SHEDDING OF SURFACE PROTEINS BY PORCINE THYROID CELLS

1980 ◽  
Vol 85 (2) ◽  
pp. 245-251 ◽  
Author(s):  
A. BRENNAN ◽  
P. M. POVEY ◽  
B. REES SMITH ◽  
R. HALL
Keyword(s):  
Cell Surface ◽  
Sodium Dodecyl Sulphate ◽  
Culture Medium ◽  
Cell Metabolism ◽  
Sodium Dodecyl ◽  
Surface Proteins ◽  
Half Time ◽  
Thyroid Cells ◽  
Peptide Cleavage ◽  
Membrane Bound

Isolated porcine thyroid cells were surface-labelled with 125I using the lactoperoxidase technique. Samples of the cells were then cultured and harvested at various intervals for up to 7 days. The labelled proteins remaining on the cells or shed into the culture medium were analysed by electrophoresis on polyacrylamide gels run in sodium dodecyl sulphate. These studies indicated that the several different surface proteins of the thyroid cells were lost from the cell surface at similar rates (half-time of approximately 28 h) as the result, at least in part, of a process which depended on active cell metabolism. In addition, the gel profiles obtained from analysis of both medium and membrane-bound labelled proteins were similar and this suggested that peptide cleavage was not involved in the shedding of the majority of these proteins.

scholarly journals Cell-surface radioiodination with the sparingly soluble catalyst Iodogen. Difference between the dividing and non-dividing populations of rodent thymocytes

1981 ◽  
Vol 194 (1) ◽  
pp. 351-355 ◽  
Author(s):  
J G Salisbury ◽  
J M Graham
Keyword(s):  
Cell Surface ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Surface Proteins ◽  
New Method

The surface proteins of dividing and non-dividing subpopulations of rat and mouse thymocytes have been labelled by using a new method of radioiodination. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and autoradiography of the labelled proteins shows distinct differences in labelling between the mouse and rat cells and also, in the case of the rat, between the dividing and non-dividing populations.

Structure and expression of two temperature-specific surface proteins in the ciliated protozoan Tetrahymena thermophila

1986 ◽  
Vol 6 (9) ◽  
pp. 3240-3245
Author(s):  
G A Bannon ◽  
R Perkins-Dameron ◽  
A Allen-Nash
Keyword(s):  
Cell Surface ◽  
Specific Surface ◽  
Incubation Temperature ◽  
Tetrahymena Thermophila ◽  
Sodium Dodecyl ◽  
Surface Protein ◽  
Surface Proteins ◽  
Ciliated Protozoan ◽  
Temperature Dependent ◽  
Molecular Sizes

The presence of specific proteins (known as immobilization antigens) on the surface of the ciliated protozoan Tetrahymena thermophila is under environmental regulation. There are five different classes (serotypes) of surface proteins which appear on the cell surface when T. thermophila is cultured under different conditions of temperature or incubation medium; three of these are temperature dependent. The appearance of these proteins on the cell surface is mutually exclusive. We used polyclonal antibodies raised against 30 degrees C (designated SerH3)- and 40 degrees C (designated SerT)-specific surface antigens to study their structure and expression. We showed that these surface proteins contain at least one disulfide bridge. On sodium dodecyl sulfate-denaturing polyacrylamide gels, the nonreduced 30 degrees C- and 40 degrees C-specific surface proteins migrated with molecular sizes of 69 and 36 kilodaltons, respectively. The reduced forms of the proteins migrated with molecular sizes of 58 and 30 kilodaltons, respectively. The synthesis of the surface proteins responded rapidly and with a time course similar to that of the incubation temperature. The synthesis of each surface protein was greatly reduced within 1 h and undetectable by 2 h after a shift to the temperature at which the protein is not expressed. Surface protein synthesis resumed by the end of 1 h after a shift to the temperature at which the protein is expressed. The temperature-dependent induction of these surface proteins appears to be dependent on the synthesis of new mRNA, as indicated by a sensitivity to actinomycin D. Surface protein syntheses were mutually exclusive except at a transition temperature. At 35 degrees C both surface proteins were synthesized by a cell population. These data support the potential of this system as a model for the study of the effects of environmental factors on the genetic regulation of cell surface proteins.

