulation by the nuclear receptor PPARα. To confirm this hypothesis, we treated wild-type and Ppara-null mice with the specific PPARα agonist fenofibrate and examined the amounts of sulfatides and CST gene expression in various tissues. Fenofibrate treatment increased sulfatides and CST mRNA levels in the kidney, heart, liver, and small intestine in a PPARα-dependent manner. However, these effects of fenofibrate were absent in the brain or colon. Fenofibrate treatment did not affect the mRNA level of arylsulfatase A, which is the key enzyme for catalyzing desulfation of sulfatides, in any of these six tissues. Analyses of the DNA-binding activity and conventional gene expression targets of PPARα has demonstrated that fenofibrate treatment activated PPARα in the kidney, heart, liver, and small intestine but did not affect the brain or colon. These findings suggest that PPARα activation induces CST gene expression and enhances sulfatide synthesis in mice, which suggests that PPARα is a possible transcriptional regulator for the mouse CST gene.
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Affiliation(s)
- Takero Nakajima
- Department of Metabolic Regulation, Institute of Pathogenesis and Disease Prevention, Shinshu University Graduate School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano, 390-8621, Japan
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19
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Acosta DM, Soprano LL, Ferrero MR, Esteva MI, Riarte A, Couto AS, Duschak VG. Structural and immunological characterization of sulphatides: relevance of sulphate moieties in Trypanosoma cruzi glycoconjugates. Parasite Immunol 2013; 34:499-510. [PMID: 22738032 DOI: 10.1111/j.1365-3024.2012.01378.x] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/01/2022]
Abstract
Sulphoglycosphingolipids, present on the surface of diverse cells, participate in the regulation of various cellular events. However, little is known about the structure and the role of sulphoglycosphingolipids in trypanosomatids. Herein, sulphated dihexosylceramide structures - composed mainly of sphingosine as the long chain base acylated with stearic acid - have been determined for the first time in Trypanosoma cruzi epimastigotes by UV-MALDI-TOF-MS analysis. Interestingly, inhibition ELISA assays using cruzipain as antigen and polyclonal rabbit antibodies specific for cruzipain, the major cysteine proteinase of T. cruzi, or for its C-terminal domain, have demonstrated (i) that sulphate epitopes are shared between cruzipain and sulphatides of T. cruzi, (ii) that cross-reactivity maps to the C-terminal domain and (iii) the existence of other antigenic determinants in the glycolipidic structures. These features provide evidence that sulphate groups are antigenic in sulphate-containing parasite glycoconjugates. Furthermore, IgG2 antibody levels inversely correlate with disease severity in chronic Chagas disease patients, suggesting that IgG2 antibodies specific for sulphated epitopes might be associated with protective immunity and might be considered as potential surrogates of the course of chronic Chagas disease.
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Affiliation(s)
- D M Acosta
- Instituto Nacional de Parasitología Dr Mario Fatala Chaben, ANLIS-Malbrán, Ministerio de Salud de la Nación, Buenos Aires, Argentina
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20
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Honke K. Biosynthesis and biological function of sulfoglycolipids. PROCEEDINGS OF THE JAPAN ACADEMY. SERIES B, PHYSICAL AND BIOLOGICAL SCIENCES 2013; 89:129-138. [PMID: 23574804 PMCID: PMC3669731 DOI: 10.2183/pjab.89.129] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 01/18/2013] [Accepted: 02/19/2013] [Indexed: 06/02/2023]
Abstract
Sulfation confers negative charge on glycolipids and the attached sulfate group presents a part of determinants for the molecular interactions. Mammalian sulfoglycolipids are comprised of two major members, sulfatide (SO3-3Gal-ceramide) and seminolipid (SO3-3Gal-alkylacylglycerol). Sulfatide is abundant in the myelin sheath and seminolipid is unique to the spermatogenic cells. The carbohydrate moiety of sulfatide and seminolipid is biosynthesized via sequential reactions catalyzed by common enzymes: ceramide galactosyltransferase (CGT) and cerebroside sulfotransferase (CST). To elucidate the biological function of sulfoglycolipids, we have purified CST, cloned the CST gene, and generated CST-knockout mice. CST-null mice completely lack sulfoglycolipids all over the body. CST-null mice manifest some neurological disorders due to myelin dysfunction, an aberrant enhancement of oligodendrocyte terminal differentiation, and an arrest of spermatogenesis. CST-deficiency ameliorates L-selectin-dependent monocyte infiltration in the renal interstitial inflammation, indicating that sulfatide is an endogenous ligand of L-selectin. Studies on the molecular mechanisms underlying the biological events for which sulfoglycolipids are essential are ongoing
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Affiliation(s)
- Koichi Honke
- Department of Biochemistry and Kochi System Glycobiology Center, Kochi University Medical School, Kochi, Japan.
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The Enigmatic Role of Sulfatides: New Insights into Cellular Functions and Mechanisms of Protein Recognition. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2013; 991:27-40. [DOI: 10.1007/978-94-007-6331-9_3] [Citation(s) in RCA: 32] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
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Takahashi T, Ito K, Fukushima K, Takaguchi M, Hayakawa T, Suzuki Y, Suzuki T. Sulfatide negatively regulates the fusion process of human parainfluenza virus type 3. J Biochem 2012; 152:373-80. [DOI: 10.1093/jb/mvs080] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
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Takahashi T, Suzuki T. Role of sulfatide in normal and pathological cells and tissues. J Lipid Res 2012; 53:1437-50. [PMID: 22619219 DOI: 10.1194/jlr.r026682] [Citation(s) in RCA: 211] [Impact Index Per Article: 16.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Sulfatide is 3-O-sulfogalactosylceramide that is synthesized by two transferases (ceramide galactosyltransferase and cerebroside sulfotransferase) from ceramide and is specifically degraded by a sulfatase (arylsulfatase A). Sulfatide is a multifunctional molecule for various biological fields including the nervous system, insulin secretion, immune system, hemostasis/thrombosis, bacterial infection, and virus infection. Therefore, abnormal metabolism or expression change of sulfatide could cause various diseases. Here, we discuss the important biological roles of sulfatide in the nervous system, insulin secretion, immune system, hemostasis/thrombosis, cancer, and microbial infections including human immunodeficiency virus and influenza A virus. Our review will be helpful to achieve a comprehensive understanding of sulfatide, which serves as a fundamental target of prevention of and therapy for nervous disorders, diabetes mellitus, immunological diseases, cancer, and infectious diseases.
