Cell surface glycoproteins change during gastrulation in Pleurodeles waltlii

1986 ◽  
Vol 82 (1) ◽  
pp. 23-40
Author(s):  
J.F. Riou ◽  
T. Darribere ◽  
J.C. Boucaut
Keyword(s):  
Cell Surface ◽  
Gel Electrophoresis ◽  
Lectin Binding ◽  
Galactose Oxidase ◽  
Two Dimensional ◽  
Isolated Cells ◽  
Two Dimensional Gel Electrophoresis ◽  
Sodium Metaperiodate ◽  
Cell Surface Glycoproteins ◽  
Surface Glycoproteins

Identification of the cell surface glycoproteins is an essential requirement for elucidating the mechanisms that mediate intercellular adhesion and cell migration in gastrulating amphibian embryos. In our experiments the glycoproteins present at the surface of isolated cells at blastula and gastrula stages were labelled with 3H-labelled sodium borohydride. The procedure included the oxidation of either galactosyl and, or, N-acetylgalactosaminyl residues with galactose oxidase or sialic acid with sodium metaperiodate. The tritiated components were analysed by two-dimensional gel electrophoresis. On the surface of isolated embryonic cells, 23 glycoproteins have been identified. The most important observation was the labelling of 15 new glycoproteins during gastrulation. At the late blastula stage, eight glycoproteins containing galactosyl or N-acetyl-galactosaminyl residues are labelled, while 23 are labelled at the late gastrula stage. In other experiments, glycoproteins labelled by [3H]fucose or [3H]mannose or precipitated by concanavalin A(ConA)-, or wheat-germ agglutinin (WGA)-Sepharose were studied by two-dimensional gel electrophoresis in order to provide supplementary data on the synthesis and lectin-binding properties of major cell surface glycoproteins. It appears that approximately 100 different glycoproteins are revealed by these methods. Among them, 13 have been identified as major cell surface glycoproteins, 12 being labelled by [3H]mannose, four by [3H]fucose, 10 bearing an affinity for ConA and five for WGA. These 13 glycoproteins are synthesized throughout gastrulation without qualitative variation.

The accessibility of certain proteins on embryonic chick neural retina cells to iodination and tryptic removal is altered by calcium

1982 ◽  
Vol 55 (1) ◽  
pp. 85-103
Author(s):  
J.H. Cook ◽  
J. Lilien
Keyword(s):  
Cell Surface ◽  
Gel Electrophoresis ◽  
Adhesive System ◽  
Neural Retina ◽  
Galactose Oxidase ◽  
Two Dimensional ◽  
Embryonic Chick ◽  
Two Dimensional Gel Electrophoresis ◽  
Oligosaccharide Moiety ◽  
Specific Labelling

We have used cell-surface-specific labelling techniques and two-dimensional gel electrophoresis to identify proteins on embryonic chick neural retina cells and to determine the effects of Ca2+ on their accessibility to labelling and tryptic removal. A number of proteins on these cells are, in the presence of Ca2+, relatively inaccessible to iodination and/or tryptic removal. Of these, a glycoprotein of Mr approx. 130 × 10(3), with a pI of approx. 4.8, is the major cell-surface-iodinatable species that is retained during trypsinization in the presence of Ca2+. The removal of Ca2+ renders this glycoprotein much more accessible to both procedures. Its accessibility to these probes decreases on re-addition of Ca2+. The accessibility of its oligosaccharide moiety to galactose oxidase is, however, unaltered by the removal of Ca2+. These characteristics, together with immunological data presented elsewhere suggest that this glycoprotein may be a component of the Ca2+-dependent adhesive system that can be demonstrated on these cells.

scholarly journals Identification of phagocytosis-associated surface proteins of macrophages by two-dimensional gel electrophoresis.

