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Genes required for glycolipid synthesis and lipoteichoic acid anchoring in Staphylococcus aureus.

Staphylococcus aureus lipoteichoic acid (LTA) is composed of a linear 1,3-linked polyglycerolphosphate chain and is tethered to the bacterial membrane by a glycolipid (diglucosyl-diacylglycerol [Glc2-DAG]). Glc2-DAG is synthesized in the bacterial cytoplasm by YpfP, a processive enzyme that transfers glucose to diacylglycerol (DAG), using UDP-glucose as its substrate. Here we present evidence that the S. aureus alpha-phosphoglucomutase (PgcA) and UTP:alpha-glucose 1-phosphate uridyltransferase (GtaB) homologs are required for the synthesis of Glc2-DAG. LtaA (lipoteichoic acid protein A), a predicted membrane permease whose structural gene is located in an operon with ypfP, is not involved in Glc2-DAG synthesis but is required for synthesis of glycolipid-anchored LTA. Our data suggest a model in which LtaA facilitates the transport of Glc2-DAG from the inner (cytoplasmic) leaflet to the outer leaflet of the plasma membrane, delivering Glc2-DAG as a substrate for LTA synthesis, thereby generating glycolipid-anchored LTA. Glycolipid anchoring of LTA appears to play an important role during infection, as S. aureus variants lacking ltaA display defects in the pathogenesis of animal infections.

Animals↗

Characterization of novel long-chain 1,2-diols in Thermus species and demonstration that Thermus strains contain both glycerol-linked and diol-linked glycolipids.

In this study, we purified and characterized tetra- and triglycosyl glycolipids (GL-1 and GL-2, respectively) from two different colonial forms of Thermus scotoductus X-1, from T. filiformis Tok4 A2, and from T. oshimai SPS-11. Acid hydrolysis of the purified glycolipids liberated, in addition to the expected long-chain fatty acids, two components which were identified by gas chromatography-mass spectrometry as 16-methylheptadecane-1,2-diol and 15-methylheptadecane-1,2-diol. Fast atom bombardment mass spectrometry of the intact glycolipids indicated that a major proportion consisted of components with glycan head groups linked to long-chain 1,2-diols rather than to glycerol, although in all cases glycerol-linked compounds containing similar glycan head groups were also present. As in other Thermus strains, the polar head group of GL-1 from T. filiformis Tok4 A2 and from T. scotoductus X-1 colony type t2 was a glucosylgalactosyl-(N-acyl)glucosaminylglucosyl moiety. However, GL-2 from T. scotoductus X-1 colony type t1 and from T. oshimai SPS-11 was a truncated analog which lacked the nonreducing terminal glucose. Long-chain 1,2-diols have been previously reported in the polar lipids of Thermomicrobium roseum and (possibly) Chloroflexus aurantiacus, but to our knowledge, this is the first report of their detection in other bacteria and the first account of the structural determination of long-chain diol-linked glycolipids.

Fatty Acids↗

Isolation and characterization of glycolipids from some photosynthetic bacteria.

Glycosyl glycerides have been found in substantial amounts in Chloropseudomonas ethylicum but could not be detected in two strains of Rhodopseudomonas palustris. Rhodospirillum molischianum possibly contains small amounts of monoglycosyl diglyceride. The glycolipids of C. ethylicum have been separated into two components. One of these, glycolipid I, is a monogalactosyl diglyceride. Glycolipid II, upon acid hydrolysis, yields galactose, rhamnose, and a third, unidentified sugar. The glycolipids or total lipids of the photosynthetic bacteria examined contained saturated and monounsaturated, but none of the more highly unsaturated, fatty acids.

Bacteria↗

Enzyme-linked immunosorbent assay of glycolipid antigens for identification of mycobacteria.

