Functional production of mammalian concentrative nucleoside transporters in Saccharomyces cerevisiae.
The transport of nucleosides and nucleobases in the yeast Saccharomyces cerevisiae is reviewed and the use of this organism to study recombinant mammalian concentrative nucleoside transport (CNT) proteins is described. A selection strategy based on the ability of an expressed nucleoside transporter cDNA to mediate thymidine uptake by yeast under a selective condition that depletes endogenous thymidylate was used to assess the transport capacity of heterologous transporter proteins. The pyrimidine-nucleoside selective concentrative transporters from human (hCNT1) and rat (rCNT1) complemented the imposed thymidylate depletion in S. cerevisiae, as did N-terminally truncated versions of hCNT1 and rCNT1 lacking up to 31 amino acids. Transporter-mediated rescue of S. cerevisiae by both nucleoside transporters was inhibited by cytidine, uridine and adenosine, but not by guanosine or inosine. This work represents the development of a new model system for the functional production of recombinant nucleoside transporters of the CNT family of membrane proteins.
Vickers
M F
MF
University of Alberta, and Cross Cancer Institute, Edmonton, Canada.
Young
J D
JD
Baldwin
S A
SA
Ellison
M J
MJ
Cass
C E
CE
D000818:Animals; D001692:Biological Transport; D018076:DNA, Complementary / Q000378:metabolism; D005816:Genetic Complementation Test; D006801:Humans; D026901:Membrane Transport Proteins / Q000378:metabolism*; D033702:Nucleobase Transport Proteins; D033703:Nucleoside Transport Proteins; D009705:Nucleosides / Q000378:metabolism*; D033704:Nucleotide Transport Proteins; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism
D018076:DNA, Complementary; C043181:FCY2 protein, S cerevisiae; C413444:FUI1 protein, S cerevisiae; C035528:FUR4 protein, S cerevisiae; D026901:Membrane Transport Proteins; D033702:Nucleobase Transport Proteins; D033703:Nucleoside Transport Proteins; D009705:Nucleosides; D033704:Nucleotide Transport Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C065171:cif nucleoside transporter