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Andrei Chabes

Publications and source records attributed to Andrei Chabes.

6 recordsLinked to original sources
PubMed2025

Decoding nucleoside supplementation: how thymidine outperforms ribonucleosides in accelerating mammalian replication forks.

Disruptions in deoxynucleoside triphosphate (dNTP) supply impair DNA replication and lead to genomic instability. While exogenous ribonucleosides (rNuc) have been suggested to alleviate replication stress by increasing dNTP levels, their precise metabolic effects remain unclear. Here, we show that rNuc supplementation primarily elevates CTP and UTP levels, with only modest increases in dCTP, and has minimal impact on replication fork speed across multiple mammalian cell lines. In contrast, thymidine (dThd), either alone or in combination with rNuc-as in EmbryoMax Nucleosides-significantly increases dTTP and dGTP levels, leading to accelerated replication fork progression. Notably, dThd, rather than rNuc, drives fork acceleration and counteracts fork slowdown caused by elevated dUTP, consistent with primer extension assays showing that dUTP transiently inhibits Pol ϵ-mediated DNA synthesis at template adenines. These results clarify the distinct roles of nucleosides in nucleotide metabolism, providing a mechanistic basis for how dThd promotes fork progression and preserves genomic stability.

DNA Replication↗
PubMed2007

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

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.

Alleles↗
PubMed2003

Survival of DNA damage in yeast directly depends on increased dNTP levels allowed by relaxed feedback inhibition of ribonucleotide reductase.

In eukaryotes, DNA damage elicits a multifaceted response that includes cell cycle arrest, transcriptional activation of DNA repair genes, and, in multicellular organisms, apoptosis. We demonstrate that in Saccharomyces cerevisiae, DNA damage leads to a 6- to 8-fold increase in dNTP levels. This increase is conferred by an unusual, relaxed dATP feedback inhibition of ribonucleotide reductase (RNR). Complete elimination of dATP feedback inhibition by mutation of the allosteric activity site in RNR results in 1.6-2 times higher dNTP pools under normal growth conditions, and the pools increase an additional 11- to 17-fold during DNA damage. The increase in dNTP pools dramatically improves survival following DNA damage, but at the same time leads to higher mutation rates. We propose that increased survival and mutation rates result from more efficient translesion DNA synthesis at elevated dNTP concentrations.

Adenosine Triphosphate↗
PubMed2002

Cid13 is a cytoplasmic poly(A) polymerase that regulates ribonucleotide reductase mRNA.

Fission yeast Cid13 and budding yeast Trf4/5 are members of a newly identified nucleotidyltransferase family conserved from yeast to man. Trf4/5 are thought to be essential DNA polymerases. We report that Cid13 is a poly(A) polymerase. Unlike conventional poly(A) polymerases, which act in the nucleus and indiscriminately polyadenylate all mRNA, Cid13 is a cytoplasmic enzyme that specifically targets suc22 mRNA that encodes a subunit of ribonucleotide reductase (RNR). cid13 mutants have reduced dNTP pools and are sensitive to hydroxyurea, an RNR inhibitor. We propose that Cid13 defines a cytoplasmic form of poly(A) polymerase important for DNA replication and genome maintenance.

Cell Nucleus↗
PubMed2002

Yeast DNA damage-inducible Rnr3 has a very low catalytic activity strongly stimulated after the formation of a cross-talking Rnr1/Rnr3 complex.

The ribonucleotide reductase system in Saccharomyces cerevisiae includes four genes (RNR1 and RNR3 encoding the large subunit and RNR2 and RNR4 encoding the small subunit). RNR3 expression, nearly undetectable during normal growth, is strongly induced by DNA damage. Yet an rnr3 null mutant has no obvious phenotype even under DNA damaging conditions, and the contribution of RNR3 to ribonucleotide reduction is not clear. To investigate the role of RNR3 we expressed and characterized the Rnr3 protein. The in vitro activity of Rnr3 was less than 1% of the Rnr1 activity. However, a strong synergism between Rnr3 and Rnr1 was observed, most clearly demonstrated in experiments with the catalytically inactive Rnr1-C428A mutant, which increased the endogenous activity of Rnr3 by at least 10-fold. In vivo, the levels of Rnr3 after DNA damage never reached more than one-tenth of the Rnr1 levels. We propose that heterodimerization of Rnr3 with Rnr1 facilitates the recruitment of Rnr3 to the ribonucleotide reductase holoenzyme, which may be important when Rnr1 is limiting for dNTP production. In complex with inactive Rnr1-C428A, the activity of Rnr3 is controlled by effector binding to Rnr1-C428A. This result indicates cross-talk between the Rnr1 and Rnr3 polypeptides of the large subunit.

Allosteric Regulation↗