Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.4.99.7 (sialyltransferase)
 1,534 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mouse cells transformed by a temperature-sensitive mutant of simian virus 40 belonging to complementation group A lost their ability to regulate cell growth when grown at the permissive temperature (35 degrees) but showed the low saturation density of cell growth at the restrictive temperature (39.5 degrees) that is characteristic of normal cells in vitro. Biochemical analysis of the membranes of cells grown under the restrictive and the permissive conditions demonstrated no qualitative temperature-dependent differences either in neutral glycolipids or in acidic glycolipids of the cells. Plasma membrane glycoproteins labeled with radioactive glucosamine showed significantly different patterns on both polyacrylamide gel electrophoresis and electrofocusing. When the levels of glycoprotein glycosyltransferases of the cells were examined, the level of sialyltransferase (CMP-N-acetylneuraminytransferase,EC 2.4.99.1) of the cells grown at the restrictive temperature was low compared with that of cells grown at the permissive temperature. Our results indicate that the level of sialyltransferase is under the control of the gene A function of simian virus 40 and consequently is related to alterations in the cell surface glycoproteins.
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PMID:Alterations in surface glycoproteins and level of sialyltransferase of cells transformed by a temperature-sensitive mutant of simian virus 40. 18 85

A large family (14 children) with congenital goiter whose parents are first cousins was studied. Thyroid tissue was obtained, after 125I in vivo labeling, from one of the siblings (JBM). Gel filtration of thyroid proteins indicated that thyroglobulin (Tg) eluted as a single symmetrical peak in the same position as authentic 19S Tg. Gel electrophoresis in a 7.5% sodium dodecyl sulfate-polyacrylamide gel revealed a major band with the same mobility and immunoreactivity as normal 19S Tg. Hydrolysis of the patient's Tg indicated that most of the radioactivity was mono- and diiodotyrosines. The yield of T4 from JBM Tg (26 pmol/mg protein) was 5-fold less than normal thyroid tissue (140 pmol/mg protein) and approximately half of that in thyroid tissue from endemic goiter (51 pmol/mg). Total T3 released from JBM Tg was similar to the other two tissues. When the carbohydrate content of normal and patient Tg was analyzed, there was no differences in glucosamine, galactose or mannose content. However, unlike normal and endemic-goiter Tg, that had a mean sialic acid content of 7.3 and 5.6 micrograms/mg protein, respectively, the sialic acid concentration of the patients Tg was only 0.3 microgram/mg. Sialyltransferase activity was readily demonstrated in homogenate from normal thyroid or endemic goiter, but no sialyltransferase activity was detectable in a homogenate of JBM-thyroid tissue. We conclude that the finding of severely hyposialylated Tg is linked to a defect in iodotyrosine coupling seen in this patient with a possibly abnormal migration of Tg into the follicular lumen.
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PMID:Hyposialylated thyroglobulin in a patient with congenital goiter and hypothyroidism. 172 28

Incubation of rat jejunal slices in Krebs-Ringer bicarbonate buffer (KRB) required the presence of heat-inactivated horse serum (HHS) in order to show time-dependent release of sialyltransferase into the medium. Sialyltransferase activity could not be detected in the medium when KRB alone or KRB supplemented with either albumin or glycerol was used in the incubations. The viability of the jejunal slices for up to 4 h of incubation was determined by studying the incorporation of glucosamine and leucine into acid-insoluble proteins. Supplementation of KRB with HHS had no beneficial effect on the rate of incorporation of leucine and glucosamine into proteins. KRB medium obtained after different periods of incubation contained higher trypsin-like activity than KRB medium containing HHS. Various antiproteases present as supplements to KRB resulted in the release of sialyltransferase activity from the jejunal slices. Among these antiproteases, alpha 1-proteinase inhibitor (alpha 1-PI) was the most effective. Also, HHS added to KRB immediately following incubation resulted in partial restoration of sialyltransferase activity in the medium, suggesting the presence of anti-proteolytic factors in HHS. The addition of increasing concentrations of heparin to incubations containing HHS caused a decrease in the medium sialyltransferase activity. The heparin-binding fraction (HBF) from HHS, when added to incubations, was able to protect the sialyltransferase released into medium. However, HHS depleted of its heparin-binding fraction by heparin-agarose affinity chromatography was unable to protect the sialyltransferase. HBF was separated into high- and low-molecular-mass fractions (fractions A and B respectively) by gel-filtration chromatography. The capacity to protect the released sialyltransferase was contained in fraction B. Fraction A contained multiple bands on SDS/PAGE and did not protect the enzyme. Fraction B contained a major protein band on the gel which corresponded to the migration of a similar band in human alpha 1-PI. HBF as well as fraction B isolated from HHS showed anti-trypsin-like activity. The results presented indicate that HHS contains a heparin-binding protein(s) similar to human alpha 1-PI which plays a role in the protection of sialyltransferase released from jejunal slices.
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PMID:Heparin-binding serum protein(s) is required for the protection of sialyltransferase released during the incubation of rat jejunal slices. 176 33

