Metabolic expression of thiol-derivatized sialic acids on the cell surface and their quantitative estimation by flow cytometry

2006 ◽  
Vol 1 (4) ◽  
pp. 1840-1851 ◽  
Author(s):  
Srinivasa-Gopalan Sampathkumar ◽  
Mark B Jones ◽  
Kevin J Yarema
2020 ◽  
Vol 9 (6) ◽  
pp. 1816
Author(s):  
Philip Rosenstock ◽  
Kaya Bork ◽  
Chiara Massa ◽  
Philipp Selke ◽  
Barbara Seliger ◽  
...  

Sialic acids are terminal sugars on the cell surface that are found on all cell types including immune cells like natural killer (NK) cells. The attachment of sialic acids to different glycan structures is catalyzed by sialyltransferases in the Golgi. However, the expression pattern of sialyltransferases in NK cells and their expression after activation has not yet been analyzed. Therefore, the present study determines which sialyltransferases are expressed in human NK cells and if activation with IL-2 changes the sialylation of NK cells. The expression of sialyltransferases was analyzed in the three human NK cell lines NK-92, NKL, KHYG-1 and primary NK cells. NK-92 cells were cultured in the absence or presence of IL-2, and changes in the sialyltransferase expression were measured by qPCR. Furthermore, specific sialylation was investigated by flow cytometry. In addition, polySia and NCAM were measured by Western blot analyses. IL-2 leads to a reduced expression of ST8SIA1, ST6GAL1 and ST3GAL1. α-2,3-Sialylation remained unchanged, while α-2,6-sialylation was increased after IL-2 stimulation. Moreover, an increase in the amount of NCAM and polySia was observed in IL-2-activated NK cells, whereas GD3 ganglioside was decreased. In this study, all sialyltransferases that were expressed in NK cells could be identified. IL-2 regulates the expression of some sialyltransferases and leads to changes in the sialylation of NK cells.


1990 ◽  
Vol 36 (3) ◽  
pp. 183-192 ◽  
Author(s):  
A. R. Hardham ◽  
E. Suzaki

Glycoconjugates on the surface of zoospores and cysts of the pathogenic fungus Phytophthora cinnamomi have been studied using fluorescein isothiocyanate labelled lectins for fluorescence microscopy and flow cytometry, and ferritin- and gold-labelled lectins for ultrastructural analysis. Of the five lectins used, only concanavalin A (ConA) binds to the surface of the zoospores, including the flagella and water expulsion vacuole. This suggests that of accessible saccharides, glucosyl or mannosyl residues predominate on the outer surface of the zoospore plasma membrane. Early in encystment, a system of flat disc-like cisternae, which underlie the zoospore plasma membrane, vesiculate. These and other small peripheral vesicles quickly disappear. After the induction of encystment, ConA is no longer localised close to the plasma membrane but binds to material loosely associated with the cell surface. Quantitative measurements by flow cytometry indicate that the ConA-binding material is gradually lost from the cell surface. The cyst wall is weakly labelled, but the site of germ tube emergence stains intensely. During the first 2 min after the induction of encystment, material that binds soybean agglutinin, Helix pommatia agglutinin, and peanut agglutinin appears on the surface of the fungal cells. The distribution of this material, rich in galactosyl or N-acetyl-D-galactosaminosyl residues, is initially patchy, but by 5 min the material evenly coats most of the cell surface. Labelling of zoospores in which intracellular sites are accessible indicates that the soybean agglutinin binding material is stored in vesicles that lie beneath the plasma membrane. Quantitation of soybean agglutinin labelling shows that maximum binding occurs 2–3 min after the induction of encystment. Key words: cell surface, flow cytometry, lectins, pathogenic fungi, Phytophthora cinnamomi.


2016 ◽  
Vol 61 ◽  
pp. S183
Author(s):  
E. Shestakova ◽  
E. Dudko ◽  
A. Grishanina ◽  
V. Kirsanov ◽  
N. Vichljantzeva ◽  
...  

1986 ◽  
Vol 24 (2) ◽  
pp. 145-154 ◽  
Author(s):  
E.T. Souza ◽  
F.C. Silva-Filho ◽  
W. De Souza ◽  
C.S. Alviano ◽  
J. Angluster ◽  
...  
Keyword(s):  

2011 ◽  
Vol 134 (2) ◽  
pp. 113-121 ◽  
Author(s):  
Laia Llinàs ◽  
Adriana Lázaro ◽  
Jose de Salort ◽  
Jessica Matesanz-Isabel ◽  
Jordi Sintes ◽  
...  

1995 ◽  
Vol 182 (6) ◽  
pp. 1997-2006 ◽  
Author(s):  
H Kishimoto ◽  
R T Kubo ◽  
H Yorifuji ◽  
T Nakayama ◽  
Y Asano ◽  
...  

Recent studies indicate that there may be functional uncoupling of the TCR-CD3 complex and suggest that the TCR-CD3 complex is composed of two parallel signal-transducing units, one made of gamma delta epsilon chains and the other of zeta chains. To elucidate the molecular mechanisms that may explain the functional uncoupling of TCR and CD3, we have analyzed their expression by using flow cytometry as well as immunochemical means both before and after stimulation with anti-TCR-beta, anti-CD3 epsilon, anti-CD2, staphylococcal enterotoxin B, and ionomycin. We present evidence that TCR physically dissociates from CD3 after stimulation of the TCR-CD3 complex. Stimulation with anti-CD3 resulted in down-modulation of TCR within 45 min whereas CD3 epsilon was still expressed on the cell surface as detected by flow cytometry. However, the cell surface expression of TCR and CD3 was not affected when cells were stimulated with anti-TCR-beta under the same conditions. In the case of anti-CD3 treatment of T cells, the TCR down-modulation appeared to be due to the internalization of TCR, as determined by immunoelectron microscopy. Immunochemical analysis of cells after stimulation with either anti-TCR or anti-CD3 mAbs revealed that the overall protein levels of TCR and CD3 were similar. More interestingly, the dissociation of the TCR-CD3 complex was observed with both treatments and occurred in a manner that the TCR and the associated TCR-zeta chain dissociated as a unit from CD3. These results provide the first report of physical dissociation of TCR and CD3 after stimulation through the TCR-CD3 complex. The results also suggest that the signal transduction pathway triggered by TCR may differ from that induced by CD3.


Author(s):  
Benoîte Bourdin ◽  
Emilie Segura ◽  
Marie-Philippe Tétreault ◽  
Sylvie Lesage ◽  
Lucie Parent

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