Impact of the reduced folate carrier on the accumulation of active thiamin metabolites in murine leukemia cells.
The thiamin transporter encoded by SLC19A2 and the reduced folate carrier (RFC1) share 40% homology at the protein level, but the thiamin transporter does not mediate transport of folates. By using murine leukemia cell lines that express no, normal, or high levels of RFC1, we demonstrate that RFC1 does not mediate thiamin influx. However, high level RFC1 expression substantially reduced accumulation of the active thiamin coenzyme, thiamin pyrophosphate (TPP). This decreased level of TPP, synthesized intracellularly from imported thiamin, resulted from RFC1-mediated efflux of TPP. This conclusion was supported by the following observations. (i) Efflux of intracellular TPP was increased in cells with high expression of RFC1. (ii) Methotrexate inhibits TPP influx. (iii) TPP competitively inhibits methotrexate influx. (iv) Loading cells, which overexpress RFC1 to high levels of methotrexate to inhibit competitively RFC1-mediated TPP efflux, augment TPP accumulation. (v) There was an inverse correlation between thiamin accumulation and RFC1 activity in cells grown at a physiological concentration of thiamin. The modulation of thiamin accumulation by RFC1 in murine leukemia cells suggests that this carrier may play a role in thiamin homeostasis and could serve as a modifying factor in thiamin nutritional deficiency as well as when the high affinity thiamin transporter is mutated.
Zhao
R
R
Department of Medicine and Molecular Pharmacology, and the Albert Einstein Comprehensive Cancer Center, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Gao
F
F
Wang
Y
Y
Diaz
G A
GA
Gelb
B D
BD
Goldman
I D
ID
D000818:Animals; D001692:Biological Transport; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D007939:Leukemia L1210 / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins; D008727:Methotrexate / Q000493:pharmacokinetics*; D051379:Mice; D016296:Mutagenesis; D011994:Recombinant Proteins / Q000378:metabolism; D013831:Thiamine / Q000031:analogs & derivatives / Q000378:metabolism*; D013835:Thiamine Pyrophosphate / Q000378:metabolism*; D014162:Transfection; D014407:Tumor Cells, Cultured
D002352:Carrier Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D011994:Recombinant Proteins; C120043:SLC19A2 protein, human; C492292:Slc19a2 protein, mouse; D013835:Thiamine Pyrophosphate; D013831:Thiamine; D008727:Methotrexate