LABELLING OF THE THYROID CELL SURFACE WITH 125I

1978 ◽  
Vol 77 (3) ◽  
pp. 353-NP ◽  
Author(s):  
PAMELA M. POVEY ◽  
B. REES SMITH ◽  
R. HALL
Keyword(s):  
Cell Surface ◽  
Species Differences ◽  
Surface Membrane ◽  
Sodium Dodecyl ◽  
Thyroid Cell ◽  
Thyroid Cells ◽  
Molecular Weights ◽  
Electrophoretic Patterns ◽  
High Viability ◽  
Method Evidence

The surface membrane proteins of cultured porcine thyroid cells have been labelled with 125I by the lactoperoxidase method. Evidence that the labelling was restricted to the cell surface was supported by the high viability of the cells in suspension, the high proportion of labelled material in the particulate fraction after homogenization and electronmicroscopic autoradiographic studies. The labelled proteins were analysed by electrophoresis on polyacrylamide gels containing sodium dodecyl sulphate and this indicated the presence of ten major labelled protein bands with approximate molecular weights of 175 000, 155 000, 135 000, 88 000, 80 000, 52 300, 39 000, 30 000, 21 000 and 14 300. Comparison of the electrophoretic patterns obtained with cultured human and porcine thyroid cells suggested that there were species differences in the proportions of lower-molecular-weight proteins.

scholarly journals Cell-surface discoidin in aggregating cells of Dictyostelium discoideum

1982 ◽  
Vol 204 (3) ◽  
pp. 787-794 ◽  
Author(s):  
I C Madley ◽  
M J Cook ◽  
B D Hames
Keyword(s):  
Membrane Protein ◽  
Cell Surface ◽  
Dictyostelium Discoideum ◽  
Sodium Dodecyl Sulphate ◽  
Sodium Dodecyl ◽  
Electrophoretic Analysis ◽  
Integral Membrane Protein ◽  
Polyacrylamide Gel ◽  
Two Dimensional ◽  
Surface Receptors

Both discoidin I and discoidin II have been detected on the surface of aggregating (10 h developmental stage) cells of Dictyostelium discoideum NC4 by radioiodination of the cell-surface followed by immunoprecipitation and sodium dodecyl sulphate/polyacrylamide-gel-electrophoretic analysis. Approx. 92% of cell-surface discoidin I and 72% of cell-surface discoidin II can be eluted with 0.5 M-galactose, showing that most of each endogenous lectin is not present as integral membrane protein but rather is bound to cell-surface discoidin receptors. Two-dimensional polyacrylamide-gel-electrophoretic analysis of discoidin I suggests that the native tetramer may be a hetero-multimer composed of both Ia and Ib subunits. Cell-surface discoidin I also contains both types of subunit, but it is not clear whether both subunits have corresponding cell-surface receptors.

Isolation and partial characterization of host cell surface agglutinin and its role in attachment of a biotrophic mycoparasite

1991 ◽  
Vol 37 (5) ◽  
pp. 377-383 ◽  
Author(s):  
M. S. Manocha ◽  
Y. Chen
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Surface Proteins ◽  
Carbohydrate Binding ◽  
Protein Extract ◽  
Host Cell Wall ◽  
Host Cell Surface ◽  
Faint Band

Cell surface proteins obtained by alkaline extraction from isolated cell walls of Mortierella pusilla and M. candelabrum, host and nonhost, respectively, of the mycoparasite Piptocephalis virginiana, were tested for their ability to agglutinate mycoparasite spores. The host cell wall protein extract had a high agglutinating activity (788 agglutination units/mg) compared with that of the nonhost extract (21 agglutination units/mg). Sodium dodecyl sulfate – polyacrylamide gel electrophoresis of the crude extract of the host revealed four bands, a, b, c, and d, with respective Mr of 117 000, 100 000, 85 000 and 64 000; these bands except for a faint band c, were absent from the nonhost surface. Deletion of proteins b or c from the crude protein extract of the host significantly reduced its agglutinating activity. Proteins b and c, purified by a series of procedures, were shown to be glycoproteins with glucose and N-acetylglucosamine as major saccharides. The agglutinating activity of a mixture of pure proteins b and c was over 500 times that of either glycoprotein alone, suggesting an involvement of both glycoproteins in the agglutination process. Further characterization showed that the two glycoproteins were heat-resistant with respect to their agglutinin function, which could be totally inhibited by three sugars: arabinose, glucose and N-acetyglucosamine. It is suggested that glycoproteins b and c are the two subunits of a carbohydrate-binding agglutinin present at the host cell surface and involved in agglutination and attachment of the mycoparasite germ tubes. Key words: agglutinin, attachment, cell surface, sugars, glycoproteins, mycoparasitism.

scholarly journals Structure and expression of two temperature-specific surface proteins in the ciliated protozoan Tetrahymena thermophila.

