Cell surface glycosyl transferase activities in liver cells of developing chicken embryos

Isolated sinusoidal endothelial rat liver cells (EC) in suspension bound and internalized ovalbumin, a mannose-terminated glycoprotein, in a saturable manner. The binding and uptake were Ca2+-dependent and were effectively inhibited by alpha-methyl mannoside and yeast mannan, but not by galactose or asialoglycoproteins. This corresponds to the binding specificity described for the mannose receptor of macrophages and non-parenchymal liver cells. Binding studies indicated a surface pool of 20,000-25,000 mannose receptors per cell, with a dissociation constant of 6 x 10(-8) M. Uptake and degradation of ovalbumin by isolated EC were inhibited by weak bases and ionophores which inhibit acidification of endocytic vesicles and dissociation of receptor-ligand complexes. Cycloheximide had no effect on uptake or degradation. Degradation, but not uptake, was inhibited by leupeptin. We conclude that ovalbumin dissociates from the mannose receptors in the endosomal compartment and the receptors are recycled to the cell surface, while the ovalbumin is directed to the lysosomes for degradation. A fraction of the internalized ovalbumin was recycled intact to the cell surface and escaped degradation (retroendocytosis). The rate of internalization of ovalbumin by isolated EC was very fast, with a Ke (endocytotic rate constant) of 4.12 min-1, which corresponds to a half-life of 10 s for the surface pool of receptor-ligand complexes. To our knowledge, this is the highest Ke reported for a receptor-mediated endocytosis system.

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Endocytosis of lysosomal enzymes by non-parenchymal rat liver cells. Comparative study of lysosomal-enzyme uptake by hepatocytes and non-parenchymal liver cells

Biochemical Journal ◽

10.1042/bj1820329 ◽

1979 ◽

Vol 182 (2) ◽

pp. 329-335 ◽

Cited By ~ 28

Author(s):

Kurt Ullrich ◽

Volkmar Gieselmann ◽

Günther Mersmann ◽

Kurt Von Figura

Keyword(s):

Cell Surface ◽

Rat Liver ◽

Lysosomal Enzymes ◽

High Efficiency ◽

Indirect Evidence ◽

Liver Cells ◽

Cell Surface Receptors ◽

Surface Receptors ◽

Parenchymal Liver ◽

Rat Liver Cells

Cultured non-parenchymal rat liver cells internalize human urine α-N-acetylglucosaminidase, human skin β-N-acetylglucosaminidase and pig kidney α-mannosidase. Different heat-stabilities of endocytosed and endogenous α-mannosidase activity provided indirect evidence that the increase in intracellular activity resulted from uptake. The high efficiency and the saturation kinetics of uptake indicated that these enzymes become internalized by adsorptive endocytosis. Competition experiments with glycoproteins bearing known carbohydrates at their non-reducing terminals, with mannans, methyl glycosides and monosaccharides, established that the uptake of these three lysosomal enzymes is mediated by the binding to cell-surface receptors that recognize mannose and N-acetylglucosamine residues. The decreased uptake after treatment of these enzymes with either β-N-acetylglucosaminidase or α-mannosidase was in accordance with the results of the inhibition experiments. Removal of oligosaccharides of the high-mannose type by treatment with endoglucosaminidase H inhibited uptake almost completely, suggesting that the sugars recognized by cell-surface receptors of non-parenchymal liver cells are located in the outer core of these oligosaccharides. A comparison of the uptake of these three lysosomal enzymes by parenchymal and non-parenchymal rat liver cells indicates that infused α-N-acetylglucosaminidase is taken up preferentially by hepatocytes, whereas α-mannosidase and β-N-acetylglucosaminidase are localized predominantly in non-parenchymal rat liver cells.

