PMID- 10353719
OWN - NLM
STAT- MEDLINE
DCOM- 19990805
LR  - 20190915
IS  - 0829-8211 (Print)
IS  - 0829-8211 (Linking)
VI  - 76
IP  - 5
DP  - 1998
TI  - Stable expression of a recombinant sodium-dependent, pyrimidine-selective
      nucleoside transporter (CNT1) in a transport-deficient mouse leukemia cell line.
PG  - 843-51
AB  - Previous studies of nucleoside transport in mammalian cells have identified two
      types of activities: the equilibrative nucleoside transporters and concentrative,
      Na+-nucleoside cotransporters. Characterization of the concentrative nucleoside
      transporters has been hampered by the presence in most cells and tissues of
      multiple transporters with overlapping permeant specificities. With the recent
      cloning of cDNAs encoding rat and human members of the concentrative nucleoside
      transporter (CNT) family, it is now possible to study the concentrative
      transporters in isolation by use of functional expression systems. We report here
      the isolation of a nucleoside transport-deficient subline of L1210 mouse leukemia
      (L1210/DNC3) that is a suitable recipient for stable expression of cloned
      nucleoside transporter cDNAs. We have used L1210/DNC3 as the recipient in gene
      transfer studies to develop a stable cell line (L1210/DU5) that produces the
      recombinant concentrative nucleoside transporter with selectivity for pyrimidine 
      nucleosides (CNT1) that was initially identified in rat intestine (Q.Q. Huang,
      S.Y. Yao, M.W. Ritzel, A.R.P. Paterson, C.E. Cass, and J.D. Young. 1994. J. Biol.
      Chem. 269: 17,757-17,760). L1210/DU5 was used to examine the permeant selectivity
      of recombinant rat CNT1 by comparing a series of nucleoside analogs with respect 
      to (i) inhibition of inward fluxes of [3H]thymidine, (ii) initial rates of
      transport of 3H-analog, and (iii) cytotoxicity to L1210/DU5 versus the parental
      transport-deficient cell line. By all three criteria, recombinant CNT1
      transported 5-fluoro-2'-deoxyuridine and 5-fluorouridine well and cytosine
      arabinoside poorly. Although some purine nucleosides
      (2'-deoxyadenosinedeoxyadeno-2'-deoxyadenosine, 7-deazaadenosine) were potent
      inhibitors of CNT1, they were poor permeants when uptake was measured directly by
      analysis of isotopic fluxes or indirectly by comparison of cytotoxicity ratios.
      We conclude that comparison of analog cytotoxicity to L1210/DU5 versus L1210/DNC3
      is a reliable indirect predictor of transportability, suggesting that
      cytotoxicity assays with a panel of such cell lines, each with a different
      recombinant nucleoside transporter, would be a valuable tool in the development
      of antiviral and antitumor nucleoside analogs.
FAU - Crawford, C R
AU  - Crawford CR
AD  - Department of Molecular Pharmacology, St. Jude Children's Research Hospital,
      Memphis, TN 38105, USA.
FAU - Cass, C E
AU  - Cass CE
FAU - Young, J D
AU  - Young JD
FAU - Belt, J A
AU  - Belt JA
LA  - eng
GR  - CA21765/CA/NCI NIH HHS/United States
GR  - CA55056/CA/NCI NIH HHS/United States
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - Canada
TA  - Biochem Cell Biol
JT  - Biochemistry and cell biology = Biochimie et biologie cellulaire
JID - 8606068
RN  - 0 (Carrier Proteins)
RN  - 0 (Membrane Transport Proteins)
RN  - 0 (Nucleosides)
RN  - 0 (Recombinant Fusion Proteins)
RN  - 0 (cif nucleoside transporter)
SB  - IM
MH  - Animals
MH  - Carrier Proteins/*chemistry/*metabolism
MH  - Inhibitory Concentration 50
MH  - Ion Transport
MH  - Leukemia/*metabolism
MH  - *Membrane Transport Proteins
MH  - Mice
MH  - Nucleosides/pharmacology
MH  - Recombinant Fusion Proteins/*metabolism
MH  - Time Factors
MH  - Transfection
MH  - Tumor Cells, Cultured
EDAT- 1999/06/03 00:00
MHDA- 1999/06/03 00:01
CRDT- 1999/06/03 00:00
PHST- 1999/06/03 00:00 [pubmed]
PHST- 1999/06/03 00:01 [medline]
PHST- 1999/06/03 00:00 [entrez]
AID - 10.1139/bcb-76-5-843 [doi]
PST - ppublish
SO  - Biochem Cell Biol. 1998;76(5):843-51. doi: 10.1139/bcb-76-5-843.