Modulation of the equilibrative nucleoside transporter by inhibitors of DNA synthesis

• Published in: British journal of cancer Vol. 72; no. 4; pp. 939 - 942
• Main Authors: PRESSACCO, J, WILEY, J. S, JAMIESON, G. P, ERLICHMAN, C, HEDLEY, D. W
• Format: Journal Article
• Language: English
• Published: Basingstoke Nature Publishing Group 01.10.1995
• Subjects: Antimetabolites - pharmacology; Biological and medical sciences; Carrier Proteins - analysis; Carrier Proteins - drug effects; Cytarabine - pharmacology; DNA - biosynthesis; Fluoresceins; Fluorouracil - pharmacology; Human viral diseases; Humans; Infectious diseases; Medical sciences; Membrane Proteins - analysis; Membrane Proteins - drug effects; Nucleoside Transport Proteins; Purine Nucleosides; Thymine Nucleotides - metabolism; Tumor Cells, Cultured; Viral diseases; Viral diseases of the genital and urinary system; Human; Exploration; Papovaviridae; Uterine cervix; Transitory; Female genital diseases; Infection; Papillomavirus; Virus; Polymerase chain reaction; Human papillomavirus; Viral disease; Adolescent; Young adult; Diagnosis; Uterine cervix diseases
• ISSN: 0007-0920; 1532-1827
• DOI: 10.1038/bjc.1995.437

Expression of the equilibrative, S-(p-nitrobenzyl)-6-thioinosine (NBMPR)-sensitive nucleoside transporter (es), a component of the nucleoside salvage pathway, was measured during unperturbed growth and following exposure to various antimetabolites at growth-inhibitory concentrations. The probe 5-(SAENTA-x8)-fluorescein is a highly modified form of adenosine incorporating a fluorescein molecule. It binds. with high affinity and specificity to the (es) nucleoside transporter at a 1:1 stoichiometry, allowing reliable estimates of es expression by flow cytometry. Using a dual labelling technique which combined the vital DNA dye Hoechst-33342 and 5-(SAENTA-x8)-fluorescein, we found that surface expression of es approximately doubled between G1 and G2 + M phases of the cell cycle. To address the question of whether es expression could be modulated in cells exposed to drugs which inhibit de novo synthesis of nucleotides, cells were exposed to antimetabolite drugs having different modes of action. Hydroxyurea and 5-fluorouracil (5-FU), which inhibit the de novo synthesis of DNA precursors, produced increases in the expression of es. In contrast, cytosine arabinoside (ara-C) and aphidicolin, which directly inhibit DNA synthesis, produced no significant increase in es expression. Thymidine (TdR), which is an allosteric inhibitor of ribonucleotide reductase that depletes dATP, dCTP and dGTP pools while repleting the dTTP pool, had no significant effect on es expression. These data suggest that surface expression of the es nucleoside transporter is regulated by a mechanism which is sensitive to the supply of deoxynucleotides. Because 5-FU (which specifically depletes dTTP pools) causes a large increase in expression whereas TdR (which depletes all precursors except dTTP) does not, this mechanism might be particularly sensitive to dTTP pools.