Genetic analysis and functional characterization of polymorphisms in the human concentrative nucleoside transporter, CNT2.
The concentrative nucleoside transporter CNT2 (SPNT1; SLC28A2) plays a role in the absorption and disposition of naturally occurring nucleosides, as well as nucleoside analog drugs. The aim of the present study was to characterize genetic variation in SLC28A2, the gene encoding CNT2, and to functionally analyse non-synonymous variants of CNT2, as a first step towards understanding whether genetic variation in this nucleoside transporter contributes to variation in response to nucleoside analogs. As part of a larger study, DNA samples from an ethnically diverse population (100 African-Americans, 100 European-Americans, 30 Asians, 10 Mexicans and seven Pacific Islanders) were screened and 10 coding region variants of CNT2 were identified. The non-synonymous variants were then constructed and characterized in Xenopus laevis oocytes. Six non-synonymous variants were identified, and all were able to transport guanosine. The four common variants (>1% in the sample population) were further characterized with the anti-viral nucleoside analog drug ribavirin. No differences were observed among the four common variants in the uptake kinetics of 3H-ribavirin (Km in microM: 35.6+/-9.27 for CNT2-reference, 40.7+/-6.47 for CNT2-P22L, 31.2+/-15.8 for CNT2-S75R, 26.7+/-6.13 for CNT2-S245T and 49.9+/-14.6 for CNT2-F355S). The variant CNT2-F355S exhibited a change in specificity for the naturally occurring nucleosides, inosine and uridine. All non-synonymous variants of CNT2 took up guanosine, and the four variants examined showed no significant difference in ribavirin kinetics. However, CNT2-F355S (3% allele frequency in the African-American sample) was found to alter specificity for naturally occurring nucleosides, which may have implications for nucleoside homeostasis.
Owen
Ryan P
RP
Department of Biopharmaceutical Sciences, University of California, San Francisco, San Francisco, California, USA.
Gray
Jennifer H
JH
Taylor
Travis R
TR
Carlson
Elaine J
EJ
Huang
Conrad C
CC
Kawamoto
Michiko
M
Johns
Susan J
SJ
Stryke
Doug
D
Ferrin
Thomas E
TE
Giacomini
Kathleen M
KM
D000483:Alleles; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005091:Exons; D014644:Genetic Variation; D006151:Guanosine / Q000737:chemistry / Q000378:metabolism; D006239:Haplotypes; D006801:Humans; D020128:Inhibitory Concentration 50; D007288:Inosine / Q000737:chemistry; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000235:genetics*; D008957:Models, Genetic; D033703:Nucleoside Transport Proteins / Q000235:genetics; D009705:Nucleosides / Q000235:genetics; D009865:Oocytes / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011110:Polymorphism, Genetic; D012254:Ribavirin / Q000737:chemistry / Q000494:pharmacology; D012680:Sensitivity and Specificity; D014529:Uridine / Q000737:chemistry; D014982:Xenopus laevis
D000998:Antiviral Agents; D026901:Membrane Transport Proteins; D033703:Nucleoside Transport Proteins; D009705:Nucleosides; C065171:cif nucleoside transporter; D006151:Guanosine; D012254:Ribavirin; D007288:Inosine; D004247:DNA; D014529:Uridine