Enzyme-linked immunosorbent assays which are based on species- or type-specific glycolipids antigens and in which rabbit antisera are prepared with homologous strains are capable of distinguishing among serological variants of the Mycobacterium avium-Mycobacterium intracellulare-Mycobacterium scrofulaceum complex, Mycobacterium chelonei subspecies chelonei and abscessus, Mycobacterium simiae I and II, Mycobacterium kansasii, Mycobacterium szulgai, Mycobacterium xenopi, and Mycobacterium fortuitum biovariant peregrinum. The immunoreactive glycolipids can be divided into two classes. Those resistant to alkali, the C-mycoside glycopeptidolipids, are present in the M. avium-M. intracellulare-M. scrofulaceum, the M. chelonei subspecies chelonei and abscessus, and the M. simiae I and II complexes and in M. fortuitum biovariant peregrinum. The alkali-labile glycolipid antigens, the lipooligosaccharides, are present in M. kansasii, M. szulgai, and M. xenopi. In one study, the combination of enzyme-linked immunosorbent assay and alkaline susceptibility was compared with seroagglutination in the identification of 60 clinical isolates of nontuberculous mycobacteria: 45 showed perfect concordance, 9 could be narrowed to one, two, or three possibilities, and the rest did not correspond. In a second study involving 43 clinical isolates that were untypable by seroagglutination or were autoagglutinable, the results of enzyme-linked immunosorbent assay and thin-layer chromatography of glycolipid antigens were compared: 21 showed clear concordance. The results demonstrate that enzyme-linked immunosorbent assay is particularly useful in assessing the antigenicity of lipids, and sensitivity, ease, and rapidity recommend it as an adjunct to seroagglutination and thin-layer chromatography for the identification of nontuberculous mycobacteria.

Animals↗

Verotoxin receptor glycolipid in human renal tissue.

Infection with verotoxin producing Escherichia coli has been strongly implicated in the etiology of the hemolytic uremic syndrome (HUS). We have previously shown that this toxin specifically binds to a glycolipid receptor-globotriosyl ceramide (Gb3). We have therefore quantitated the level of this glycolipid by HPLC in human renal cortex and medulla as a function of age. We have also measured the binding of verotoxin to Gb3 isolated from each renal tissue sample. Gb3 was a major component of the glycolipid fraction of all renal samples analyzed. The levels were found to be higher in the cortex than medulla, correlating with the clinical incidence of renal lesions in HUS, but reduced in the kidneys of infants as compared to adults. Verotoxin binding was directly proportional to the renal Gb3 content. Thus, human renal tissue is a rich source of the verotoxin receptor glycolipid. However, changes in receptor concentration cannot explain the age-related incidence of HUS.

Aging↗

Glycolipid expression in prostatic tissue and analysis of the antigen recognized by antiprostatic monoclonal antibody APG1.

The expression patterns of glycolipid from prostatic hyperplasia, prostatic cancer and normal prostate tissue were observed. A further analysis of antigen recognized by mouse monoclonal antibody APG1, which was gained by immunizing glycolipids extracted from human prostate cancer, was also performed. In cancer tissue, both of the lactosyl and globoside series glycolipids were found to be generally reduced, although in the ganglioside series, GM3 and GD3 were not reduced and only the glycolipids with longer chains than GD2 were found to be reduced. These results indicated that the inhibition of sugar chain elongation, but not sialylation, was the main synthetic change occurring with carcinogenesis of the human prostate. APG1 reacted with only two bands near GM2 and GD2 of the ganglioside fraction on a thin-layer chromatography plate, but it did not react with any of the known gangliosides of the ganglioside series including GM2 and GD2. Histochemically, APG1 showed intense reaction only in frozen tissue sections of human prostate, and the reactivity decreased with the increasing grade of cancer. Therefore, this antigen was considered to be a prostate-specific and differentiated antigen reacting with nonganglioseries gangliosides.

Antibodies, Monoclonal↗

Mannosylerythritol lipid, a yeast extracellular glycolipid, shows high binding affinity towards human immunoglobulin G.

