Constitutively high dNTP concentration inhibits cell cycle progression and the DNA damage checkpoint in yeast Saccharomyces cerevisiae

doi:10.1073/pnas.0610585104

Constitutively high dNTP concentration inhibits cell cycle progression and the DNA damage checkpoint in yeast Saccharomyces cerevisiae

Andrei Chabes*,^†,^‡ and
Bruce Stillman*,^‡

*Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724; and
^†Department of Medical Biochemistry and Biophysics, Umeå University, SE-901 87 Umeå, Sweden

Contributed by Bruce Stillman, November 29, 2006 (received for review November 2, 2006)

Abstract

In eukaryotic cells the concentration of dNTP is highest in S phase and lowest in G₁ phase and is controlled by ribonucleotide reductase (RNR). RNR activity is eliminated in all eukaryotes in G₁ phase by a variety of mechanisms: transcriptional regulation, small inhibitory proteins, and protein degradation. After activation of RNR upon commitment to S phase, dATP feedback inhibition ensures that the dNTP concentration does not exceed a certain maximal level. It is not apparent why limitation of dNTP concentration is necessary in G₁ phase. In principle, dATP feedback inhibition should be sufficient to couple dNTP production to utilization. We demonstrate that in Saccharomyces cerevisiae constitutively high dNTP concentration transiently arrests cell cycle progression in late G₁ phase, affects activation of origins of replication, and inhibits the DNA damage checkpoint. We propose that fluctuation of dNTP concentration controls cell cycle progression and the initiation of DNA replication.

Footnotes

^‡To whom correspondence may be addressed. andrei.chabes{at}medchem.umu.se or stillman{at}cshl.edu
Author contributions: A.C. and B.S. designed research; A.C. performed research; A.C. and B.S. contributed new reagents/analytic tools; A.C. and B.S. analyzed data; and A.C. and B.S. wrote the paper.
The authors declare no conflict of interest.
This article contains supporting information online at www.pnas.org/cgi/content/full/0610585104/DC1.
Abbreviations:

RNR,

ribonucleotide reductase;

pre-RC,

prereplicative complex;

gal,

galactose;

MMS,

methyl methanesulfonate;

pre-IC,

preinitiation complex;

TMPK,

thymidylate kinase;

YPD,

1% yeast extract/2% peptone/2% dextrose;

YPGal,

1% yeast extract/2% peptone/2% gal;

YPRaf,

1% yeast extract/2% peptone/2% raffinose.