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Affiliation(s)
- Tadanobu Takahashi
- Department of Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka and Global COE Program for Innovation in Human Health Sciences, 52-1 Yada, Suruga-ku, Shizuoka-shi, Shizuoka 422-8526, Japan
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Kimura T, Nakajima T, Kamijo Y, Tanaka N, Wang L, Hara A, Sugiyama E, Tanaka E, Gonzalez FJ, Aoyama T. Hepatic Cerebroside Sulfotransferase Is Induced by PPARα Activation in Mice. PPAR Res 2012; 2012:174932. [PMID: 22645601 PMCID: PMC3356938 DOI: 10.1155/2012/174932] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/04/2012] [Accepted: 02/16/2012] [Indexed: 11/23/2022] Open
Abstract
Sulfatides are one of the major sphingoglycolipids in mammalian serum and are synthesized and secreted mainly from the liver as a component of lipoproteins. Recent studies revealed a protective role for serum sulfatides against arteriosclerosis and hypercoagulation. Although peroxisome proliferator-activated receptor (PPAR) α has important functions in hepatic lipoprotein metabolism, its association with sulfatides has not been investigated. In this study, sulfatide levels and the expression of enzymes related to sulfatide metabolism were examined using wild-type (+/+), Ppara-heterozygous (+/-), and Ppara-null (-/-) mice given a control diet or one containing 0.1% fenofibrate, a clinically used hypolipidemic drug and PPARα activator. Fenofibrate treatment increased serum and hepatic sulfatides in Ppara (+/+) and (+/-) mice through a marked induction of hepatic cerebroside sulfotransferase (CST), a key enzyme in sulfatide synthesis, in a PPARα-dependent manner. Furthermore, increases in CST mRNA levels were correlated with mRNA elevations of several known PPARα target genes, and such changes were not observed for other sulfatide-metabolism enzymes in the liver. These results suggest that PPARα activation enhances hepatic sulfatide synthesis via CST induction and implicate CST as a novel PPARα target gene.
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Affiliation(s)
- Takefumi Kimura
- Department of Metabolic Regulation, Institute on Aging and Adaptation, Shinshu University Graduate School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
- Department of Gastroenterology, Shinshu University School of Medicine, Matsumoto 390-8621, Japan
| | - Takero Nakajima
- Department of Metabolic Regulation, Institute on Aging and Adaptation, Shinshu University Graduate School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
| | - Yuji Kamijo
- Department of Metabolic Regulation, Institute on Aging and Adaptation, Shinshu University Graduate School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
- Department of Nephrology, Shinshu University School of Medicine, Matsumoto 390-8621, Japan
| | - Naoki Tanaka
- Department of Metabolic Regulation, Institute on Aging and Adaptation, Shinshu University Graduate School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
- Laboratory of Metabolism, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
| | - Lixuan Wang
- Department of Metabolic Regulation, Institute on Aging and Adaptation, Shinshu University Graduate School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
| | - Atsushi Hara
- Department of Metabolic Regulation, Institute on Aging and Adaptation, Shinshu University Graduate School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
| | - Eiko Sugiyama
- Department of Nutritional Science, Nagano Prefectural College, Nagano 380-8525, Japan
| | - Eiji Tanaka
- Department of Gastroenterology, Shinshu University School of Medicine, Matsumoto 390-8621, Japan
| | - Frank J. Gonzalez
- Laboratory of Metabolism, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
| | - Toshifumi Aoyama
- Department of Metabolic Regulation, Institute on Aging and Adaptation, Shinshu University Graduate School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
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Sheng X, Nakajima T, Wang L, Zhang X, Kamijo Y, Takahashi K, Tanaka N, Sugiyama E, Kyogashima M, Aoyama T, Hara A. Attenuation of kidney injuries maintains serum sulfatide levels dependent on hepatic synthetic ability: a possible involvement of oxidative stress. TOHOKU J EXP MED 2012; 227:1-12. [PMID: 22499158 DOI: 10.1620/tjem.227.1] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022]
Abstract
Serum sulfatides are the major glycosphingolipids in lipoproteins. Although serum sulfatides are mainly synthesized and secreted by the liver, they are significantly decreased when the kidneys are impaired. Our recent experimental study using a murine protein-overload nephropathy model suggested a hypothetical mechanism whereby serum sulfatides were reduced due to kidney dysfunction. This was the result of decreased hepatic expression of a sulfatide synthetic enzyme, cerebroside sulfotransferase (CST), which is associated with systemic enhancement of oxidative stress. However, there is a possibility that the experimental process, protein-overload itself, directly affected the sulfatide metabolism and oxidative stress in the liver. To determine whether kidney dysfunction actually reduces the hepatic synthesis of sulfatides via oxidative stress, we examined sulfatide levels, the hepatic content of metabolic sulfatide enzymes, and the degree of oxidative stress in protein-overload mice subjected to renoprotective therapy using clofibrate, a representative hypolipidemic medicine. Protein-overload mice exhibited marked kidney injuries, enhancement of hepatic oxidative stress, decreased levels of serum and hepatic sulfatides, and decreased expression of hepatic CST. The clofibrate treatment attenuated kidney damage and hepatic oxidative stress while maintaining serum/hepatic sulfatide levels and hepatic CST content in the mice. Because clofibrate monotherapy without protein-overload treatment only minimally affected these hepatic parameters, the hepatic synthesis of sulfatides appeared to be strongly influenced by kidney dysfunction and subsequent oxidative stress. This study suggests that the crosstalk between kidney dysfunction and hepatic sulfatide metabolism is mediated by oxidative stress. These results should help to understand the phenomenon in patients with end-stage kidney disease.