1986 ◽  
Vol 6 (9) ◽  
pp. 3240-3245 ◽  
Author(s):  
G A Bannon ◽  
R Perkins-Dameron ◽  
A Allen-Nash
Keyword(s):  
Cell Surface ◽  
Specific Surface ◽  
Incubation Temperature ◽  
Tetrahymena Thermophila ◽  
Sodium Dodecyl ◽  
Surface Protein ◽  
Surface Proteins ◽  
Ciliated Protozoan ◽  
Temperature Dependent ◽  
Molecular Sizes

The presence of specific proteins (known as immobilization antigens) on the surface of the ciliated protozoan Tetrahymena thermophila is under environmental regulation. There are five different classes (serotypes) of surface proteins which appear on the cell surface when T. thermophila is cultured under different conditions of temperature or incubation medium; three of these are temperature dependent. The appearance of these proteins on the cell surface is mutually exclusive. We used polyclonal antibodies raised against 30 degrees C (designated SerH3)- and 40 degrees C (designated SerT)-specific surface antigens to study their structure and expression. We showed that these surface proteins contain at least one disulfide bridge. On sodium dodecyl sulfate-denaturing polyacrylamide gels, the nonreduced 30 degrees C- and 40 degrees C-specific surface proteins migrated with molecular sizes of 69 and 36 kilodaltons, respectively. The reduced forms of the proteins migrated with molecular sizes of 58 and 30 kilodaltons, respectively. The synthesis of the surface proteins responded rapidly and with a time course similar to that of the incubation temperature. The synthesis of each surface protein was greatly reduced within 1 h and undetectable by 2 h after a shift to the temperature at which the protein is not expressed. Surface protein synthesis resumed by the end of 1 h after a shift to the temperature at which the protein is expressed. The temperature-dependent induction of these surface proteins appears to be dependent on the synthesis of new mRNA, as indicated by a sensitivity to actinomycin D. Surface protein syntheses were mutually exclusive except at a transition temperature. At 35 degrees C both surface proteins were synthesized by a cell population. These data support the potential of this system as a model for the study of the effects of environmental factors on the genetic regulation of cell surface proteins.

Absence of lipophosphoglycan-like glycoconjugates in Entamoeba dispar

Parasitology ◽  
2000 ◽  
Vol 120 (1) ◽  
pp. 31-35 ◽  
Author(s):  
A. BHATTACHARYA ◽  
R. ARYA ◽  
C. G. CLARK ◽  
J. P. ACKERS
Keyword(s):  
Cell Surface ◽  
Entamoeba Histolytica ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Invasive Disease ◽  
Pathogenic Species ◽  
Human Gut ◽  
Entamoeba Dispar

Invasive amoebiasis is the result of infection of Entamoeba histolytica. The closely related Entamoeba dispar can colonize the human gut but does not cause invasive disease. In this study, E. dispar was analysed for the presence of the lipophosphoglycan-like (LPG) glycoconjugate known to be present on the cell surface of E. histolytica. E. dispar cells were radio-isotope labelled with [3H]galactose or [3H]inositol. The acidic glycoconjugates were extracted and analysed by hydrophobic chromatography over phenyl–Sepharose and by sodium dodecyl sulphate polyacrylamide gel electrophoresis. No LPG-like molecules could be identified in E. dispar in contrast to E. histolytica, suggesting that these molecules may be absent in the non-pathogenic species.

scholarly journals Incorporation of l-[3H]fucose and d-[3H]glucosamine into cell-surface-associated glycoconjugates in epidermis of cultured pig skin slices