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Role of cell surface receptor mobility in concanavalin A-induced agglutination of Novikoff hepatoma and normal rat liver cells

Journal of Cell Science ◽

10.1242/jcs.42.1.169 ◽

1980 ◽

Vol 42 (1) ◽

pp. 169-182

Author(s):

P. Mastromarino ◽

G. Neri ◽

A. Serra ◽

E.F. Walborg

Keyword(s):

Cell Surface ◽

Rat Liver ◽

Liver Cells ◽

Cell Surface Receptor ◽

Lateral Mobility ◽

Con A ◽

Novikoff Hepatoma ◽

Surface Fluorescence ◽

Rat Liver Cells ◽

Normal Rat

The relationshio between Con A-receptor mobility and Con A-induced agglutination of Novikoff hepatoma and normal rat liver cells was investigated. Novikoff cells, incubated with fluorescein-labelled Con A at 3 degrees C displayed uniform, ring-like surface fluorescence. Increasing the temperature of the cells to 37 degrees C caused capping of Con A receptors in approximately 65% of the cells, a phenomenon that could be prevented by prefixing the cells with glutaraldehyde. In spite of these variations in Con A-receptor distribution, Con A-induced agglutination was remarkably constant over a temperature range from 3 to 37 degrees C. In contrast to Novikoff cells, normal hepatocytes displayed a uniform, ringlike surface fluorescence at both 3 and 37 degrees C. No capping was observed. However, hepatocytes, similar to Novikoff cells, were agglutinable by low concentrations of Con A. These findings indicate that, in this model system, Con A-induced cytoagglutination is not dependent upon long-range lateral mobility of Con A receptors. The qualitative differences in the lateral mobility of cell-surface Con A receptors of normal and malignant rat liver cells may represent a marker for neoplastioc transformation during hepatocarcinogenesis, adaptation to growth in ascitic form, or progression of a tumour to a more malignant state.

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Cell Surface Sialoglycopeptide Metabolism and Surface Glycosyl Transferase Activity in the Ehrlich Ascites Cell

Canadian Journal of Biochemistry ◽

10.1139/o75-122 ◽

1975 ◽

Vol 53 (8) ◽

pp. 895-902 ◽

Cited By ~ 5

Author(s):

D. Irwin ◽

T. P. Anastassiades

Keyword(s):

Sialic Acid ◽

Cell Surface ◽

Transferase Activity ◽

Intact Cells ◽

Neuraminidase Treatment ◽

Glycosyl Transferase ◽

Ehrlich Ascites ◽

Ehrlich Ascites Cell ◽

Almost All

1. The in vivo incorporation of radioactivity from [3H]glucosamine into a trypsin labile, cell surface sialoglycopeptide fraction (SGP) of Ehrlich ascites cells was studied in the presence and absence of puromycin pretreatment. The results indicated a much more complete inhibition of incorporation into the surface SGP than in the average intracellular acid insoluble glycoproteins. No evidence of turnover of the carbohydrate portion of the surface SGP independent of protein synthesis could be obtained.2. However, when intact cells were incubated with labelled uridine 5′-diphosphate – N-acetyl glucosamine or cytidine 5′-monophosphate (CMP) – sialic acid there was some incorporation largely into acid insoluble material, suggesting the presence of glycosyl transferase activity at the surface. Further evidence for surface activity was obtained when neuraminidase pretreatment of intact cells stimulated incorporation of labelled CMP – sialic acid sixfold and almost all of the incorporated counts could be released by subsequent neuraminidase treatment. Furthermore, a much greater proportion of the incorporated counts could be released by papain than by trypsin treatment of the intact cells. These results suggest that the surface acceptor for exogenously added CMP – sialic acid is not identical to the endogenously synthesized trypsin labile surface SGP.

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Endocytosis of β-N-acetylglucosaminidase from sections of mucolipidosis-II and-III fibroblasts by non-parenchymal rat liver cells

Biochemical Journal ◽

10.1042/bj1820245 ◽

1979 ◽

Vol 182 (1) ◽

pp. 245-247 ◽

Cited By ~ 12

Author(s):

K Ullrich ◽

K von Figura

Keyword(s):

Cell Surface ◽

Rat Liver ◽

Liver Cells ◽

Cell Surface Receptors ◽

Surface Receptors ◽

Rat Liver Cells ◽

Mucolipidosis Ii

beta-N-Acetylglucosaminidase isolated from the secretions of fibroblasts of mucolipidosis-II and -III patients is internalized by cultured non-parenchymal rat liver cells. The rate of endocytosis compared with that of beta-N-acetylglucosaminidase from control fibroblasts was 11 and 19% for the enzyme from mucolipidosis-II and -III patients respectively. The inhibition of endocytosis by mannan indicates that the beta-N-acetylglucosaminidase from mucolipidosis-II and -III patients is recognized by cell-surface receptors specific for mannose.

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