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Affiliation(s)
- Xiaona Sheng
- Department of Metabolic Regulation, Institute on Aging and Adaptation, Shinshu University Graduate School of Medicine, Matsumoto, Japan
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Pugacheva EM, Suzuki T, Pack SD, Kosaka-Suzuki N, Yoon J, Vostrov AA, Barsov E, Strunnikov AV, Morse HC, Loukinov D, Lobanenkov V. The structural complexity of the human BORIS gene in gametogenesis and cancer. PLoS One 2010; 5:e13872. [PMID: 21079786 PMCID: PMC2975627 DOI: 10.1371/journal.pone.0013872] [Citation(s) in RCA: 47] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2010] [Accepted: 10/11/2010] [Indexed: 12/14/2022] Open
Abstract
BACKGROUND BORIS/CTCFL is a paralogue of CTCF, the major epigenetic regulator of vertebrate genomes. BORIS is normally expressed only in germ cells but is aberrantly activated in numerous cancers. While recent studies demonstrated that BORIS is a transcriptional activator of testis-specific genes, little is generally known about its biological and molecular functions. METHODOLOGY/PRINCIPAL FINDINGS Here we show that BORIS is expressed as 23 isoforms in germline and cancer cells. The isoforms are comprised of alternative N- and C-termini combined with varying numbers of zinc fingers (ZF) in the DNA binding domain. The patterns of BORIS isoform expression are distinct in germ and cancer cells. Isoform expression is activated by downregulation of CTCF, upregulated by reduction in CpG methylation caused by inactivation of DNMT1 or DNMT3b, and repressed by activation of p53. Studies of ectopically expressed isoforms showed that all are translated and localized to the nucleus. Using the testis-specific cerebroside sulfotransferase (CST) promoter and the IGF2/H19 imprinting control region (ICR), it was shown that binding of BORIS isoforms to DNA targets in vitro is methylation-sensitive and depends on the number and specific composition of ZF. The ability to bind target DNA and the presence of a specific long amino terminus (N258) in different isoforms are necessary and sufficient to activate CST transcription. Comparative sequence analyses revealed an evolutionary burst in mammals with strong conservation of BORIS isoproteins among primates. CONCLUSIONS The extensive repertoire of spliced BORIS variants in humans that confer distinct DNA binding and transcriptional activation properties, and their differential patterns of expression among germ cells and neoplastic cells suggest that the gene is involved in a range of functionally important aspects of both normal gametogenesis and cancer development. In addition, a burst in isoform diversification may be evolutionarily tied to unique aspects of primate speciation.
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Affiliation(s)
- Elena M Pugacheva
- Laboratory of Immunopathology, National Institute of Allergy and Infectious Disease, National Institutes of Health, Rockville, Maryland, United States of America.
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Expression of a testis-specific form of Gal3st1 (CST), a gene essential for spermatogenesis, is regulated by the CTCF paralogous gene BORIS. Mol Cell Biol 2010; 30:2473-84. [PMID: 20231363 DOI: 10.1128/mcb.01093-09] [Citation(s) in RCA: 61] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Previously, it was shown that the CTCF paralogous gene, BORIS (brother of the regulator of imprinted sites) is expressed in male germ cells, but its function in spermatogenesis has not been defined. To develop an understanding of the functional activities of BORIS, we generated BORIS knockout (KO) mice. Mice homozygous for the null allele had a defect in spermatogenesis that resulted in small testes associated with increased cell death. The defect was evident as early as postnatal day 21 and was manifested by delayed production of haploid cells. By gene expression profiling, we found that transcript levels for Gal3st1 (also known as cerebroside sulfotransferase [CST]), known to play a crucial role in meiosis, were dramatically reduced in BORIS KO testes. We found that CST is expressed in testis as a novel testis-specific isoform, CST form F(TS), that has a short exon 1f. We showed that BORIS bound to and activated the promoter of CST form F(TS). Mutation of the BORIS binding site in the promoter reduced the ability of BORIS to activate the promoter. These findings define transcriptional regulation of CST expression as a critical role for BORIS in spermatogenesis.
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Zhang X, Nakajima T, Kamijo Y, Li G, Hu R, Kannagi R, Kyogashima M, Aoyama T, Hara A. Acute kidney injury induced by protein-overload nephropathy down-regulates gene expression of hepatic cerebroside sulfotransferase in mice, resulting in reduction of liver and serum sulfatides. Biochem Biophys Res Commun 2009; 390:1382-8. [PMID: 19895791 DOI: 10.1016/j.bbrc.2009.10.164] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2009] [Accepted: 10/30/2009] [Indexed: 11/27/2022]
Abstract
Sulfatides, possible antithrombotic factors belonging to sphingoglycolipids, are widely distributed in mammalian tissues and serum. We recently found that the level of serum sulfatides was significantly lower in hemodialysis patients than that in normal subjects, and that the serum level closely correlated to the incidence of cardiovascular disease. These findings suggest a relationship between the level of serum sulfatides and kidney function; however, the molecular mechanism underlying this relationship remains unclear. In the present study, the influence of kidney dysfunction on the metabolism of sulfatides was examined using an established murine model of acute kidney injury, protein-overload nephropathy in mice. Protein-overload treatment caused severe proximal tubular injuries within 4days, and this treatment obviously decreased both serum and hepatic sulfatide levels. The sphingoid composition of serum sulfatides was very similar to that of hepatic ones at each time point, suggesting that the serum sulfatide level is dependent on the hepatic secretory ability of sulfatides. The treatment also decreased hepatic expression of cerebroside sulfotransferase (CST), a key enzyme in sulfatide metabolism, while it scarcely influenced the expression of the other sulfatide-metabolizing enzymes, including arylsulfatase A, ceramide galactosyltransferase, and galactosylceramidase. Pro-inflammatory responses were not detected in the liver of these mice; however, potential oxidative stress was increased. These results suggest that down-regulation of hepatic CST expression, probably affected by oxidative stress from kidney injury, causes reduction in liver and serum sulfatide levels. This novel mechanism, indicating the crosstalk between kidney injury and specific liver function, may prove useful for helping to understand the situation where human hemodialysis patients have low levels of serum sulfatides.
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Affiliation(s)
- Xiaowei Zhang
- Department of Metabolic Regulation, Institute on Aging and Adaptation, Shinshu University Graduate School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan
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Nagai KI, Tadano-Aritomi K, Niimura Y, Ishizuka I. Development and application of a system for seminolipid metabolism using mouse seminiferous tubules. Glycoconj J 2009; 27:181-7. [DOI: 10.1007/s10719-009-9250-0] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/24/2009] [Revised: 06/11/2009] [Accepted: 06/11/2009] [Indexed: 10/20/2022]
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The Role and Metabolism of Sulfatide in the Nervous System. Mol Neurobiol 2008; 37:93-103. [DOI: 10.1007/s12035-008-8022-3] [Citation(s) in RCA: 113] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/25/2008] [Accepted: 04/09/2008] [Indexed: 12/16/2022]
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Abstract
Sulfatide is abundantly expressed in various mammalian organs, including the intestines and trachea, in which influenza A viruses (IAVs) replicate. However, the function of sulfatide in IAV infection remains unknown. Sulfatide is synthesized by two transferases, ceramide galactosyltransferase (CGT) and cerebroside sulfotransferase (CST), and is degraded by arylsulfatase A (ASA). In this study, we demonstrated that sulfatide enhanced IAV replication through efficient translocation of the newly synthesized IAV nucleoprotein (NP) from the nucleus to the cytoplasm, by using genetically produced cells in which sulfatide expression was down-regulated by RNA interference against CST mRNA or overexpression of the ASA gene and in which sulfatide expression was up-regulated by overexpression of both the CST and CGT genes. Treatment of IAV-infected cells with an antisulfatide monoclonal antibody (MAb) or an anti-hemagglutinin (HA) MAb, which blocks the binding of IAV and sulfatide, resulted in a significant reduction in IAV replication and accumulation of the viral NP in the nucleus. Furthermore, antisulfatide MAb protected mice against lethal challenge with pathogenic influenza A/WSN/33 (H1N1) virus. These results indicate that association of sulfatide with HA delivered to the cell surface induces translocation of the newly synthesized IAV ribonucleoprotein complexes from the nucleus to the cytoplasm. Our findings provide new insights into IAV replication and suggest new therapeutic strategies.