BACKGROUND: There have been many attempts to develop new materials with stability and high affinity towards immunoglobulins. Some of glycolipids such as gangliosides exhibit a high affinity toward immunoglobulins. However, it is considerably difficult to develop these glycolipids into the practical separation ligand due to their limited amounts. We thus focused our attention on the feasible use of "mannosylerythritol lipid A", a yeast glycolipid biosurfactant, as an alternative ligand for immunoglobulins, and undertook the investigation on the binding between mannosylerythritol lipid A (MEL-A) and human immunoglobulin G (HIgG). RESULTS: In ELISA assay, MEL-A showed nearly the same binding affinity towards HIgG as that of bovine ganglioside GM1. Fab of human IgG was considered to play a more important role than Fc in the binding of HIgG by MEL-A. The bound amount of HIgG increased depending on the attached amount of MEL-A onto poly (2-hydroxyethyl methacrylate) (polyHEMA) beads, whereas the amount of human serum albumin slightly decreased. Binding-amount and -selectivity of HIgG towards MEL-A were influenced by salt species, salt concentration and pH in the buffer solution. The composite of MEL-A and polyHEMA, exhibited a significant binding constant of 1.43 x 10(6) (M(-1)) for HIgG, which is approximately 4-fold greater than that of protein A reported. CONCLUSIONS: MEL-A shows high binding-affinity towards HIgG, and this is considered to be due to "multivalent effect" based on the binding molar ratio. This is the first report on the binding of a natural human antibody towards a yeast glycolipid.

Animals↗

Inflammatory cytokines inhibit upregulation of glycolipid expression by Schwann cells in vitro.

OBJECTIVE: To determine whether products of inflammatory cells can inhibit differentiation and synthesis of myelin glycolipids by Schwann cells. BACKGROUND: Infiltration of the peripheral nervous system by inflammatory cells is a feature of acquired demyelinating neuropathies. It is not clear what role these cells have in causing demyelination or inhibiting myelin synthesis. METHODS: Nonmyelinating rat Schwann cells were incubated with 1) different concentrations of activated supernatants (AS) from mitogen-activated inflammatory cells; 2) 8-bromo cyclic adenosine monophosphate (8Br cAMP), known to induce Schwann cell differentiation and synthesis of glycolipids; 3) 8Br cAMP and varying concentrations of AS; 4) 8Br cAMP and cytosine arabinoside (Ara C), which inhibits Schwann cell proliferation; 5) 8Br cAMP, AS, and Ara C; or 6) additional medium. RESULTS: AS inhibits the capacity of cAMP to induce Schwann cell expression of myelin-associated glycolipids. Inhibition of glycolipid expression was independent of the capacity of these AS to induce Schwann cell proliferation. CONCLUSIONS: These data suggest that inflammatory mediators are capable of inhibition of Schwann cell differentiation and synthesis of myelin.

8-Bromo Cyclic Adenosine Monophosphate↗

Putative inhibitory effects of chrysotile, crocidolite, and amosite mineral fibers on the more complex surface membrane glycolipids and glycoproteins.

Syrian hamster embryo cells were treated with galactose oxidase, followed by reduction with tritiated sodium borohydride at pH 7.4. The labeling patterns of galactosyl and N-acetyl galactosaminyl residues on the cell surface were altered in comparing scraped vs. unscraped and buffer vs. media-soaked cells treated with galactose oxidase. From these preliminary studies, the procedure to be used in most of the asbestos treatment studies was to treat cells in situ, in buffer with galactose oxidase, and then to label treated scraped cells with NaB(3)H(4). After 20 hr interaction between chrysotile asbestos and Syrian hamster cell cultures, an alteration in surface labeling of glycolipids and glycoproteins was observed. Tritiated disialogangliosides (G(Dla)) and the higher molecular weight labeled glycoproteins were significantly reduced by asbestos treatment. Similar chrysotile asbestos-treated cultures were grown in monolayers in MEM (Eagles) with 10% fetal bovine serum for 2, 24, 48, and 72 hr and then surface-labeled with galactose oxidase-. NaB(3)H(4) in phosphate buffer. Little or no difference was observed between surface-labeled lipid or protein distribution in untreated cells and those treated with asbestos for 2 hr. Asbestos-induced polar and neutral glycolipid pattern changes were observed at 24, 48, and 72 hr. Disialo- and trisialogangliosides (the more complex gangliosides) were decreased 85%, whereas globoside GL-4 was decreased by 60% at 72 hr. An overall decrease of labeled glycoproteins was observed at 24-48 hr. By 72 hr there was a complete loss of labeled protein bands with 80,000 dalton molecular mass. Since the changes in glycoproteins and glycolipids occur only after extended exposure of the cells to asbestos, the present studies support the concept that a metabolic rather than immediate masking effect is involved. Comparisons of treatment of Syrian hamster embryo cells with various asbestos fibers for 48 hr in the order of decreasing reduction in complex gangliosides were crocidolite>chrysotile (intermediate)>amosite. Effects of the above fibers on high molecular weight glycoproteins labeling followed the same order. The labeling pattern is reminiscent of the increased simplification of glycolipids and glycoproteins found in transformed cells. In the case of asbestos which appears to have no independent mutagenic capability, it is more likely that the membrane changes induced by asbestos serve to allow other mutagens to pass into the cell so as to act on the nuclear structure.

