S phase block following MEC1ATR inactivation occurs without severe dNTP depletion

Inactivation of Mec1, the budding yeast ATR, results in a permanent S phase arrest followed by chromosome breakage and cell death during G2/M. The S phase arrest is proposed to stem from a defect in Mec1-mediated degradation of Sml1, a conserved inhibitor of ribonucleotide reductase (RNR), causing a...

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Main Authors: Caroline Earp, Samuel Rowbotham, Gábor Merényi, Andrei Chabes, Rita S. Cha
Format: Article
Language:English
Published: The Company of Biologists 2015-12-01
Series:Biology Open
Subjects:
Online Access:http://bio.biologists.org/content/4/12/1739
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author Caroline Earp
Samuel Rowbotham
Gábor Merényi
Andrei Chabes
Rita S. Cha
author_facet Caroline Earp
Samuel Rowbotham
Gábor Merényi
Andrei Chabes
Rita S. Cha
author_sort Caroline Earp
collection DOAJ
description Inactivation of Mec1, the budding yeast ATR, results in a permanent S phase arrest followed by chromosome breakage and cell death during G2/M. The S phase arrest is proposed to stem from a defect in Mec1-mediated degradation of Sml1, a conserved inhibitor of ribonucleotide reductase (RNR), causing a severe depletion in cellular dNTP pools. Here, the casual link between the S phase arrest, Sml1, and dNTP-levels is examined using a temperature sensitive mec1 mutant. In addition to S phase arrest, thermal inactivation of Mec1 leads to constitutively high levels of Sml1 and an S phase arrest. Expression of a novel suppressor, GIS2, a conserved mRNA binding zinc finger protein, rescues the arrest without down-regulating Sml1 levels. The dNTP pool in mec1 is reduced by ∼17% and GIS2 expression restores it, but only partially, to ∼93% of a control. We infer that the permanent S phase block following Mec1 inactivation can be uncoupled from its role in Sml1 down-regulation. Furthermore, unexpectedly modest effects of mec1 and GIS2 on dNTP levels suggest that the S phase arrest is unlikely to result from a severe depletion of dNTP pool as assumed, but a heightened sensitivity to small changes in its availability.
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spelling doaj.art-8d91a02359ab4df6b3e3f909010b498c2022-12-21T21:58:35ZengThe Company of BiologistsBiology Open2046-63902015-12-014121739174310.1242/bio.015347015347S phase block following MEC1ATR inactivation occurs without severe dNTP depletionCaroline Earp0Samuel Rowbotham1Gábor Merényi2Andrei Chabes3Rita S. Cha4 Stem Cell Biology and Developmental Genetics, National Institute for Medical Research, MRC, London NW7 1AA, UK Stem Cell Biology and Developmental Genetics, National Institute for Medical Research, MRC, London NW7 1AA, UK Department of Medical Biochemistry and Biophysics, Umeå University, Umeå SE 901 87, Sweden Department of Medical Biochemistry and Biophysics, Umeå University, Umeå SE 901 87, Sweden Stem Cell Biology and Developmental Genetics, National Institute for Medical Research, MRC, London NW7 1AA, UK Inactivation of Mec1, the budding yeast ATR, results in a permanent S phase arrest followed by chromosome breakage and cell death during G2/M. The S phase arrest is proposed to stem from a defect in Mec1-mediated degradation of Sml1, a conserved inhibitor of ribonucleotide reductase (RNR), causing a severe depletion in cellular dNTP pools. Here, the casual link between the S phase arrest, Sml1, and dNTP-levels is examined using a temperature sensitive mec1 mutant. In addition to S phase arrest, thermal inactivation of Mec1 leads to constitutively high levels of Sml1 and an S phase arrest. Expression of a novel suppressor, GIS2, a conserved mRNA binding zinc finger protein, rescues the arrest without down-regulating Sml1 levels. The dNTP pool in mec1 is reduced by ∼17% and GIS2 expression restores it, but only partially, to ∼93% of a control. We infer that the permanent S phase block following Mec1 inactivation can be uncoupled from its role in Sml1 down-regulation. Furthermore, unexpectedly modest effects of mec1 and GIS2 on dNTP levels suggest that the S phase arrest is unlikely to result from a severe depletion of dNTP pool as assumed, but a heightened sensitivity to small changes in its availability.http://bio.biologists.org/content/4/12/1739Mec1ATRSml1RNRdNTPDNA replicationReplication arrest
spellingShingle Caroline Earp
Samuel Rowbotham
Gábor Merényi
Andrei Chabes
Rita S. Cha
S phase block following MEC1ATR inactivation occurs without severe dNTP depletion
Biology Open
Mec1
ATR
Sml1
RNR
dNTP
DNA replication
Replication arrest
title S phase block following MEC1ATR inactivation occurs without severe dNTP depletion
title_full S phase block following MEC1ATR inactivation occurs without severe dNTP depletion
title_fullStr S phase block following MEC1ATR inactivation occurs without severe dNTP depletion
title_full_unstemmed S phase block following MEC1ATR inactivation occurs without severe dNTP depletion
title_short S phase block following MEC1ATR inactivation occurs without severe dNTP depletion
title_sort s phase block following mec1atr inactivation occurs without severe dntp depletion
topic Mec1
ATR
Sml1
RNR
dNTP
DNA replication
Replication arrest
url http://bio.biologists.org/content/4/12/1739
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AT samuelrowbotham sphaseblockfollowingmec1atrinactivationoccurswithoutseveredntpdepletion
AT gabormerenyi sphaseblockfollowingmec1atrinactivationoccurswithoutseveredntpdepletion
AT andreichabes sphaseblockfollowingmec1atrinactivationoccurswithoutseveredntpdepletion
AT ritascha sphaseblockfollowingmec1atrinactivationoccurswithoutseveredntpdepletion