We found that rhabdomyosarcoma (RMS) subcellular membranes contain sialyltransferase activities for LcOse4Cer and GgOse4Cer acceptors. Chromatographic analyses and neuraminidase lability of the sialyltransferase products indicated that the principal site of sialylation was the non-reducing terminal galactosyl moiety. In order to control for the effects of cell density in culture, metastatic S4T18 RMS cells and nonmetastatic F9-4/21 RMS cells were harvested at 2 X 10(4) to 6 X 10(4) per cm2 prior to analyses. Irrespective of metastatic potential, we found that sialyltransferase-specific activities were influenced by cell densities. F9-4/21 cells, for example, at a density of 6 X 10(4), produced membranes with sialyltransferase-specific activities to LcOse4Cer 1.9-fold higher than cells at 2.1 X 10(4)/cm2. Metastatic potential (predetermined in vivo) appeared to be correlated with an accelerated effect of cell density on the sialyltransferase activity to LcOse4Cer. Metastatic S4T18 cells at 6.3 X 10(4)/cm2 yielded membranes with sialyltransferase-specific activities 5.4-fold higher than membranes from cells at 1.9 X 10(4)/cm2. Conversely, fucosyltransferase activities in the presence of LcOse4Cer were highest in non-metastatic F9-4/21 cells at low cell densities. Quantitative analyses of monosialoganglioside fractions of RMS cells were in agreement with the sialyl-transferase studies. HPLC and HPTLC analyses demonstrated the presence of glucosamine-containing monosialoganglioside with Rf identical with the radioactive products of LcOse4Cer sialylation, which increased 4.5-fold on a per mg protein basis as cell densities increased in S4T18 cells in culture from 1.9 X 10(4)/cm2 to 6.3 X 10(4)/cm2. Plasma membrane marker Na+, K+, ATPase-specific activity also increased in RMS metastatic cells in a manner comparable to that described for the sialyl-transferase activity to LcOse4Cer. Our results suggest that metastatic potential is expressed in the rate of sialylation at specific membrane sites of RMS intercellular contact. We propose a process of selection for metastasis whereby specific cell surface non-reducing galactosyl termini are recognized by intercellular transferases and lectins in the primary tumor, and the corresponding labile sialylated sites (on disseminated cells) are recognized by host neuraminidases.
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PMID:Monosialoganglioside biosynthesis by subcellular membranes of rhabdomyosarcoma cell lines differing in metastatic potential. 233

Cell line MDA 886Ln was established from a laryngeal lymph node metastasis. When grown as a multicellular tumor spheroid (MTS), it exhibits squamous differentiation. We studied the effects of beta-all-trans retinoic acid (RA) on the growth, differentiation and glycoprotein content of this MTS model for squamous carcinomas of the head and neck. The growth of MTSs was inhibited in a dose-dependent manner by 10(-6) to 10(-10) M RA. Growth inhibition occurred between 3 and 5 days of RA treatment (10(-6)M). Immunohistochemical and electrophoretic analyses revealed that RA suppressed the morphological markers of squamous differentiation (squames), involucrin expression, and keratin expression. Gly-coprotein expression was examined by metabolic labelling using 3H-glucosamine, in situ labelling of polyacrylamide gels with 125I-labelled wheat-germ agglutinin (WGA), localization of fluorescein isothionate-WGA in frozen sections, and determination of sialyltransferase activity. Treatment using 10(-6) M RA altered glycoprotein expression both biochemically and morphologically, and WGA was shown to bind preferentially to sialic acid residues. The sensitivity of this MTS model to RA treatment and its ability to be analyzed through morphological, immunohistochemical and biochemical techniques suggest that it will prove useful in studying the relationships between growth, differentiation and RA-induced alterations in squamous carcinomas.
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PMID:Modulation of growth, differentiation and glycoprotein synthesis by beta-all-trans retinoic acid in a multicellular tumor spheroid model for squamous carcinoma of the head and neck. 247 9