Animals↗

Glycolipids as immune modulatory tools.

NKT cells are a subset of regulatory lymphocytes characterized by co-expression of the NK cell receptor-CD161 and an invariant TCR-alpha chain (Valpha14-Jalpha28). They are most abundant in the liver, spleen, and bone marrow. NKT lymphocytes have been implicated in the regulation of autoimmune processes in both mice and humans. Activation of NKT lymphocytes leads to rapid amplification of either IFNgamma or IL4, endowing these cells with the capability to mediate both pro-inflammatory and anti-inflammatory immune responses. Activation of this subset of cells is associated with significant liver damage in the Concanavalin A immune mediated hepatitis model. Administration of CD1d ligand has a protective role in collagen-induced arthritis in mice. Disease amelioration was associated with a shift in the immune balance from a pathological Th1 type response towards a protective Th2 type response. In humans, patients with SLE, scleroderma, diabetes, multiple sclerosis, and rheumatoid arthritis have lower numbers of peripheral NKT cells. NKT lymphocytes promote tumor rejection in experimental models of tumor immunotherapy. In contrast, NKT lymphocyte-related anti-tumor activity is associated with pro-inflammatory Th1-type immune responses. NKT cells were shown to have a role in suppression of hepatocellular carcinoma (HCC) via immune regulation towards tumor derived antigens, and adoptive transfer of dendritic cells pulsed ex vivo with the same antigens. NKT lymphocytes are activated by interaction of their TCR with glycolipids presented by CD1d, a nonpolymorphic, MHC class I-like molecule expressed by antigen presenting cells, and also by hepatocytes. Several possible ligands for NKT cells have recently been suggested including CD1d bound Glucocerebroside. Glucocerebroside (GC, beta-glucosylceramide), a naturally occurring glycolipid, is a metabolic intermediate in the anabolic and catabolic pathways of complex glycosphingolipids. Its synthesis from ceramide is catalyzed by the enzyme glucosylceramide synthase. Inherited deficiency of glucocerebrosidase, a lysosomal hydrolase, results in Gaucher's disease. Patients with Gaucher's disease have altered humoral and cellular immune profiles and increased peripheral blood NKT lymphocytes. CD1d-bound glucocerebroside does not activate NKT cells directly, and may inhibit activation of NKT cells by alpha-GalCer. On the other hand, glucosylceramide-synthase deficiency was shown to lead to defective ligand presentation by CD1d, with secondary inhibition of NKT cell activation. Recent studies have suggested that a number of glycolipids, including GC, have an immune modulatory effect in several immune mediated disorders. The ability to alter NKT lymphocyte function in various settings and the potential application of natural glycolipids for treatment are discussed.

Animals↗

Glycolipids as the major autoantigens of cytoplasmatic islet cell antibodies.