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Yaghootfam A, Sorkalla T, Häberlein H, Gieselmann V, Kappler J, Eckhardt M. Cerebroside Sulfotransferase Forms Homodimers in Living Cells. Biochemistry 2007; 46:9260-9. [PMID: 17658888 DOI: 10.1021/bi700014q] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2023]
Abstract
Cerebroside sulfotransferase (CST) catalyzes the 3'-sulfation of galactose residues in several glycolipids. Its major product in the mammalian brain is sulfatide, which is an essential myelin component. Using epitope-tagged variants, murine CST was found to localize to the Golgi apparatus, but in contrast to previous assumptions, not to the trans-Golgi network. An examination of enhanced green fluorescent protein (EGFP)-tagged CST suggests that CST forms homodimers and that dimerization is mediated by the lumenal domain of the enzyme, as shown by immunoprecipitation and density gradient centrifugation. In order to verify that dimerization of CST observed by biochemical methods reflects the behavior of the native protein within living cells, the mobility of CST-EGFP was examined using fluorescence correlation spectroscopy. These experiments confirmed the homodimerization of CST-EGFP fusion proteins in vivo. In contrast to full-length CST, a fusion protein of the amino-terminal 36 amino acids of CST fused to EGFP was exclusively found as a monomer but nevertheless showed Golgi localization.
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Affiliation(s)
- Afshin Yaghootfam
- Institut für Physiologische Chemie, Rheinische Friedrich-Wilhelms-Universität Bonn, Germany
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Garcia J, Callewaert N, Borsig L. P-selectin mediates metastatic progression through binding to sulfatides on tumor cells. Glycobiology 2006; 17:185-96. [PMID: 17043066 DOI: 10.1093/glycob/cwl059] [Citation(s) in RCA: 63] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
Hematogenous carcinoma metastasis is associated with tumor cell emboli formation, which is now known to be facilitated by selectins. P-selectin-mediated interactions of platelets with cancer cells are based mostly on mucin- and glycosaminoglycan-type selectin ligands. We previously showed that mouse colon carcinoma cells (MC-38) carry P-selectin ligands of nonmucin origin, which were not identified. Here we show that P-selectin ligands recognized on MC-38 cells are sulfated glycolipids, thereby facilitating experimental metastasis in a syngeneic mouse model. Metabolic inhibition of sulfation by incubation of cells with sodium chlorate almost completely abrogated P-selectin binding. Metabolic labeling of MC-38 cells with (35)S sulfate revealed only a single band as detected by high-performance thin layer chromatography analysis of a total lipid extract. Matrix-assisted laser desorption/ionization tandem time-of-flight/time-of-flight analysis (MALDI-TOF-TOF) analysis of the purified sulfate-containing lipid fraction identified the selectin ligand to be a sulfated galactosylceramide SM4 (HSO(3)-3Galbeta-1Cer). Modulation of glycolipid biosynthesis in MC-38 cells altered P-selectin binding, thereby confirming sulfoglycolipids to be major P-selectin ligands. In addition, P-selectin was also found to recognize lactosylceramide sulfate SM3 (HSO(3)-3Galbeta-4Glcbeta-1Cer) and gangliotriaosylceramide sulfate SM2 [GalNAcbeta-4(HSO(3)-3)Galbeta-4Glcbeta-1Cer] in human hepatoma cells. Finally, the enzymatic removal of sulfation from the cell surface of MC-38 cells resulted in decreased P-selectin binding and led to attenuation of metastasis. Thus, SM4 sulfatide serves as a native ligand for P-selectin contributing to cell-cell interactions and to facilitation of metastasis.
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Affiliation(s)
- Josep Garcia
- Center for Integrative Human Physiology, University of Zürich, Zürich, Switzerland
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Wu XZ, Zhang L, Shi BZ, Hu P. Inhibitory effects of N-(4-hydrophenyl) retinamide on liver cancer and malignant melanoma cells. World J Gastroenterol 2005; 11:5763-9. [PMID: 16270382 PMCID: PMC4479673 DOI: 10.3748/wjg.v11.i37.5763] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the effect of N-(4-hydrophenyl) retinamide (4-HPR), the derivative of retinoic acid, on inhibition of migration, invasion, cell growth, and induction of apoptosis in hepatocellular carcinoma cells (HCCs) and malignant melanoma cells.
METHODS: 4-HPR was chemically synthesized. Cellular migration and invasion were assayed by Borden chamber experiment. Cell growth was assayed by MTT chromometry. Apoptosis effect was measured using Hoechst 32258 staining and flow cytometry. Gene transfection was performed with lipofectamine.
RESULTS: We observed that the migration of HCC and melanoma cells was significantly suppressed by 4-HPR and the migration cells were reduced to 585.03 (control 20127.2, P < 0.05, n = 4) in SMMC 7721-k3 HCC, and to 25425.04 (control 30230.1, P < 0.05, n = 4) in melanoma cells after 6-h incubation with 4-HPR. The invasion through reconstituted basement membrane was also significantly reduced by 4-HPR treatment to 11.23.3 in SMMC 7721-k3 HCC (control 2713.1), and to 24.33.2 in melanoma cells (control 67.510.1, P < 0.05, n = 3). Cell growth, especially in melanoma cells, was also significantly inhibited. Furthermore, 3 mmol/L of 4-HPR induced apoptosis in B16 melanoma cells (37.110.94%) more significantly than all-trans retinoic acid (P < 0.05), but it failed to induce apoptosis in SMMC 7721-k3 HCC. The mechanism for 4-HPR-induced apoptosis was not clear, but we observed that 4-HPR could regulate p27kip1, and overexpression of cerebroside sulfotransferase (CST) diminished the apoptosis induced by 4-HPR in melanoma cells.
