Residual nitrobenzylthioinosine-resistant nucleoside transport in a transport mutant (AE1) of S49 murine T-lymphoma cells

1987 ◽  
Vol 7 (1) ◽  
pp. 160-166
Author(s):  
P G Plagemann ◽  
C Woffendin
Keyword(s):  
Mammalian Cells ◽  
Single Step ◽  
Nucleoside Transport ◽  
Wild Type ◽  
Lymphoma Cells ◽  
Defective Mutant ◽  
T Lymphoma Cells ◽  
Residual Transport ◽  
T Lymphoma ◽  
Thymidine Transport

The uptake of various nucleosides by S49 mouse T-lymphoma cells and that by a single-step nucleoside transport-defective mutant thereof (AE1) were compared. Residual nucleoside entry into AE1 cells occurred via two routes, nonmediated permeation and saturable, non-concentrative transport with broad substrate specificity and a Michaelis-Menten constant approximating that for thymidine transport in wild-type cells. However, in contrast to nucleoside transport in wild-type cells, residual nucleoside transport in AE1 cells was resistant to inhibition by nitrobenzylthioinosine. In its properties the latter resembled nitrobenzylthioinosine-resistant nucleoside transport observed in other types of mammalian cells. It amounted to less than 1% of the total nucleoside transport activity of wild-type S49 cells. The results indicate that nitrobenzylthioinosine-resistant and -sensitive nucleoside transports are genetically distinguishable. In wild-type cells, the salvage of thymidine, when present at concentrations higher than 1 to 10 microM, was limited by phosphorylation, because of the saturation of thymidine kinase. In AE1 cells, entry into the cells mainly limited thymidine salvage, but at high thymidine concentrations the combined entry via residual transport and nonmediated permeation was sufficiently rapid to support intracellular thymidine phosphorylation at rates comparable to those observed in wild-type cells.

scholarly journals Residual nitrobenzylthioinosine-resistant nucleoside transport in a transport mutant (AE1) of S49 murine T-lymphoma cells.

1987 ◽  
Vol 7 (1) ◽  
pp. 160-166 ◽  
Author(s):  
P G Plagemann ◽  
C Woffendin
Keyword(s):  
Mammalian Cells ◽  
Single Step ◽  
Nucleoside Transport ◽  
Wild Type ◽  
Lymphoma Cells ◽  
Defective Mutant ◽  
T Lymphoma Cells ◽  
Residual Transport ◽  
T Lymphoma ◽  
Thymidine Transport

The uptake of various nucleosides by S49 mouse T-lymphoma cells and that by a single-step nucleoside transport-defective mutant thereof (AE1) were compared. Residual nucleoside entry into AE1 cells occurred via two routes, nonmediated permeation and saturable, non-concentrative transport with broad substrate specificity and a Michaelis-Menten constant approximating that for thymidine transport in wild-type cells. However, in contrast to nucleoside transport in wild-type cells, residual nucleoside transport in AE1 cells was resistant to inhibition by nitrobenzylthioinosine. In its properties the latter resembled nitrobenzylthioinosine-resistant nucleoside transport observed in other types of mammalian cells. It amounted to less than 1% of the total nucleoside transport activity of wild-type S49 cells. The results indicate that nitrobenzylthioinosine-resistant and -sensitive nucleoside transports are genetically distinguishable. In wild-type cells, the salvage of thymidine, when present at concentrations higher than 1 to 10 microM, was limited by phosphorylation, because of the saturation of thymidine kinase. In AE1 cells, entry into the cells mainly limited thymidine salvage, but at high thymidine concentrations the combined entry via residual transport and nonmediated permeation was sufficiently rapid to support intracellular thymidine phosphorylation at rates comparable to those observed in wild-type cells.

Incomplete nucleoside transport deficiency with increased hypoxanthine transport capability in mutant T-lymphoblastoid cells

1986 ◽  
Vol 6 (4) ◽  
pp. 1296-1303
Author(s):  
B Aronow ◽  
P Hollingsworth ◽  
J Patrick ◽  
B Ullman
Keyword(s):  
Binding Site ◽  
Wild Type Mouse ◽  
Single Step ◽  
Nucleoside Transport ◽  
Wild Type ◽  
Surface Binding ◽  
T Lymphoma Cells ◽  
Residual Transport ◽  
T Lymphoma ◽  
Mutant Cells

From a mutagenized population of wild-type mouse (S49) T-lymphoma cells, a clone, 80-5D2, was isolated in a single step by virtue of its ability to survive in 80 nM 5-fluorouridine. Unlike previously isolated nucleoside transport-deficient cell lines (A. Cohen, B. Ullman, and D. W. Martin, Jr., J. Biol. Chem. 254:112-116, 1979), 80-5D2 cells were only slightly less sensitive to growth inhibition by a variety of cytotoxic nucleosides and were capable of proliferating in hypoxanthine-amethopterin-thymidine-containing medium. The molecular basis for the phenotype of 80-5D2 cells was incomplete deficiency in the ability of the mutant cells to translocate nucleosides across the plasma membrane. Interestingly, mutant cells were more capable than wild-type cells of transporting the nucleobase hypoxanthine. Residual transport of adenosine into 80-5D2 cells was just as sensitive to inhibition by nucleosides and more sensitive to inhibition by hypoxanthine than that in wild-type cells, indicating that the phenomena of ligand binding and translocation can be uncoupled genetically. The 80-5D2 cells lacked cell surface binding sites for the potent inhibitor of nucleoside transport p-nitrobenzylthioinosine (NBMPR) and, consequently, were largely resistant to the physiological effects of NBMPR. However, the altered transporter retained its sensitivity to dipyridamole, another inhibitor of nucleoside transport. The biochemical phenotype of the 80-5D2 cell line supports the hypothesis that the determinants that comprise the nucleoside carrier site, the hypoxanthine carrier site, the NBMPR binding site, and the dipyridamole binding site of the nucleoside transport function of mouse S49 cells are genetically distinguishable.

scholarly journals Incomplete nucleoside transport deficiency with increased hypoxanthine transport capability in mutant T-lymphoblastoid cells.

