Determination of deoxyribonucleoside triphosphate pool sizes in ribonucleotide reductase cDNA transfected human KB cells

Biochem Pharmacol. 1998 May 15;55(10):1657-65. doi: 10.1016/s0006-2952(98)00042-2.


Ribonucleotide reductase (RR) is a rate-limiting enzyme in DNA synthesis, which is responsible for controlling deoxyribonucleoside triphosphate (dNTP) pool size. It has been shown that transfection of RR M2 cDNA in human KB cells (M2-D clone) results in overexpression for the M2 subunit and resistance to hydroxyurea (HU). In this study, dNTP pool assays were performed to measure the pool sizes in six cell lines: two controls, three transfectants, and drug-induced HU-resistant (HUR) cells. Total dNTP levels among the six cell lines rose in the following order: KB wild-type, KB vector-only transfectant, M1 cDNA transfectant, M2 cDNA transfectant, M1/M2 cDNA transfectant, and HU-induced resistant clone. The dCTP levels of the cells mimicked the total dNTP pools on a smaller scale. The significant increases in the dCTP pool sizes of the M2-D, X-D, and HUR clones were proportional to their respective increases in RR activity. Relative to all other transfectants, the M1-D clone demonstrated lower dCTP levels but increased dATP pools. The M1-D clone demonstrated a significant resistance to dNTP inhibition of RR activity compared with the control KB wild-type cells. In contrast, a profound inhibition of dCTP and a decreased sensitivity to dATP inhibition was observed in M2-D, X-D, and HUR clones. In summary, M2 cDNA transfectants and HUR clones had increased RR activity as well as expanded dNTP pools, particularly dCTP, when compared with wild-type KB cells. These data provide evidence for the intertwined relationship between RR activity and dNTP pools.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Clone Cells
  • DNA, Complementary
  • Deoxyribonucleotides / metabolism*
  • Drug Resistance, Neoplasm / genetics
  • Humans
  • Hydroxyurea / pharmacology
  • RNA, Messenger / genetics
  • Ribonucleotide Reductases / genetics*
  • Transfection
  • Tumor Cells, Cultured


  • DNA, Complementary
  • Deoxyribonucleotides
  • RNA, Messenger
  • Ribonucleotide Reductases
  • Hydroxyurea