1982 ◽  
Vol 92 (2) ◽  
pp. 283-288 ◽  
Author(s):  
F D Howard ◽  
H R Petty ◽  
H M McConnell
Keyword(s):  
Cell Surface ◽  
Gel Electrophoresis ◽  
Protein Composition ◽  
Surface Protein ◽  
Surface Proteins ◽  
Two Dimensional ◽  
Cell Surface Protein ◽  
Two Dimensional Gel Electrophoresis ◽  
Reducing Conditions ◽  
Membrane Components

Two-dimensional PAGE (P. Z. O'Farrell, H. M. Goodman, and P. H. O'Farrell. 1977. Cell. 12:1133-1142) has been employed to assess the effects of antibody-dependent phagocytosis on the cell surface protein composition of RAW264 macrophages. Unilamellar phospholipid vesicles containing 1% dinitrophenyl-aminocaproyl-phosphatidylethanolamine (DNP-cap-PE) were used as the target particle. Macrophages were exposed to anti-DNP antibody alone, vesicles alone, or vesicles in the presence of antibody for 1 h at 37 degrees C. Cell surface proteins were then labeled by lactoperoxidase-catalyzed radioiodination at 4 degrees C. After detergent solubilization, membrane proteins were analyzed by two-dimensional gel electrophoresis. The resulting pattern of spots was compared to that of standard proteins. We have identified several surface proteins, not apparently associated with the phagocytic process, which are present either in a multichain structure or in several discretely charged forms. After phagocytosis, we have observed the appearance of two proteins of 45 and 50 kdaltons in nonreducing gels. In addition, we have noted the disappearance of a 140-kdalton protein in gels run under reducing conditions. These alterations would not be detected in the conventional one-dimensional gel electrophoresis. This evidence shows that phagocytosis leads to a modification of cell surface protein composition. Our results support the concept of specific enrichment and depletion of membrane components during antibody-dependent phagocytosis.

scholarly journals Antisera inhibiting mammalian cell spreading and possible cell surface antigens involved.

1980 ◽  
Vol 86 (3) ◽  
pp. 866-873 ◽  
Author(s):  
P Hsieh ◽  
N Sueoka
Keyword(s):  
Cell Surface ◽  
Cell Line ◽  
Cell Lines ◽  
Gel Electrophoresis ◽  
Critical Role ◽  
Cell Spreading ◽  
Surface Proteins ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Cell Surface Proteins

Antiserum against a rat neuronal tumor cell line (B103) has been prepared in rabbit by intravenous injection of live cells. This immune serum (anti-B103) precipitates a few cell surface proteins recognizable by two-dimensional gel electrophoresis as common radioiodinatable spots in 15 different rat neural cell lines and in mouse and rat fibroblast cell lines. The apparent molecular weight of one major common protein (II4) is estimated by SDS gel electrophoresis to be somewhere between 80,000 and 90,000 and another protein (I3) to be 120,000. These two proteins are consistently recognized in various cell lines by this antiserum. Furthermore, at a 1:20 dilution, this serum causes monolayer cells to round up usually within 0.5 h and detach from the plate within 3 h. It also inhibits spreading of freshly plated cells. These effects of the antiserum are reversible. Upon absorption of the antiserum with cells (e.g., absorbed with a glial cell line, B27), the serum no longer causes the rounding up of the monolayer cells, it does not inhibit cell spreading, and it does not immune-precipitate the two common proteins from the cell surface of various cell lines. Antisera against several other rat cell lines also precipitate the same common proteins (II4 and I3) from the cell surface and prevent cell spreading. These data suggest that the antibody acts first at the cell surface and then inhibits cell spreading or rounding up of spread cells. The consistent pattern of the immunoprecipitated cell surface proteins on the two-dimensional gel electrophoresis makes these two common surface proteins (II4 or I3 or both) possible candidates for target proteins to which the antibody binds. Thus, they may play a critical role in cell spreading.

scholarly journals Differences between the carbohydrate units of cell-surface glycoproteins of moust B- and T-lymphocytes

1979 ◽  
Vol 181 (2) ◽  
pp. 451-456 ◽  
Author(s):  
T Krusius ◽  
J Finne ◽  
L C Andersson ◽  
C G Gahmberg
Keyword(s):  
T Lymphocytes ◽  
Cell Surface ◽  
Affinity Chromatography ◽  
Concanavalin A ◽  
Cell Types ◽  
Galactose Oxidase ◽  
Neuraminidase Treatment ◽  
Cell Surface Glycoproteins ◽  
B And T Lymphocytes ◽  
Surface Glycoproteins

