Transport of adenosine by recombinant purine- and pyrimidine-selective sodium/nucleoside cotransporters from rat jejunum expressed in Xenopus laevis oocytes.
Two major Na+-dependent nucleoside transporter subtypes implicated in adenosine transport in mammalian cells are distinguished functionally on the basis of substrate specificity: one is selective for pyrimidine nucleosides but also binds adenosine, and the other has selectivity for purine nucleosides but also binds uridine. Transportability of adenosine by the purine-selective system has been established by measurements of [3H]adenosine fluxes, whereas the conclusion that adenosine is permeant of the pyrimidine-selective system is based on inhibition assays. We investigated adenosine transport mediated by a recombinant pyrimidine-selective rat jejunal/kidney Na+/nucleoside cotransporter (rCNT1) expressed in Xenopus laevis oocytes and compared it with that mediated by a recombinant purine-selective rat jejunal/liver Na+/nucleoside cotransporter (rCNT2). Adenosine fluxes mediated by rCNT1 were 1 order of magnitude lower than those mediated by rCNT2. In kinetic studies, rCNT1 transported adenosine with an apparent Km value for influx (26 microM) similar to that for uridine but with a very much lower Vmax value, and the Vmax/Km ratios were 0.003 and 0.57 for adenosine and uridine, respectively. Recombinant rCNT1 mediated efflux of [3H]uridine from preloaded oocytes, demonstrating a capacity for bidirectional transport of nucleoside permeants. Uridine efflux was stimulated by extracellular uridine and inhibited by extracellular adenosine, suggesting that the rate of conversion of rCNT1 from its outward-facing conformation to its inward-facing conformation was increased when the transporter was complexed with uridine and decreased when it was complexed with adenosine. Thus, although rCNT1 binds adenosine and uridine with similar affinities, it kinetically favors transport of uridine.
Yao
S Y
SY
Department of Physiology, University of Alberta, Edmonton, Canada.
Ng
A M
AM
Ritzel
M W
MW
Gati
W P
WP
Cass
C E
CE
Young
J D
JD
D000241:Adenosine / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism*; D018076:DNA, Complementary; D007583:Jejunum / Q000378:metabolism*; D008969:Molecular Sequence Data; D011684:Purine Nucleosides / Q000378:metabolism*; D011741:Pyrimidine Nucleosides / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D012964:Sodium / Q000378:metabolism*; D027981:Symporters; D014982:Xenopus laevis
D002352:Carrier Proteins; D018076:DNA, Complementary; D011684:Purine Nucleosides; D011741:Pyrimidine Nucleosides; D011994:Recombinant Proteins; D027981:Symporters; C073034:sodium-nucleoside cotransporter; D012964:Sodium; D000241:Adenosine