1980 ◽  
Vol 190 (1) ◽  
pp. 65-77 ◽  
Author(s):  
I A King ◽  
A Tabiowo ◽  
R H Williams
Keyword(s):  
Molecular Weight ◽  
Cell Surface ◽  
Hydrophobic Interactions ◽  
Sodium Dodecyl ◽  
Apparent Molecular Weight ◽  
Membrane Glycoproteins ◽  
Molecular Weight Range ◽  
Electron Microscope Autoradiography ◽  
Pig Skin ◽  
Membrane Bound

1. Electron microscope autoradiography indicated that L-[3H]fucose and D-[3H]glucosamine were both incorporated into cell-surface-associated glycoconjugates in the epidermis of cultured pig skin slices. 2. Acid hydrolysis and paper chromatography of skin homogenates confirmed that there was little metabolic conversion of the labeled precursors to other sugars. 3. Epidermis was separated from dermis using CaCl2, and was extracted with 8 M-urea/5% (w/v) sodium dodecyl sulphate and was then analysed by gel electrophoresis. The major component labelled with D-[3H]glucosamine had an apparent molecular weight in excess of 200 000. This material was not labelled with L-[3H]fucose. Lower molecular-weight components were labelled to a similar extent with both L-[3H]fucose and D-[3H]glucosamine. 4. The high molecular-weight material labelled with D-[3H]glucosamine was released into the medium when the epidermal cells were dispersed with trypsin, indicating that it was either surface-associated or was extracellular. It was also labelled with D-[14C]glucuronic acid, 35SO4(2-) and to a small extent with 14C-labelled amino acids indicating that it contained glycosaminoglycans derived from epidermal proteoglycans. This was confirmed by the fact that it was degraded by testicular hyaluronoglucosidase. It was not present in isolated membranes but was recovered in the soluble fraction from epidermal homogenates. It is therefore only very loosely bound at the cell surface or is present in the extracellular spaces. 5. Membrane-bound [3H]glycoproteins were identified after differential centrifugation of epidermal homogenates. The radioactivity profiles of membrane glycoproteins were similar whether L-[3H]fucose or D-[3H]glucosamine were used and both consisted of a major heterogeneous peak in the apparent mol.wt. range 70 000–150 000. [3H]Glycoproteins in this molecular-weight range were also major components of a plasma-membrane-enriched fraction. These glycoproteins were probably bound to the membrane by hydrophobic interactions, since they were only solubilized by treatment with detergent or organic solvent. They contained terminal sialic acid residues, since they were degraded by neuraminidase.

A calcium-dependent bacterial surface protein is involved in the attachment of rhizobia to peanut roots

2003 ◽  
Vol 49 (6) ◽  
pp. 399-405 ◽  
Author(s):  
Marta Dardanelli ◽  
Jorge Angelini ◽  
Adriana Fabra
Keyword(s):  
Cell Surface ◽  
Calcium Binding ◽  
Fluorescent Protein ◽  
Sodium Dodecyl ◽  
Surface Protein ◽  
Infection Process ◽  
Surface Proteins ◽  
Phase Cell ◽  
Nodule Development ◽  
Bradyrhizobium Sp

As part of a project to characterize molecules involved in the crack-entry infection process leading to nodule development, a microscopic assay was used to visualize the attachment of cells of Bradyrhizobium sp. strains SEMIA 6144 and TAL 1000 (labelled by introducing a plasmid expressing constitutively the green fluorescent protein GFP-S65T) to Arachis hypogaea L. (peanut). Qualitative and quantitative results revealed that attachment was strongly dependent on the growth phase of the bacteria. Optimal attachment occurred when bacteria were at the late log or early stationary phase. Cell surface proteins from the Bradyrhizobium sp. strains inhibited the attachment when supplied prior to the attachment assay. Root incubation with a 14-kDa protein (eluted from sodium dodecyl sulphate – gel electrophoresis of the cell surface fraction) prior to the attachment assay resulted in a strong decrease of attachment. The adhesin appeared to be a calcium-binding protein, since cells treated with EDTA were found to be able to bind to adhesin-treated peanut roots. Since this protein has properties identical to those reported for rhicadhesin, we propose that this adhesin is also involved in the attachment process of rhizobia to root legumes that are infected by the crack-entry process.Key words: peanut, crack entry, rhizobia, attachment, adhesin.

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