Previous studies have shown that treatment of S91-C2 murine melanoma cells with beta-all-trans-retinoic acid (RA) results in growth inhibition, enhanced activity of sialyltransferase, and increased glycosylation of a Mr 160,000 cell surface sialoglycoprotein (gp160). None of these effects could be detected in mutant clones (e.g., S91-C154) selected from the S91-C2 cells for resistance to RA-induced growth inhibition. These findings suggest that modulation by RA of gp160 might be related causally to growth inhibition. In this study we examined the possible role of gp160 in growth regulation using specific antibodies to this glycoprotein. Metabolic labeling of S91-C2 cells with either [3H]glucosamine or [35S]methionine revealed that the cells shed into the growth medium a gp160-like glycoprotein, in addition to several other macromolecules. The gp160-like glycoprotein was isolated from concentrated conditioned medium after preparative polyacrylamide slab gel electrophoresis in the presence of sodium dodecylsulfate by excision of the corresponding protein band. Rabbits were immunized with this material and immunoblotting revealed that their sera contained antibodies that bound specifically to gp160 in extracts of untreated or RA-treated S91-C2 cells. Indirect immunofluorescence staining followed by fluorescence-activated cell sorter analysis demonstrated that the anti-gp160 antibodies bound to the surface of both untreated and RA-treated S91-C2 cells and that the treated cells bound more of the antibodies than untreated ones. In contrast, these antibodies bound to the same extent to untreated and RA-treated resistant S91-C154 cells. The growth of S91-C2 cells in the presence of anti-gp160 antibodies in semisolid medium as well as in monolayer cultures was inhibited in a dose-dependent fashion. Fifty % growth inhibition was obtained at an immunoglobulin concentration of 10 micrograms/ml. The growth of cells exposed concurrently to RA and anti-gp160 antibodies was also inhibited strongly in semisolid medium, but the antibodies caused only a small increase in the inhibitory effect of RA in monolayer cultures. No inhibition by the antibodies of either anchorage-independent growth or anchorage-dependent growth of S91-C154 cells, grown in the absence or presence of RA, was observed. These results support the suggestion that cell surface gp160 might be involved in growth regulation in the S91-C2 cells.
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PMID:Growth inhibition of murine melanoma cells by antibodies to a cell surface glycoprotein implicated in retinoic acid action. 355 69

Because alterations in cell membrane sialoglycoconjugates can affect the behavior of neoplastic cells, we investigated the effects of in vitro treatment with antimetabolites used in cancer therapy on the expression of membrane sialic acid in cultured HL-60 leukemic cells. In these studies, cells were incubated with Vibrio cholerae neuraminidase to remove surface sialic acid. Reappearance of membrane sialic acid during drug treatment was followed (a) by measuring changes in radioactive surface labeling of viable cells with sodium metaperiodate-sodium[3H]-borohydride, (b) by measuring the decline in accessible surface galactosyl receptor sites which occurred coincident with membrane sialic acid replacement, and (c) by measuring the incorporation of [3H]glucosamine into membrane-associated neuraminidase-labile sialic acid. We were especially interested in learning whether drugs that affect intracellular pools of cytidine triphosphate (CTP), an important nucleotide intermediate in sialylation reactions, could inhibit regeneration of membrane sialic acid. 3-Deazauridine, a competitive inhibitor of CTP synthetase, depleted CTP pools and curtailed surface membrane resialylation with little or no effect on synthesis of de novo sialic acid from precursor sugars. The addition of cytidine restored CTP pools and sialic acid regeneration. Acivicin, a glutamine antagonist, also depleted CTP pools and curtailed surface membrane resialylation. In addition, it retarded de novo synthesis of sialic acid. The addition of cytidine restored intracellular CTP pools and sialic acid regeneration. However, both cytidine and guanosine were required to restore sialic acid synthesis from precursor sugars. 1-beta-D-Arabinofuranosylcytosine, a competitive inhibitor of sialic acid synthetase and of sialyltransferase, inhibited both de novo sialic acid synthesis and membrane resialylation. Only the latter effect was reversed by the addition of exogenous cytidine. Hydroxyurea, an agent shown previously to inhibit glycoconjugate production in hamster fibroblasts, curtailed membrane resialylation and de novo synthesis of sialic acid without depleting CTP pools. Doxorubicin, at levels that caused marked arrest of cell proliferation, had no effect on sialic acid synthesis or expression on the membrane surface. These data suggest that antimetabolites, apart from their cytotoxic effects or effects on cellular growth, may directly inhibit the expression of membrane sialic acid.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Effects of pyrimidine antagonists on sialic acid regeneration in HL-60 cells. 385 65

