Hydroxyurea arrests DNA replication by a mechanism that preserves basal dNTP pools.
The relationship between dNTP levels and DNA synthesis was investigated using alpha factor-synchronized yeast treated with the ribonucleotide reductase inhibitor hydroxyurea (HU). Although HU blocked DNA synthesis and prevented the dNTP pool expansion that normally occurs at G1/S, it did not exhaust the levels of any of the four dNTPs, which dropped to about 80% of G1 levels. When dbf4 yeast that are ts for replication initiation were allowed to preaccumulate dNTPs at 37 degrees C before being released to 25 degrees C in the presence of HU, they synthesized 0.3 genome equivalents of DNA and then arrested as dNTPs approached sub-G1 levels. Accumulation of dNTPs at G1/S was not a prerequisite for replication initiation, since dbf4 cells incubated in HU at 25 degrees C were able to replicate when subsequently switched to 37 degrees C in the absence of HU. The replication arrest mechanism was not dependent on the Mec1/Rad53 pathway, since checkpoint-deficient rad53 cells also failed to exhaust basal dNTPs when incubated in HU. The persistence of basal dNTP levels in HU-arrested cells and partial bypass of the arrest in cells that had preaccumulated dNTPs suggest that cells have a mechanism for arresting DNA chain elongation when dNTP levels are not maintained above a critical threshold.
Koç
Ahmet
A
Department of Biochemistry and Biophysics, Oregon State University, Corvallis 97331, USA.
Wheeler
Linda J
LJ
Mathews
Christopher K
CK
Merrill
Gary F
GF
D004261:DNA Replication / Q000187:drug effects*; D004271:DNA, Fungal / Q000187:drug effects / Q000502:physiology; D003854:Deoxyribonucleotides / Q000378:metabolism*; D016193:G1 Phase; D005838:Genotype; D006918:Hydroxyurea / Q000494:pharmacology*; D007700:Kinetics; D016196:S Phase; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000187:drug effects / Q000235:genetics*
D004271:DNA, Fungal; D003854:Deoxyribonucleotides; D006918:Hydroxyurea