Carbohydrate units of cell-surface glycoproteins of mouse B- and T-lymphocytes, labelled in their sialic acid residues by the periodate/NaB3H4 method and in their galactose residues by the galactose oxidase/NaB3H4 method after neuraminidase treatment, have been studied. Glycopeptides were prepared from the labelled cells by Pronase digestion and fractionated by concanavalin A affinity chromatography into two fractions (A and B). Alkali-labile oligosaccharides were isolated after mild NaOH/NaBH4 treatment by gel filtration. The alkali-labile oligosaccharides were further analysed by t.l.c. To study the relative proportion of neutral mannose-rich carbohydrate units (fraction C) in lymphocyte glycoproteins, glycopeptides were also prepared from unlabelled cells and subjected to concanavalin A affinity chromatography after N-[3H]acetylation of their peptide moiety. The major alkali-labile oligosaccharide component of both cell types was identified as galactosyl-(beta 1 leads to 3)-N-acetylgalactosaminitol. T-Lymphocytes were characterized by a high proportion of this oligosaccharide and a lower proportion of alkali-stable fraction A glycopeptides, whereas the opposite was observed for B-lymphocytes. The relative proportions of the concanavalin A-binding fractions B and C were similar in both cell types. The differences observed may correlate with the different surface properties of B- and T-lymphocytes.

Developmental regulation of cell-surface glycoproteins in clonal cultures of human skeletal muscle satellite cells

1989 ◽  
Vol 67 (2-3) ◽  
pp. 128-136 ◽  
Author(s):  
Shashikant Champaneria ◽  
Paul C. Holland ◽  
George Karpati ◽  
Claude Guérin
Keyword(s):  
Skeletal Muscle ◽  
Cell Surface ◽  
Satellite Cells ◽  
Human Skeletal Muscle ◽  
Developmental Regulation ◽  
Lectin Binding ◽  
Primary Cultures ◽  
Surface Proteins ◽  
Cell Surface Glycoproteins ◽  
Surface Glycoproteins

Pure populations of myogenic cells were obtained by cloning satellite cells from human skeletal muscle biopsies. Cell-surface glycoproteins at various stages of myogenesis were analysed by one- and two-dimensional gel electrophoresis. A total of 14 distinct proteins were detectable at the cell surface, on the basis of their susceptibility to desialation by exogenous neuraminidase or their iodination by exogenous lactoperoxidase. Reproducible changes in lectin binding or iodination of eight of these proteins occurred during myogenesis. Only two of the developmentally regulated proteins were components of the detergent-insoluble extracellular matrix fraction. Developmental regulation of these two proteins was unaffected by growth of cultures in 5-bromo-2′-deoxyuridine to inhibit myogenesis. In contrast, developmental regulation of the other cell-surface proteins was inhibited by growth in 5-bromo-2′-deoxyuridine, suggesting that changes in these proteins are tightly coupled to satellite cell differentiation. These studies represent the first systematic analysis of the surface proteins of pure, clonally derived, primary cultures of normal myogenic cells.Key words: satellite cells, myogenesis, myoblast, glycoproteins, cell surface.

Diversity of Glycoprotein Deficiencies in Glanzmann’s Thrombasthenia

1985 ◽  
Vol 54 (03) ◽  
pp. 626-629 ◽  
Author(s):  
M Meyer ◽  
F H Herrmann
Keyword(s):  
High Resolution ◽  
Gel Electrophoresis ◽  
Type Ii ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Structural Variant ◽  
Glanzmann’S Thrombasthenia ◽  
Crossed Immunoelectrophoresis ◽  
Glanzmann's Thrombasthenia ◽  
Platelet Proteins

SummaryThe platelet proteins of 9 thrombasthenic patients from 7 families were analysed by high resolution two-dimensional gel electrophoresis (HR-2DE) and crossed immunoelectrophoresis (CIE). In 7 patients both glycoproteins (GPs) IIb and Ilia were absent or reduced to roughly the same extent. In two related patients only a trace of GP Ilb-IIIa complex was detected in CIE, but HR-2DE revealed a glycopeptide in the position of GP Ilia in an amount comparable to type II thrombasthenia. This GP Ilia-like component was neither recognized normally by anti-GP Ilb-IIIa antibodies nor labeled by surface iodination. In unreduced-reduced two-dimensional gel electrophoresis two components were observed in the region of GP Ilia. The assumption of a structural variant of GP Ilia in the two related patients is discussed.

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