CONCLUSION: 4-HPR is a potent inhibitor of HCC migration and inducer of melanoma cell apoptosis. CST and p27kip1expression might be associated with 4-HPR-induced apoptosis.
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Affiliation(s)
- Xing-Zhong Wu
- Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai 200032, China.
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Sandhoff R, Grieshaber H, Djafarzadeh R, Sijmonsma TP, Proudfoot AEI, Handel TM, Wiegandt H, Nelson PJ, Gröne HJ. Chemokines bind to sulfatides as revealed by surface plasmon resonance. Biochim Biophys Acta Mol Cell Biol Lipids 2005; 1687:52-63. [PMID: 15708353 DOI: 10.1016/j.bbalip.2004.11.011] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2004] [Revised: 11/05/2004] [Accepted: 11/08/2004] [Indexed: 11/18/2022]
Abstract
Chemokines bind to sulfated cell surface glycosaminoglycans and thereby modulate signaling mediated by G-protein-coupled seven-transmembrane domain chemokine receptors. Similar to glycosaminoglycans, sulfated oligosaccharides are also exposed on the cell surface by sulfatides, a class of glycosphingolipids. We have now identified sulfated glycosphingolipids (sulfatides) as novel binding partners for chemokines. Using surface plasmon resonance (SPR), the binding of proinflammatory and homeostatic chemokines to glycosphingolipids, in particular sulfatides, was investigated. Chemokines were immobilized while glycosphingolipids or additional phospholipids incorporated into liposomes were applied as soluble analytes. A specific affinity of the chemokines MCP-1/CCL2, IL-8/CXCL8, SDF-1alpha/CXCL12, MIP-1alpha/CCL3 and MIP-1beta/CCL4 to the sulfatides SM4s, SM3, SM2a and SB2, SB1a was detected. No significant interactions with the chemokines were observed for gangliosides, neutral glycosphingolipids or phospholipids. Chemokine receptors have been associated with the detergent-insoluble fraction supposed to contain 'rafts', i.e., glycosphingolipid enriched microdomains of the cell surface. Accordingly, the data suggest that early chemokine receptor signaling may take place in the vicinity of sulfated glycosphingolipids on the cell surface, whereby these sulfatides could modulate the chemokine receptor-mediated cell activation signal.
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Affiliation(s)
- Roger Sandhoff
- German Cancer Research Center, Department of Cellular and Molecular Pathology, INF 280, 69120 Heidelberg, Germany.
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Honke K, Zhang Y, Cheng X, Kotani N, Taniguchi N. Biological roles of sulfoglycolipids and pathophysiology of their deficiency. Glycoconj J 2005; 21:59-62. [PMID: 15467400 DOI: 10.1023/b:glyc.0000043749.06556.3d] [Citation(s) in RCA: 53] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
Mammalian sulfoglycolipids are comprised of two major members, sulfatide (SO(3)-3Gal-ceramide) and seminolipid (SO(3)-3Gal-alkylacylglycerol). Sulfatide is abundant in the myelin sheath and seminolipid is expressed on the spermatogenic cells. Cerebroside sulfotransferase (CST)-deficient mice generated by gene targeting completely lack sulfatide and seminolipid all over the body. CST-null mice manifest some neurological disorders due to myelin dysfunction, an aberrant enhancement of oligodendrocyte terminal differentiation, and an arrest of spermatogenesis, indicating that sulfation of glycolipids is essential for myelin formation and spermatogenesis. Moreover, CST-deficiency ameliorates L-selectin-dependent monocyte infiltration in the kidney after ureteral obstruction, an experimental model of renal interstitial inflammation, indicating that sulfatide is an endogenous ligand of L-selectin. Studies on the molecular mechanisms by which sulfoglycolipids participate in these biological processes are ongoing.
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Affiliation(s)
- Koichi Honke
- Department of Molecular Genetics, Kochi University Medical School, Kohasu, Oko-cho, Nankoku, Kochi 783-8505, Japan.
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Nagai KI, Tadano-Aritomi K, Iida-Tanaka N, Yoshizawa H, Ishizuka I. Metabolism of sulfolipids in isolated renal tubules from rat. Comp Biochem Physiol B Biochem Mol Biol 2005; 140:487-95. [PMID: 15694597 DOI: 10.1016/j.cbpc.2004.11.013] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2004] [Revised: 11/16/2004] [Accepted: 11/17/2004] [Indexed: 11/24/2022]
Abstract
Proximal-rich tubules were prepared from rat kidneys by using collagenase treatment. The isolated rat renal tubules were compared with the intact kidney on the following characteristics. (1) Composition of the sulfoglycolipid. (2) Sulfoglycolipid metabolism based on incorporation of [35S]sulfate or some properties of sulfoglycolipid metabolism, including the activities of anabolic and catabolic enzymes. The results indicated following characteristics of the isolated renal tubules in comparison to the kidney in vivo. (1) The sulfoglycolipid compositions are qualitatively similar, except that the content of glucosyl sulfatide, Gg3Cer II3-sulfate, and GM4 was slightly higher in the isolated tubules. (2) The apparent half-lives (15-55 min) of sulfoglycolipids in the isolated tubules could indicate the existence of a rapid turnover pool of these lipids. (3) The sulfotransferase and sulfatase activities related to sulfoamphiphiles in the renal tubule were similar to those reported for the whole kidney. Based on the above criteria, we conclude that the isolated rat renal tubule should be a useful metabolic system for clarification of the short-term physiological events, up to 90 min, of proximal tubular sulfoglycolipids. By using the present system, we showed that biosynthesis of the renal total sulfoglycolipid was significantly elevated in rats deprived of water for 24 h.
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Affiliation(s)
- Ken-ichi Nagai
- Department of Biochemistry, Teikyo University School of Medicine, 2-11-1, Kaga, Itabashi, Tokyo 173-8605, Japan
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Abstract
Metachromatic leukodystrophy (MLD) is a lysosomal storage disorder caused by the deficiency of arylsulphatase A (ASA). This leads to the accumulation of the sphingolipid 3-O-sulphogalactosylceramide (sulphatide) and progressive demyelination in the nervous system of MLD patients. The mechanisms and development of pathology in the disease are still largely unknown. In this study we investigate how the inability to degrade sulphatide affects the formation of myelin in ASA-deficient (ASA-/-) mice. In mice at 2 weeks of age there was a substantial reduction in myelin basic protein (MBP) mRNA and protein. This was confirmed by an immunohistochemical analysis. MBP mRNA and protein, however, reach normal levels at 3 weeks of age. Proteolipid protein (PLP) and MAL mRNA were also reduced in ASA-/- mice at 2 weeks of age; whereas the level of PLP mRNA was normal at 26 weeks of age, MAL mRNA expression remained reduced up to this age. In situ hybridization revealed no significant changes in the number of myelinating oligodendrocytes or oligodendrocyte precursor cells in ASA-/- mice. These results suggest that oligodendrocyte differentiation was normal in ASA-/- mice. No differences were found in the expression of the sulphatide synthesizing enzymes cerebroside sulphotransferase and UDP-galactose : ceramide galactosyltransferase. Our data demonstrate a delay in myelin formation in ASA-/- mice. This raises the possibility that similar alterations in MLD patients may contribute to the pathology of the disease.
