Functional characterization of a recombinant sodium-dependent nucleoside transporter with selectivity for pyrimidine nucleosides (cNT1rat) by transient expression in cultured mammalian cells.
Identifieur interne : 003C88 ( Main/Exploration ); précédent : 003C87; suivant : 003C89Functional characterization of a recombinant sodium-dependent nucleoside transporter with selectivity for pyrimidine nucleosides (cNT1rat) by transient expression in cultured mammalian cells.
Auteurs : X. Fang [Canada] ; F E Parkinson ; D A Mowles ; J D Young ; C E CassSource :
- The Biochemical journal [ 0264-6021 ] ; 1996.
English descriptors
- KwdEn :
- Adenosine (metabolism), Amino Acid Sequence, Animals, Base Sequence, Biological Transport, Carrier Proteins (drug effects), Carrier Proteins (genetics), Carrier Proteins (metabolism), Cells, Cultured, Dilazep (pharmacology), Dipyridamole (pharmacology), Genes, myc, Guanosine, Haplorhini, Intestines, Kidney (cytology), Membrane Transport Proteins, Molecular Sequence Data, Nucleosides (pharmacology), Pyrimidine Nucleosides (metabolism), Rats, Recombinant Proteins (metabolism), Thioinosine (analogs & derivatives), Thioinosine (pharmacology), Transfection, Uridine (metabolism).
- MESH :
- chemical , analogs & derivatives : Thioinosine.
- chemical , drug effects : Carrier Proteins.
- chemical , genetics : Carrier Proteins.
- chemical , metabolism : Adenosine, Carrier Proteins, Pyrimidine Nucleosides, Recombinant Proteins, Uridine.
- chemical , pharmacology : Dilazep, Dipyridamole, Nucleosides, Thioinosine.
- cytology : Kidney.
- Amino Acid Sequence, Animals, Base Sequence, Biological Transport, Cells, Cultured, Genes, myc, Guanosine, Haplorhini, Intestines, Membrane Transport Proteins, Molecular Sequence Data, Rats, Transfection.
Abstract
We have demonstrated that monkey kidney (COS-1) cells have a single type of nucleoside transport process, which, because it was equilibrative, sodium-independent and could be inhibited by nitrobenzylthioinosine (NBMPR), was identified as the 'equilibrative sensitive' or 'es' transporter. Using NBMPR or dilazep to inhibit the endogenous nucleoside transport activity, we have transiently expressed a cDNA that encodes an inhibitor-insensitive, concentrative nucleoside transporter protein (cNT1rat) of rat intestine in COS-1 cells. The production of recombinant cNT1rat was examined by immunoblotting using an epitope-tagged construct and by analysis of inward fluxes of 3H-labelled nucleosides. Recombinant cNT1rat was sodium-dependent and selective for pyrimidine nucleosides, with approximately Km values of 21 microM, 12.5 microM and 15 microM for uridine, thymidine and adenosine, respectively. Although adenosine exhibited high affinity for the recombinant transporter, its Vmax value was low. A variety of anti-viral and anti-cancer nucleoside drugs inhibited cNT1rat-mediated uptake of uridine by transfected COS-1 cells although to different extents (Floxidine > Idoxuridine > Zidovudine > Zalcitabine > Cytarabine > Gemcitabine), suggesting that the concentrative pyrimidine-selective nucleoside transporters, of which cNT1rat is a representative, may play a role in cellular uptake of these drugs. The cNT1rat/COS-1 expression system is a useful tool for analysis of cNT1rat-mediated transport processes.
PubMed: 8713072
Affiliations:
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Fang</name><affiliation wicri:level="1"><nlm:affiliation>Department of Biochemistry, University of Alberta, Edmonton, Canada.</nlm:affiliation><country xml:lang="fr">Canada</country><wicri:regionArea>Department of Biochemistry, University of Alberta, Edmonton</wicri:regionArea><wicri:noRegion>Edmonton</wicri:noRegion></affiliation></author><author><name sortKey="Parkinson, F E" sort="Parkinson, F E" uniqKey="Parkinson F" first="F E" last="Parkinson">F E Parkinson</name></author><author><name sortKey="Mowles, D A" sort="Mowles, D A" uniqKey="Mowles D" first="D A" last="Mowles">D A Mowles</name></author><author><name sortKey="Young, J D" sort="Young, J D" uniqKey="Young J" first="J D" last="Young">J D Young</name></author><author><name sortKey="Cass, C E" sort="Cass, C E" uniqKey="Cass C" first="C E" last="Cass">C E Cass</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="1996">1996</date><idno type="RBID">pubmed:8713072</idno><idno type="pmid">8713072</idno><idno type="wicri:Area/PubMed/Corpus">001869</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">001869</idno><idno type="wicri:Area/PubMed/Curation">001869</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Curation">001869</idno><idno type="wicri:Area/PubMed/Checkpoint">001869</idno><idno type="wicri:explorRef" wicri:stream="Checkpoint" wicri:step="PubMed">001869</idno><idno type="wicri:Area/Ncbi/Merge">002532</idno><idno type="wicri:Area/Ncbi/Curation">002532</idno><idno type="wicri:Area/Ncbi/Checkpoint">002532</idno><idno type="wicri:doubleKey">0264-6021:1996:Fang X:functional:characterization:of</idno><idno type="wicri:Area/Main/Merge">004366</idno><idno type="wicri:Area/Main/Curation">003C88</idno><idno type="wicri:Area/Main/Exploration">003C88</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">Functional characterization of a recombinant sodium-dependent nucleoside transporter with selectivity for pyrimidine nucleosides (cNT1rat) by transient expression in cultured mammalian cells.