A short G1 phase imposes constitutive replication stress and fork remodelling in mouse embryonic stem cells

被引:135
作者
Ahuja, Akshay K. [1 ]
Jodkowska, Karolina [2 ]
Teloni, Federico [3 ]
Bizard, Anna H. [4 ,5 ]
Zellweger, Ralph [1 ]
Herrador, Raquel [1 ]
Ortega, Sagrario [6 ]
Hickson, Ian D. [4 ,5 ]
Altmeyer, Matthias [3 ]
Mendez, Juan [2 ]
Lopes, Massimo [1 ]
机构
[1] Univ Zurich, Inst Mol Canc Res, CH-8057 Zurich, Switzerland
[2] CNIO, Mol Oncol Programme, DNA Replicat Grp, E-28029 Madrid, Spain
[3] Univ Zurich, Inst Vet Biochem & Mol Biol, CH-8057 Zurich, Switzerland
[4] Univ Copenhagen, Panum Inst, Dept Cellular & Mol Med, Ctr Chromosome Stabil, DK-2200 Copenhagen N, Denmark
[5] Univ Copenhagen, Panum Inst, Ctr Hlth Aging, DK-2200 Copenhagen N, Denmark
[6] CNIO, Biotechnol Programme, Transgen Mice Core Unit, E-28029 Madrid, Spain
基金
欧洲研究理事会;
关键词
GENOMIC INSTABILITY; SELF-RENEWAL; DARK SIDE; IN-VIVO; DNA; CYCLE; PROTEIN; MAINTENANCE; PROGRESSION; ACTIVATION;
D O I
10.1038/ncomms10660
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Embryonic stem cells (ESCs) represent a transient biological state, where pluripotency is coupled with fast proliferation. ESCs display a constitutively active DNA damage response (DDR), but its molecular determinants have remained elusive. Here we show in cultured ESCs and mouse embryos that H2AX phosphorylation is dependent on Ataxia telangiectasia and Rad3 related (ATR) and is associated with chromatin loading of the ssDNA-binding proteins RPA and RAD51. Single-molecule analysis of replication intermediates reveals massive ssDNA gap accumulation, reduced fork speed and frequent fork reversal. All these marks of replication stress do not impair the mitotic process and are rapidly lost at differentiation onset. Delaying the G1/S transition in ESCs allows formation of 53BP1 nuclear bodies and suppresses ssDNA accumulation, fork slowing and reversal in the following S-phase. Genetic inactivation of fork slowing and reversal leads to chromosomal breakage in unperturbed ESCs. We propose that rapid cell cycle progression makes ESCs dependent on effective replication-coupled mechanisms to protect genome integrity.
引用
收藏
页数:11
相关论文
共 54 条
[21]   Molecular ties between the cell cycle and differentiation in embryonic stem cells [J].
Li, Victor C. ;
Kirschner, Marc W. .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2014, 111 (26) :9503-9508
[22]   Gap 1 phase length and mouse embryonic stem cell self-renewal [J].
Li, Victor C. ;
Ballabeni, Andrea ;
Kirschner, Marc W. .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2012, 109 (31) :12550-12555
[23]   53BP1 nuclear bodies form around DNA lesions generated by mitotic transmission of chromosomes under replication stress [J].
Lukas, Claudia ;
Savic, Velibor ;
Bekker-Jensen, Simon ;
Doil, Carsten ;
Neumann, Beate ;
Pedersen, Ronni Solvhoj ;
Grofte, Merete ;
Chan, Kok Lung ;
Hickson, Ian David ;
Bartek, Jiri ;
Lukas, Jiri .
NATURE CELL BIOLOGY, 2011, 13 (03) :243-U380
[24]   How unfinished business from S-phase affects mitosis and beyond [J].
Mankouri, Hocine W. ;
Huttner, Diana ;
Hickson, Ian D. .
EMBO JOURNAL, 2013, 32 (20) :2661-2671
[25]   Chromatin association of human origin recognition complex, Cdc6, and minichromosome maintenance proteins during the cell cycle:: Assembly of prereplication complexes in late mitosis [J].
Méndez, J ;
Stillman, B .
MOLECULAR AND CELLULAR BIOLOGY, 2000, 20 (22) :8602-8612
[26]   DNA Damage Responses in Human Induced Pluripotent Stem Cells and Embryonic Stem Cells [J].
Momcilovic, Olga ;
Knobloch, Leah ;
Fornsaglio, Jamie ;
Varum, Sandra ;
Easley, Charles ;
Schatten, Gerald .
PLOS ONE, 2010, 5 (10)
[27]   Replication fork reversal in eukaryotes: from dead end to dynamic response [J].
Neelsen, Kai J. ;
Lopes, Massimo .
NATURE REVIEWS MOLECULAR CELL BIOLOGY, 2015, 16 (04) :207-220
[28]   Deregulated origin licensing leads to chromosomal breaks by rereplication of a gapped DNA template [J].
Neelsen, Kai J. ;
Zanini, Isabella M. Y. ;
Mijic, Sofija ;
Herrador, Raquel ;
Zellweger, Ralph ;
Chaudhuri, Arnab Ray ;
Creavin, Kevin D. ;
Blow, J. Julian ;
Lopes, Massimo .
GENES & DEVELOPMENT, 2013, 27 (23) :2537-2542
[29]  
Neelsen KJ, 2014, METHODS MOL BIOL, V1094, P177, DOI 10.1007/978-1-62703-706-8_15
[30]   Oncogenes induce genotoxic stress by mitotic processing of unusual replication intermediates [J].
Neelsen, Kai J. ;
Zanini, Isabella M. Y. ;
Herrador, Raquel ;
Lopes, Massimo .
JOURNAL OF CELL BIOLOGY, 2013, 200 (06) :699-708