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Affiliation(s)
- Afshin Yaghootfam
- Institut für Physiologische Chemie, Rheinische-Friedrich-Wilhelms Universität Bonn, Nussallee 11, 53115 Bonn, Germany.
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Hirahara Y, Bansal R, Honke K, Ikenaka K, Wada Y. Sulfatide is a negative regulator of oligodendrocyte differentiation: Development in sulfatide-null mice. Glia 2004; 45:269-77. [PMID: 14730700 DOI: 10.1002/glia.10327] [Citation(s) in RCA: 80] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022]
Abstract
Galactosylceramide (GalC) and its sulfated analogue, sulfatide, are major galactosphingolipid components of myelin and oligodendrocyte plasma membranes in the nervous system. We previously hypothesized that these galactolipids play functional roles in the regulation of oligodendrocyte terminal differentiation by acting as sensors/transmitters of environmental information. Evidence strongly supports this idea. First, these molecules are initially expressed on the cell surface at the interface at which oligodendrocyte progenitors first enter terminal differentiation. Second, exposure of oligodendrocyte progenitors to anti-GalC/-sulfatide (RmAb) or antisulfatide (O4), but not anti-GalC (O1), antibodies leads to the reversible arrest of oligodendrocyte lineage progression at this interface. Third, in cerebroside galactosyl transferase-null mice (Cgt(-/-)) that are unable to synthesize either GalC or sulfatide, terminal differentiation and morphological maturation of oligodendrocytes are enhanced. In the present study, we examined oligodendrocytes differentiation in cerebroside sulfotransferase-null mice (Cst(-/-)) that lack sulfatide but express GalC. We show that cerebroside sulfotransferase mRNA expression begins already in the embryonic spinal cord and progressively increases with age, that the late progenitor marker POA is not synthesized in the absence of this enzyme, and that, most notably, there is a two- to threefold enhancement in the number of terminally differentiated oligodendrocytes both in culture and in vivo, similar to that in mice lacking both GalC and sulfatide. We conclude that primarily sulfatide, rather than GalC, is a key molecule for the negative regulation of oligodendrocyte terminal differentiation.
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Affiliation(s)
- Yukie Hirahara
- Research Institute, Osaka Medical Center for Maternal and Child Health, Osaka, Japan
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41
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Blomqvist M, Kaas A, Månsson JE, Formby B, Rynmark BM, Buschard K, Fredman P. Developmental expression of the type I diabetes related antigen sulfatide and sulfated lactosylceramide in mammalian pancreas. J Cell Biochem 2003; 89:301-10. [PMID: 12704793 DOI: 10.1002/jcb.10513] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
Abstract
Previous studies have shown that sulfatide is present and functionally involved in beta cells, and that anti-sulfatide antibodies (ASA) exist during development of type I diabetes mellitus. To further explore the possible role of sulfatide in type I diabetes, developmental expression was examined in human pancreas and in pancreas of the type I diabetes models BB rat and NOD mouse compared to Lewis rat and BALB/c mouse, respectively. Sulfatide was not only expressed in adult pancreas, but also in human fetal and rodent neonatal pancreas, i.e., during the growing period of the immunological self. Sulfatide had a different expression pattern in human beings and rodents, concerning both the amounts of sulfatide and expression during development. There was no change in the sulfatide fatty acid isoform expression during development. The pancreatic expression of another sulfated glycosphingolipid, sulfated lactosylceramide, indicated that this molecule is a potential fetal/neonatal marker, which was further expressed in the type I diabetic models. In conclusion, these findings give further support to the possibility that sulfatide is a relevant autoantigen in type I diabetes and that sulfated lactosylceramide might function as a potential risk factor for disease development, at least in the animal models.
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Affiliation(s)
- Maria Blomqvist
- Institute of Clinical Neuroscience, Experimental Neuroscience Section, The Sahlgrenska Academy at Göteborg University, Sahlgrenska University Hospital/Mölndal, SE-431 80 Mölndal, Sweden.
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Okino N, Mori K, Ito M. Genomic structure and promoter analysis of the mouse neutral ceramidase gene. Biochem Biophys Res Commun 2002; 299:160-6. [PMID: 12435403 DOI: 10.1016/s0006-291x(02)02540-8] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
Abstract
We report here the molecular cloning of the mouse neutral ceramidase gene and its promoter analysis. The gene, composed of 27 exons ranging in size from 40 to 292 bp, spans more than 70 kb. Analysis of the 5(')-flanking region of the ceramidase genes revealed that the first exon of the gene of mouse liver was exactly the same as that of mouse kidney and Swiss 3T3 fibroblasts but completely different from that of mouse brain. The putative promoter regions of liver and brain ceramidase genes contained several well-characterized promoter elements such as GATA-2, C/EBP, and HNF3beta but lacked TATA and CAAT boxes, a typical feature of a housekeeping gene, although the expression is regulated in a tissue-specific manner. Interestingly, a GC box was exclusively found in the putative promoter of mouse liver whereas potential AP1 and AP4 binding sites were present in that of mouse brain. By a luciferase reporter gene assay, it was shown that the GC-rich region, which exists just upstream of the first exon, conferred the promoter activity in Swiss 3T3 cells.