It has been recently suggested that pancreatic glycolipidic extracts and acidic glycolipid fractions are able to block the binding of ICA to frozen sections of human pancreas. We study the prevalence of blocking effect by the upper-phase from human pancreatic glycolipid extracts (PGE) in thirty-eight sera ICA positive from seventeen IDDM patients and twenty-one first relatives of type 1 diabetics. Total inhibition was found in 82% and 76% insulin dependent diabetes mellitus patients and first relatives of type 1 diabetics respectively. Partial and no inhibition of ICA+ sera was seen in 6%, 12% of type 1 diabetics and 19%, 5% of the first degree relatives of type 1 diabetics respectively. Our study suggests that there is heterogeneity of cytoplasmatic islet cell antibodies and that glycolipids are the major autoantigen of ICA.

Adolescent↗

Antibodies against tumor cell glycolipids and proteins, but not mucins, mediate complement-dependent cytotoxicity.

One of several effector mechanisms thought to contribute to Ab efficacy against cancer is complement-dependent cytotoxicity (CDC). Serological analysis of a series of clinical trials conducted over a 10-year period suggested that six vaccines containing different glycolipids induced Abs mediating CDC whereas four vaccines containing carbohydrate or peptide epitopes carried almost exclusively by mucin molecules induced Abs that did not mediate CDC. To explore this further, we have now compared cell surface reactivity using flow cytometry assays (FACS), complement-fixing ability, and CDC activity of a panel of mAbs and immune sera from these trials on the same two tumor cell lines. Abs against glycolipids GM2, globo H and Lewis Y, protein KSA (epithelial cell adhesion molecule, also known as EpCAM) and mucin Ags Tn, sialylated Tn, Thomsen Friedenreich (TF), and MUC1 all reacted comparably by FACS with tumor cells expressing these Ags. Compared with the strong complement binding and CDC with Abs against glycolipids and KSA, complement binding was diminished with Abs against mucin Ags and no CDC was detected. A major difference between these two groups of Ags is proximity to the cell membrane. Glycolipids and globular glycoproteins extend less than 100 A from the cell membrane while mucins extend up to 5000 A. Although complement activation at sites remote from the cell membrane has long been known as a mechanism for resistance from complement lysis in bacteria, it is identified here for the first time as a factor which may contribute to resistance from CDC against cancer cells.

Antibodies, Monoclonal↗

[Glycolipids of rabbit spinal cord in experimental allergic encephalomyelitis].

Within the acute period of experimental allergic encephalomyelitis, when the clinical picture was quite distinct, content of glycolipids was studied biochemically and histochemically in lumbar region of rabbit spinal cord. In this region the myelin degradation was the most distinct; content of glycolipids was decreased. Localization of the centers of decreased glycolipids content, determined histochemically, was similar to the localization of the zones of degraded myelin. The decrease in content of glycolipids was due to breakdown of cerebrosides but not sulphatides. Quantitative and qualitative composition of gangliosides was not altered in the spinal cord.

Animals↗

Senile cataract-related accumulation of Lewis(x) glycolipid in human lens.

A glycosphingolipid that reacted positively to anti-stage-specific embryonic antigen-1 (SSEA-1) antiserum accumulated in human lens in association with aging and senile cataract formation. Since this antiserum recognizes Lewis(x) (Le(x)) structure, Gal beta 1-4(Fuc alpha 1-3)GlcNAc-, which is a typical tumor-associated and differentiation-related saccharide chain, the lens glycolipid was predicted to be a Lex antigen. The glycolipid purified from cataractous lens tissues was indeed a Lex glycolipid, Gal beta 1-4(Fuc alpha 1-3)GlcNAc beta 1-3Gal beta 1- 4Glc beta 1-1 ceramide. Enhanced expression of the Lex glycolipid may affect the organization of lens plasma membranes through Le(x)-Le(x) interactions, as suggested for compaction in mouse preimplantation embryos and embryonic teratocarcinomas, resulting in lens opacification, namely cataract.

Cataract↗

[The cytochemical demonstration of glycoproteins and glycolipids in Sarcocystis muris sarcocysts].