1986 ◽  
Vol 6 (4) ◽  
pp. 1296-1303 ◽  
Author(s):  
B Aronow ◽  
P Hollingsworth ◽  
J Patrick ◽  
B Ullman
Keyword(s):  
Binding Site ◽  
Wild Type Mouse ◽  
Single Step ◽  
Nucleoside Transport ◽  
Wild Type ◽  
Surface Binding ◽  
T Lymphoma Cells ◽  
Residual Transport ◽  
T Lymphoma ◽  
Mutant Cells

From a mutagenized population of wild-type mouse (S49) T-lymphoma cells, a clone, 80-5D2, was isolated in a single step by virtue of its ability to survive in 80 nM 5-fluorouridine. Unlike previously isolated nucleoside transport-deficient cell lines (A. Cohen, B. Ullman, and D. W. Martin, Jr., J. Biol. Chem. 254:112-116, 1979), 80-5D2 cells were only slightly less sensitive to growth inhibition by a variety of cytotoxic nucleosides and were capable of proliferating in hypoxanthine-amethopterin-thymidine-containing medium. The molecular basis for the phenotype of 80-5D2 cells was incomplete deficiency in the ability of the mutant cells to translocate nucleosides across the plasma membrane. Interestingly, mutant cells were more capable than wild-type cells of transporting the nucleobase hypoxanthine. Residual transport of adenosine into 80-5D2 cells was just as sensitive to inhibition by nucleosides and more sensitive to inhibition by hypoxanthine than that in wild-type cells, indicating that the phenomena of ligand binding and translocation can be uncoupled genetically. The 80-5D2 cells lacked cell surface binding sites for the potent inhibitor of nucleoside transport p-nitrobenzylthioinosine (NBMPR) and, consequently, were largely resistant to the physiological effects of NBMPR. However, the altered transporter retained its sensitivity to dipyridamole, another inhibitor of nucleoside transport. The biochemical phenotype of the 80-5D2 cell line supports the hypothesis that the determinants that comprise the nucleoside carrier site, the hypoxanthine carrier site, the NBMPR binding site, and the dipyridamole binding site of the nucleoside transport function of mouse S49 cells are genetically distinguishable.

Analysis of adenosine-mediated pyrimidine starvation using cultured wild-type and mutant mouse T-lymphoma cells

1978 ◽  
Vol 4 (2) ◽  
pp. 201-219 ◽  
Author(s):  
Lorraine J. Gudas ◽  
Amos Cohen ◽  
Buddy Ullman ◽  
David W. Martin
Keyword(s):  
Mutant Mouse ◽  
Wild Type ◽  
Lymphoma Cells ◽  
T Lymphoma Cells ◽  
T Lymphoma

scholarly journals Dipyridamole-insensitive nucleoside transport in mutant murine T lymphoma cells.

1986 ◽  
Vol 261 (31) ◽  
pp. 14467-14473 ◽  
Author(s):  
B Aronow ◽  
D Toll ◽  
J Patrick ◽  
K McCartan ◽  
B Ullman
Keyword(s):  
Nucleoside Transport ◽  
Lymphoma Cells ◽  
T Lymphoma Cells ◽  
T Lymphoma

scholarly journals Haploinsufficiency of C2GnT-I glycosyltransferase renders T lymphoma cells resistant to cell death

Blood ◽  
2006 ◽  
Vol 108 (7) ◽  
pp. 2399-2406 ◽  
Author(s):  
Paula V. Cabrera ◽  
Maho Amano ◽  
Junya Mitoma ◽  
Jessica Chan ◽  
Jonathan Said ◽  
...  
Keyword(s):  
Cell Death ◽  
Dominant Negative ◽  
Mutant Enzyme ◽  
Dominant Negative Effect ◽  
Wild Type ◽  
Lymphoma Cells ◽  
T Lymphoma Cells ◽  
Negative Effect ◽  
T Lymphoma ◽  
Altered Cell

Abstract Neoplastic T cells in mycosis fungoides (MF) are resistant to apoptotic agents, including galectin-1 that is abundant in skin. Although MF cells are typically CD7–, and thus galectin-1 resistant, CD7+ HH cells, derived from a patient with MF, were also resistant to galectin-1. HH cells demonstrate altered cell surface glycosylation, with loss of core 2 O-glycan ligands for galectin-1 created by core 2 β1,6-N-acetylglucosaminyltransferase (C2GnT-I). Loss of core 2 O-glycans on tumor cells was also seen in primary CD7+ MF lesions. Surprisingly, HH cells are heterozygous for a C2GnT-I point mutation, yet this mutation resulted in a dramatic reduction in cellular glycosyltransferase activity. Expression of wild-type C2GnT-I in human HH cells, or murine lymphoma cells that lack C2GnT-I, restored core 2 O-glycan expression and susceptibility to galectin-1, whereas mutant enzyme lacked activity and did not restore core 2 O-glycan expression or susceptibility to galectin-1. Mutant enzyme did not have a dominant negative effect by affecting dimerization or activity of wild-type enzyme; rather, C2GnT-I haploinsufficiency is sufficient for loss of core 2 O-glycan expression and galectin-1 resistance. Thus, glycosyltransferase haploinsufficiency results in altered cellular glycosylation and resistance to cell death, identifying a new survival mechanism for T-lymphoma cells.

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