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Affiliation(s)
- Nozomu Okino
- Department of Bioscience and Biotechnology, Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, 812-8581, Fukuoka, Japan
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Eckhardt M, Fewou SN, Ackermann I, Gieselmann V. N-glycosylation is required for full enzymic activity of the murine galactosylceramide sulphotransferase. Biochem J 2002; 368:317-24. [PMID: 12175333 PMCID: PMC1222978 DOI: 10.1042/bj20020946] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/19/2002] [Revised: 08/07/2002] [Accepted: 08/13/2002] [Indexed: 11/17/2022]
Abstract
3- O -Sulphogalactosylceramide (sulphatide) is a major lipid component of myelin membranes, and is required for proper myelin formation. Sulphatide is synthesized in the Golgi apparatus by galactosylceramide sulphotransferase (CST; EC 2.8.2.11). Murine and human CSTs contain two putative N-glycosylation sites (Asn-66 and Asn-312). The second site is conserved among all galactose 3-O-sulphotransferases cloned to date. In order to study the functional relevance of N-glycosylation, we generated epitope-tagged CST and soluble Protein A-CST fusion proteins lacking both N-glycosylation sites, separately or in combination. Our results show that both sites are glycosylated when CST is expressed in Chinese hamster ovary (CHO) or COS cells. Moreover, transfecting CST mutants lacking both N-glycosylation sites, or only Asn-312, reduced significantly the amount of sulphatide synthesized, whereas substituting Asn-66 with a glutamine residue did not. In contrast, activity in vitro was reduced by approx. 50% in the Asn-66-->Gln (N66Q) mutant, and was almost undetectable in N312Q and N66/312Q transfectants. Furthermore, soluble Protein A-CST expressed in the presence of tunicamycin was almost inactive, and accumulated in transfected cells. Expression of fully active CST in a CHO-glycosylation mutant lacking N-acetylglucosaminyltransferase I demonstrated that condensation of the N-linked pentamannosyl-core structure is sufficient to form a fully active enzyme.
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Affiliation(s)
- Matthias Eckhardt
- Institut für Physiologische Chemie, Rheinische-Friedrich-Wilhelms Universität Bonn, Nussallee 11, D-53115 Bonn, Germany.
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Abstract
Structural diversity of the sugar chains attached to proteins and lipids that arises from the variety of combinations of different monosaccharides, different types of linkages, branch formation and secondary modifications, such as sulfation, possesses a large amount of biological information. A number of proteoglycans, glycoproteins, and glycolipids contain sulfated carbohydrates. Their sulfate groups provide a negative charge and play a role in a specific molecular recognition process. The sulfation of oligosaccharides is catalyzed by the Golgi-associated sulfotransferases. Recent success in molecular cloning of these sulfotransferases has brought a breakthrough in the understanding of biological function of sulfated oligosaccharides in a variety of contexts. Investigations on the relationship of sulfated oligosaccharides to human diseases including hereditary deficiency, cancer, inflammation, and infection may provide hints for curing disastrous diseases.
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Affiliation(s)
- Koichi Honke
- Department of Biochemistry, Osaka University Medical School, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
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Seko A, Nagata K, Yonezawa S, Yamashita K. Down-regulation of Gal 3-O-sulfotransferase-2 (Gal3ST-2) expression in human colonic non-mucinous adenocarcinoma. Jpn J Cancer Res 2002; 93:507-15. [PMID: 12036446 PMCID: PMC5927024 DOI: 10.1111/j.1349-7006.2002.tb01285.x] [Citation(s) in RCA: 31] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022] Open
Abstract
Expression levels of sulfomucin in human colonic adenocarcinomas are lower than those in normal colonic mucosa; this should be in part caused by down-regulation of expression of sulfotransferases, but it remains unclear which Gal 3-O-sulfotransferase (Gal3ST) is responsible for the biosynthesis of sulfomucin. In this study, we first examined the substrate specificities of four Gal3STs cloned so far, and found that Galbeta1 3GlcNAcbeta1 3Galbeta1 4Glc (LNT) can be utilized only by Gal3ST-2 as an acceptor substrate. The substrate specificity of Gal3ST-2 is closely similar to those of Gal3ST activities present in human normal mucosa and adenocarcinomas, suggesting that Gal3ST-2 is the dominant Gal3ST in colon and colonic cancer. Secondly, using LNT as a substrate, we comparatively analyzed levels of Gal3ST-2 activities in non-mucinous adenocarcinoma, mucinous adenocarcinomas, and the adjacent normal mucosa. We found that levels of Gal3ST-2 activities in non-mucinous adenocarcinoma are significantly lower than those in the adjacent normal mucosa, while those in mucinous adenocarcinomas are not significantly different from those in the adjacent normal mucosa. Moreover, we showed by a competitive RT-PCR method that expression levels of transcript for Gal3ST-2 in non-mucinous adenocarcinoma are lower than those in normal mucosa. These results suggest that Gal3ST-2 is one of the enzymes responsible for biosynthesis of sulfomucin, and that expression levels of Gal3ST-2 are down-regulated in non-mucinous adenocarcinoma.
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Affiliation(s)
- Akira Seko
- Department of Biochemistry, Sasaki Institute, Chiyoda-ku, Tokyo 101-0062
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Honke K, Hirahara Y, Dupree J, Suzuki K, Popko B, Fukushima K, Fukushima J, Nagasawa T, Yoshida N, Wada Y, Taniguchi N. Paranodal junction formation and spermatogenesis require sulfoglycolipids. Proc Natl Acad Sci U S A 2002; 99:4227-32. [PMID: 11917099 PMCID: PMC123630 DOI: 10.1073/pnas.032068299] [Citation(s) in RCA: 247] [Impact Index Per Article: 10.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Mammalian sulfoglycolipids comprise two major members, sulfatide (HSO3-3-galactosylceramide) and seminolipid (HSO3-3-monogalactosylalkylacylglycerol). Sulfatide is a major lipid component of the myelin sheath and serves as the epitope for the well known oligodendrocyte-marker antibody O4. Seminolipid is synthesized in spermatocytes and maintained in the subsequent germ cell stages. Both sulfoglycolipids can be synthesized in vitro by using the isolated cerebroside sulfotransferase. To investigate the physiological role of sulfoglycolipids and to determine whether sulfatide and seminolipid are biosynthesized in vivo by a single sulfotransferase, Cst-null mice were generated by gene targeting. Cst(-/-) mice lacked sulfatide in brain and seminolipid in testis, proving that a single gene copy is responsible for their biosynthesis. Cst(-/-) mice were born healthy, but began to display hindlimb weakness by 6 weeks of age and subsequently showed a pronounced tremor and progressive ataxia. Although compact myelin was preserved, Cst(-/-) mice displayed abnormalities in paranodal junctions. On the other hand, Cst(-/-) males were sterile because of a block in spermatogenesis before the first meiotic division, whereas females were able to breed. These data show a critical role for sulfoglycolipids in myelin function and spermatogenesis.
