Initial rate kinetics and evidence for duality of mediated transport of adenosine, related purine nucleosides, and nucleoside analogues in L1210 cells.
In studies using a rapid kinetic technique, evidence was derived for multiplicity of systems mediating [3H]adenosine transport in L1210 cells. A variety of approaches were used in discriminating between transport and kinase-mediated phosphorylation. Under these conditions, two systems mediating influx were delineated which exhibited high-affinity [Km = 13.9 +/- 2 (S.E.) microM] or low-affinity [Km = 199 +/- 27 microM] for [3H]-adenosine. Both systems exhibited high capacities, but that associated with the low-affinity system (V 37 degrees max = 263 +/- 43 nmol = 99.6 +/- 12 nmol sec/g, dry weight). The relative difference in affinity of these two systems during influx was also reflected in the values for influx Ki obtained with other nucleosides and nucleoside analogues. Influx of [3H]-adenosine by each mediated system was inhibited by 6-(2-hydroxy-5-nitrobenzyl)thioguanosine, a specific transport inhibitor, and by 9-beta-D-arabinofuranosylpurine-6(1H)thione which is not phosphorylated in L1210 cells. Influx kinetics were the same in L1210 cells, in adenosine triphosphate-depleted L1210 cells (L1210/ara-C/MMPR) which have substantially reduced ability for [3H]adenosine phosphorylation, and in the presence of 2'-deoxycoformycin, a potent inhibitor of adenosine deaminase. The same multiplicity in mediated influx of [3H]adenosine was shown at 0 degrees when transport became rate limiting to total uptake. The high-affinity system mediating [3H]adenosine influx was also elucidated in L1210 cell plasma membrane vesicles in the presence or absence of 2'-deoxycoformycin. Almost all of the natural nucleosides examined competed less effectively with [3H]adenosine for influx by the high-affinity system than by the low-affinity system. These results are discussed with respect to possible pharmacological implications.
D000241:Adenosine / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism; D007700:Kinetics; D007939:Leukemia L1210 / Q000378:metabolism*; D051379:Mice; D011684:Purine Nucleosides / Q000378:metabolism*; D013696:Temperature; D013997:Time Factors; D014740:Vidarabine / Q000031:analogs & derivatives / Q000378:metabolism
D011684:Purine Nucleosides; D014740:Vidarabine; D000241:Adenosine; C024352:fludarabine