Retinoic acid inhibits the proliferation of the murine melanoma clone S91-C-2 cells, enhances the glycosylation of specific cell surface sialoglycoproteins, and stimulates sialytransferase activity. Mutant clones, selected from the S91-C-2 cells for resistance to the growth-inhibitory effect of retinoic acid, were used to explore whether cell surface modulation by retinoic acid is related to growth inhibition. Glycoprotein synthesis was assessed by analysis of [3H]glucosamine incorporation into glycoconjugates, and cell surface sialo- and galactoglycoproteins were analyzed after radiolabeling by the NaIO4:NaB3H4 and the neuraminidase plus galactose oxidase:NaB3H4 methods, respectively. The cells were solubilized and the labeled molecules were separated by polyacrylamide gel electrophoresis and identified by fluorography. Sialytransferase activity was measured in detergent-solubilized cells, using cytidine 5' -monophosphate-[14C]sialic acid as a sugar donor and asialofetuin as an exogenous acceptor. The results demonstrated that retinoic acid enhanced [3H]glucosamine incorporation into a Mr 160,000 glycoprotein in the S91-C-2 cells but not in any of the resistant mutant clones, while the pattern of [35S]methionine-labeled proteins was not modified in either the sensitive or the resistant clones. Radiolabeling of a Mr 160,000 sialoglycoprotein on the surface of S91-C-2 and of several retinoic acid-sensitive subclones of S91-C-2 was augmented by retinoic acid. A considerably smaller effect was observed on the labeling of Mr 160,000 sialoglycoprotein on one of the resistant clones, and no significant effect could be detected on the other resistant mutant clones. Sialytransferase activity was increased 2- to 3-fold by retinoic acid in the S91-C-2 cells and in several sensitive subclones, but not in any of the resistant mutant clones. Tetradecanoylphorbol acetate, which inhibits the proliferation of both retinoic acid-sensitive and retinoic acid-resistant cells, failed to increase either sialyltransferase activity or cell surface labeling of sialoglycoproteins. These findings suggest that the ability of retinoic acid to stimulate sialyltransferase activity and glycosylation of cell surface glycoproteins is related to the growth-inhibitory effect of this compound.
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PMID:Correlation of retinoic acid-enhanced sialyltransferase activity and glycosylation of specific cell surface sialoglycoproteins with growth inhibition in a murine melanoma cell system. 649 40

In L1210 leukemia cells, 6-deoxy-6-fluoro-D-galactose specifically inhibited the incorporation of [3H]-D-galactose, while that of other precursors of glycoconjugate biosynthesis, including mannose and glucosamine, was unaffected. The activation of [6-3H]-6-deoxy-6-fluoro-D-galactose to a nucleotide sugar was similar to that found for [3H]-D-galactose. The incorporation of either sugar after 1 hr was visualized by electron microscopic autoradiography to be in the Golgi region. Treatment of L1210 cells with 6-deoxy-6-fluoro-D-galactose in vitro or in vivo resulted in a specific, dose- and time-dependent decrease in the activity of cell surface sialyltransferase (ectosialyltransferase) but not of 5'-nucleotidase, a plasma membrane marker enzyme. The decrease in ectosialyltransferase activity appeared to be selective and is suggested to be due to structural modification of the cell surface galactoprotein acceptors for this enzyme. The data indicate that 6-deoxy-6-fluoro-D-galactose is an effective modifier of cellular glycoconjugate in that its incorporation into certain cell surface components results in a modification of plasma membrane structure and function.
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PMID:Effects of a membrane sugar analogue, 6-deoxy-6-fluoro-D-galactose, on the L1210 leukemic cell ectosialyltransferase system. 684 4

Total labeled glycolipids from thymocytes of leukemic AKR/J mouse thymus incubated for 24 h in the presence of the precursors [3H] galactose or [14C] glucosamine were found to exceed those from non-leukemic thymocytes. The amount of labeled compounds that co-migrated with monosialogangliosides (GM3 and GM2) and disialogangliosides (GD1a and GD1b) was higher, whereas incorporation into monosialoganglioside (GM1) and trisialoganglioside (GT1) was lower in leukemic than in non-leukemic thymocytes. Consistent with this altered pattern of ganglioside distribution was the finding of a higher activity of two of the sialyltransferase activities in homogenates of leukemic thymus as compared to normal thymus. About 2-fold higher activities of the enzymes CMP-AcNeu: lactosylceramide sialyltransferase and CMP-AcNeu: GM1 sialyltransferase were observed in leukemic thymus homogenate compared to homogenates of non-leukemic cells. The substrate affinity of sialyltransferase with GM1 as acceptor from the leukemic thymus was similar to that of the enzyme from normal thymus. Thus, our studies reveal an enzymatic basis for the enhanced rate of synthesis and altered ganglioside profile observed in malignant thymocytes, and are consistent with the general view on the pattern of acidic glycolipids in tumorigenesis.
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PMID:Glycolipid sialyltransferases in normal and neoplastic murine thymocytes. 731 48


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