</title><author><name sortKey="Fang, X" sort="Fang, X" uniqKey="Fang X" first="X" last="Fang">X. Fang</name><affiliation wicri:level="1"><nlm:affiliation>Department of Biochemistry, University of Alberta, Edmonton, Canada.</nlm:affiliation><country xml:lang="fr">Canada</country><wicri:regionArea>Department of Biochemistry, University of Alberta, Edmonton</wicri:regionArea><wicri:noRegion>Edmonton</wicri:noRegion></affiliation></author><author><name sortKey="Parkinson, F E" sort="Parkinson, F E" uniqKey="Parkinson F" first="F E" last="Parkinson">F E Parkinson</name></author><author><name sortKey="Mowles, D A" sort="Mowles, D A" uniqKey="Mowles D" first="D A" last="Mowles">D A Mowles</name></author><author><name sortKey="Young, J D" sort="Young, J D" uniqKey="Young J" first="J D" last="Young">J D Young</name></author><author><name sortKey="Cass, C E" sort="Cass, C E" uniqKey="Cass C" first="C E" last="Cass">C E Cass</name></author></analytic><series><title level="j">The Biochemical journal</title><idno type="ISSN">0264-6021</idno><imprint><date when="1996" type="published">1996</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Adenosine (metabolism)</term><term>Amino Acid Sequence</term><term>Animals</term><term>Base Sequence</term><term>Biological Transport</term><term>Carrier Proteins (drug effects)</term><term>Carrier Proteins (genetics)</term><term>Carrier Proteins (metabolism)</term><term>Cells, Cultured</term><term>Dilazep (pharmacology)</term><term>Dipyridamole (pharmacology)</term><term>Genes, myc</term><term>Guanosine</term><term>Haplorhini</term><term>Intestines</term><term>Kidney (cytology)</term><term>Membrane Transport Proteins</term><term>Molecular Sequence Data</term><term>Nucleosides (pharmacology)</term><term>Pyrimidine Nucleosides (metabolism)</term><term>Rats</term><term>Recombinant Proteins (metabolism)</term><term>Thioinosine (analogs & derivatives)</term><term>Thioinosine (pharmacology)</term><term>Transfection</term><term>Uridine (metabolism)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="analogs & derivatives" xml:lang="en"><term>Thioinosine</term></keywords><keywords scheme="MESH" type="chemical" qualifier="drug effects" xml:lang="en"><term>Carrier Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Carrier Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Adenosine</term><term>Carrier Proteins</term><term>Pyrimidine Nucleosides</term><term>Recombinant Proteins</term><term>Uridine</term></keywords><keywords scheme="MESH" type="chemical" qualifier="pharmacology" xml:lang="en"><term>Dilazep</term><term>Dipyridamole</term><term>Nucleosides</term><term>Thioinosine</term></keywords><keywords scheme="MESH" qualifier="cytology" xml:lang="en"><term>Kidney</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Amino Acid Sequence</term><term>Animals</term><term>Base Sequence</term><term>Biological Transport</term><term>Cells, Cultured</term><term>Genes, myc</term><term>Guanosine</term><term>Haplorhini</term><term>Intestines</term><term>Membrane Transport Proteins</term><term>Molecular Sequence Data</term><term>Rats</term><term>Transfection</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">We have demonstrated that monkey kidney (COS-1) cells have a single type of nucleoside transport process, which, because it was equilibrative, sodium-independent and could be inhibited by nitrobenzylthioinosine (NBMPR), was identified as the 'equilibrative sensitive' or 'es' transporter. Using NBMPR or dilazep to inhibit the endogenous nucleoside transport activity, we have transiently expressed a cDNA that encodes an inhibitor-insensitive, concentrative nucleoside transporter protein (cNT1rat) of rat intestine in COS-1 cells. The production of recombinant cNT1rat was examined by immunoblotting using an epitope-tagged construct and by analysis of inward fluxes of 3H-labelled nucleosides. Recombinant cNT1rat was sodium-dependent and selective for pyrimidine nucleosides, with approximately Km values of 21 microM, 12.5 microM and 15 microM for uridine, thymidine and adenosine, respectively. Although adenosine exhibited high affinity for the recombinant transporter, its Vmax value was low. A variety of anti-viral and anti-cancer nucleoside drugs inhibited cNT1rat-mediated uptake of uridine by transfected COS-1 cells although to different extents (Floxidine > Idoxuridine > Zidovudine > Zalcitabine > Cytarabine > Gemcitabine), suggesting that the concentrative pyrimidine-selective nucleoside transporters, of which cNT1rat is a representative, may play a role in cellular uptake of these drugs. The cNT1rat/COS-1 expression system is a useful tool for analysis of cNT1rat-mediated transport processes.</div></front></TEI><affiliations><list><country><li>Canada</li></country></list><tree><noCountry><name sortKey="Cass, C E" sort="Cass, C E" uniqKey="Cass C" first="C E" last="Cass">C E Cass</name><name sortKey="Mowles, D A" sort="Mowles, D A" uniqKey="Mowles D" first="D A" last="Mowles">D A Mowles</name><name sortKey="Parkinson, F E" sort="Parkinson, F E" uniqKey="Parkinson F" first="F E" last="Parkinson">F E Parkinson</name><name sortKey="Young, J D" sort="Young, J D" uniqKey="Young J" first="J D" last="Young">J D Young</name></noCountry><country name="Canada"><noRegion><name sortKey="Fang, X" sort="Fang, X" uniqKey="Fang X" first="X" last="Fang">X. Fang</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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