An electron cytochemical study of glycoproteins and glycolipids was made for the mature sarcocysts of Sarcocystis muris. Glycoprotein structures as branched fibrilles were seen on the surface of the sarcocyst wall. The fibrillar and granular glycoprotein structures were found in the ground substance of sarcocysts near the cyst wall and in the septae. In the plasmalemma of two types of cyst stages (merozoites and intermediate cells), glycoprotein fibrillar structures were revealed connecting these two cell types with each other. The third type cyst stages, i.e. the metrocytes, are situated separately without any fibrillar connections between them and other cyst stages being observed. This question is discussed in terms of the problem of cytodifferentiation. The fibrillar and granular glycoprotein material is scattered over the cytoplasm of the cyst stages, being especially concentrated in micronemes, rhoptries and around amylopectin granules. The control ultrathin sections were treated with saliva or pronase for the aims of protein identification in the material under study. In addition to glycoprotein, some glycolipids material was detected in the sarcocysts in the form of drops surrounded with thin glycoproteinaceous layers. Glycolipids were found in the ground substance of sarcocysts near the cyst stages and in the parasite cell cytoplasm around the micronemes and rhoptries. The data obtained are discussed in connection with the functional role glycoproteins and glycolipids play in S. muris.

Animals↗

Glycolipids in human lung carcinoma of histologically different types.

Glycolipids were isolated from primary human lung carcinoma tissue of various histologic types: adenocarcinoma, squamous cell carcinoma, and undifferentiated small cell carcinoma. Each type of carcinoma had a characteristic glycolipid pattern. The major glycolipids isolated were ceramide monohexosides, ceramide dihexosides, ceramide trihexosides, globoside, and hematoside. Squamous cell carcinoma and undifferentiated small cell carcinoma showed marked increases of ceramide monohexosides and dihexosides. Adenocarcinoma had a much higher level of the sulfatide (ceramide 3-sulfate-galactoside) as compared to squamous cell carcinoma, undifferentiated small cell carcinoma, or normal lung tissue. Embryonic tissue had more significant levels of sulfatide than did the other carcinomas. Adenocarcinoma had significantly lower levels of glycolipids due mainly to a decrease in the amount of ceramide monohexosides and dihexosides and hematoside.

Adenocarcinoma↗

Structure of sulfated glucuronyl glycolipids in the nervous system reacting with HNK-1 antibody and some IgM paraproteins in neuropathy.

Novel sulfated glucuronic acid-containing glycolipids have been identified in the nervous system. These glycolipids are highly antigenic and share antigenic determinants with several nervous system glycoproteins, such as neural cell adhesion molecules, myelin-associated glycoprotein, and ependymins. The structure of the major antigenic glycolipid from human peripheral nerve was determined by chemical and enzymatic degradation, incorporation studies, sugar analysis after permethylation, pertrimethylsilylation, and gas liquid chromatography-mass spectrometry techniques as well as fast atom bombardment-mass spectrometry of the native antigen. The following structure was established for the major antigenic glycolipid. sulfate-3-GlcA beta(1---3)Gal beta(1----4)GlcNAc beta(1----3)Gal beta(1----4)Glc beta(1----1)-ceramide. The major fatty acids in the ceramide were 18:0, 18:1, 24:0, and 24:1, with C18-sphingenine as the long chain base.

Antibodies↗

Glycolipids and gliomas. A review.

Several aspects of the relationships between glycolipids, gliomas, and problems involved in studying these are reviewed. Major problems that currently must be addressed include developing a classification scheme that more accurately predicts the biological behavior of these tumors, obtaining samples representative of the patient's glioma, and identifying appropriate control specimens. Results of analytical studies indicate that the patterns of gangliosides in human glioma tissues generally correlate with the degree of histological malignancy. Some gliomas contain unusual, and perhaps unique, glycolipids. Exogenous glycosphingolipids can alter cell shape and growth, which may involve modulation of growth factor receptors and protein kinases. Tumor glycolipids may also modulate immune functions in patients harboring gliomas. Transfection of nonneural cells with some oncogenes can alter glycolipid metabolism, which may be part of the mechanism through which these oncogenes exert their effects.

Animals↗