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Affiliation(s)
- Koichi Honke
- Department of Biochemistry, Osaka University Medical School, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan
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Kabayama K, Ito N, Honke K, Igarashi Y, Inokuchi J. Suppression of integrin expression and tumorigenicity by sulfation of lactosylceramide in 3LL Lewis lung carcinoma cells. J Biol Chem 2001; 276:26777-83. [PMID: 11352905 DOI: 10.1074/jbc.m100428200] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
To investigate the cellular functions of sulfated glycosphingolipids, we introduced the cerebroside sulfotransferase (CST) gene into J5 cells, a subclone of 3LL Lewis lung carcinoma cells. The J5 cells lack acidic glycosphingolipids but accumulate their common biosynthetic precursor, lactosylceramide. We established the stable CST transfectants, J5/CST-1 and J5/CST-2 clones, highly expressing sulfated lactosylceramide (SM3). Both clones exhibited more spherical morphology in comparison to mock transfectant, and their adhesiveness to fibronectin and laminin was significantly lower. The loss of cell-substratum interactions in these SM3-expressing cells could be attributed to decreased expression of integrins (alpha(5), alpha(6), and beta(1)) on the cell surface and their whole cellular levels. However, the levels of H-2K(b) and H-2D(b) antigens remained unchanged. Reverse transcriptase-polymerase chain reaction and Northern blot analyses for these integrins exhibited significant decrease of beta(1) gene expression in J5/CST-1 and 2, but there was no change in the levels of alpha(5) and alpha(6) transcripts. Deglycosylation by endoglycosidase H treatment clearly demonstrated that the precursor form of beta(1) integrin, possessing high mannose oligosaccharide chains, was preferentially decreased in the CST transfectants. These results demonstrate that endogenous SM3 negatively regulates beta(1) integrin expression at the transcriptional level, and the decrease of alpha integrin proteins in the CST transfectants was due to the post-transcriptional modification. We suggest the putative importance of the intracellular pre-beta(1) integrin pool for normal integrin maturation and subsequent function. Although the rates of cell proliferation in vitro for mock and CST transfectants were similar, tumorigenicity of J5/CST-1 and -2 cells inoculated into syngeneic C57/BL6 mice was greatly decreased or even absent. This was probably due to global loss of the efficient cell-matrix interactions, which are essential for the development of malignant tumors in vivo. Thus, we showed the evidence that cellular SM3 negatively regulates the cell-substratum interaction, resulting in the loss of tumorigenicity.
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Affiliation(s)
- K Kabayama
- Department of Biomembrane and Biofunctional Chemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita 12-jo, Nishi 6-chome, Kita-ku, Sapporo 060-0812, Japan
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Sekine M, Taya C, Kikkawa Y, Yonekawa H, Takenaka M, Matsuoka Y, Imai E, Izawa M, Kannagi R, Suzuki A. Regulation of mouse kidney tubular epithelial cell-specific expression of core 2 GlcNAc transferase. EUROPEAN JOURNAL OF BIOCHEMISTRY 2001; 268:1129-35. [PMID: 11179979 DOI: 10.1046/j.1432-1327.2001.01980.x] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/25/2023]
Abstract
A mouse gene, Gsl5, controls the expression of Galbeta1-4(Fucalpha1-3)GlcNAcbeta1-6(Galbeta1-3)Gb4Cer and its precursor glycolipids in the kidney by regulating transcription of beta-1,6-GlcNAc transferase. Here we report that Gsl5 controls the expression of the core 2 structure [GlcNAcbeta1-6(Galbeta1-3)GalNAcalpha1-Ser/Thr] of glycoproteins as well as the glycolipid, GlcNAcbeta1-6(Galbeta1-3)GalNAcbeta1-3Galalpha1-4Galbeta1-4Glcbeta1-ceramide. Immunohistochemical studies using an anti-(core 2-Lex) monoclonal antibody demonstrated that lysosome-like vesicles of proximal tubule cells were clearly stained in a Gsl5 wild type mouse, but not in a Gsl5 mutant strain of mice. Western blotting of microsomal fractions of kidney tissue with the same antibody confirmed the histological findings. In situ hybridization with an antisense probe to the kidney-specific mRNA demonstrated that the mRNA is localized at proximal tubule-cells in the cortex adjacent to the medulla, but not detected in glomeruli nor in collecting duct cells in the medulla. The results obtained by immunohistological staining and in situ hybridyzation are compatible and lead to the conclusion that the kidney specific mRNA is expressed in a proximal tubular cell specific manner and produces core 2 GlcNAc transferase responsible for the production of glycoproteins localized at vesicles in the proximal tubular cells. Glycosylation regulated by Gsl5 gene may modify functions of membrane glycoproteins in proximal tubular cells.
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Affiliation(s)
- M Sekine
- Departments of Membrane Biochemistry and Laboratory Animal Science, The Tokyo Metropolitan Institute of Medical Science, Japan
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Kozak M. Do the 5'untranslated domains of human cDNAs challenge the rules for initiation of translation (or is it vice versa)? Genomics 2000; 70:396-406. [PMID: 11161792 DOI: 10.1006/geno.2000.6412] [Citation(s) in RCA: 51] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
The validity of the scanning mechanism for initiation of translation has been questioned based on a compilation of human cDNA sequences that showed a high frequency of upstream ATG codons. However, closer scrutiny of those cDNAs upholds the opposite view: the 5'UTRs on most cDNAs are compatible with standard rules for initiation of translation, and those rules can be used to flag anomalous cDNAs that, upon checking, turn out to have been misinterpreted. Some of the problematic 5'UTR sequences that persist, after obvious errors in the cDNA library have been corrected, might derive from transcripts that are not intended to be translated. Examples are given of genes that, for regulatory reasons, produce transcripts that are truncated, or retain an intron, or are otherwise configured in a way that precludes translation. The existence of a cDNA proves that a gene is transcribed, but only that; not every cDNA derives from a functional mRNA. Along with providing practical guidelines for interpreting cDNA sequences, the scanning model provides a theoretical framework for understanding the effects of certain mutations in the 5'UTR that alter the translatability of mRNAs, thereby contributing to cancer and other human diseases.
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Affiliation(s)
- M Kozak
- Department of Biochemistry, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, 675 Hoes Lane, Piscataway, New Jersey 08854, USA.
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Tadano-Aritomi K, Hikita T, Fujimoto H, Suzuki K, Motegi K, Ishizuka I. Kidney lipids in galactosylceramide synthase-deficient mice: absence of galactosylsulfatide and compensatory increase in more polar sulfoglycolipids. J Lipid Res 2000. [DOI: 10.1016/s0022-2275(20)33431-3] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022] Open
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