CUL2LRR1, TRAIP and p97 control CMG helicase disassembly in the mammalian cell cycle

Fabrizio Villa; Ryo Fujisawa; Johanna Ainsworth; Kohei Nishimura; Michael Lie-A-Ling; Georges Lacaud; Karim Labib

doi:10.15252/embr.202052164

CUL2^LRR1, TRAIP and p97 control CMG helicase disassembly in the mammalian cell cycle

Fabrizio Villa, Ryo Fujisawa, Johanna Ainsworth, Kohei Nishimura, Michael Lie-A-Ling, Georges Lacaud, Karim Labib (Lead / Corresponding author)

MRC PPU

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Abstract

The eukaryotic replisome is disassembled in each cell cycle, dependent upon ubiquitylation of the CMG helicase. Studies of Saccharomyces cerevisiae, Caenorhabditis elegans and Xenopus laevis have revealed surprising evolutionary diversity in the ubiquitin ligases that control CMG ubiquitylation, but regulated disassembly of the mammalian replisome has yet to be explored. Here, we describe a model system for studying the ubiquitylation and chromatin extraction of the mammalian CMG replisome, based on mouse embryonic stem cells. We show that the ubiquitin ligase CUL2LRR1 is required for ubiquitylation of the CMG‐MCM7 subunit during S‐phase, leading to disassembly by the p97 ATPase. Moreover, a second pathway of CMG disassembly is activated during mitosis, dependent upon the TRAIP ubiquitin ligase that is mutated in primordial dwarfism and mis‐regulated in various cancers. These findings indicate that replisome disassembly in diverse metazoa is regulated by a conserved pair of ubiquitin ligases, distinct from those present in other eukaryotes.

Original language	English
Article number	e52164
Number of pages	13
Journal	EMBO Reports
Volume	22
Issue number	3
Early online date	15 Feb 2021
DOIs	https://doi.org/10.15252/embr.202052164
Publication status	Published - 3 Mar 2021

Keywords

CMG helicase
CUL2
DNA replication
TRAIP
p97 ATPase
Molecular Biology
Genetics
Biochemistry

ASJC Scopus subject areas

Genetics
Molecular Biology
Biochemistry

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.15252/embr.202052164Licence: CC BY

Final Published VersionFinal published version, 4.4 MBLicence: CC BY

Cite this

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title = "CUL2LRR1, TRAIP and p97 control CMG helicase disassembly in the mammalian cell cycle",

abstract = "The eukaryotic replisome is disassembled in each cell cycle, dependent upon ubiquitylation of the CMG helicase. Studies of Saccharomyces cerevisiae, Caenorhabditis elegans and Xenopus laevis have revealed surprising evolutionary diversity in the ubiquitin ligases that control CMG ubiquitylation, but regulated disassembly of the mammalian replisome has yet to be explored. Here, we describe a model system for studying the ubiquitylation and chromatin extraction of the mammalian CMG replisome, based on mouse embryonic stem cells. We show that the ubiquitin ligase CUL2LRR1 is required for ubiquitylation of the CMG‐MCM7 subunit during S‐phase, leading to disassembly by the p97 ATPase. Moreover, a second pathway of CMG disassembly is activated during mitosis, dependent upon the TRAIP ubiquitin ligase that is mutated in primordial dwarfism and mis‐regulated in various cancers. These findings indicate that replisome disassembly in diverse metazoa is regulated by a conserved pair of ubiquitin ligases, distinct from those present in other eukaryotes.",

keywords = "CMG helicase, CUL2, DNA replication, TRAIP, p97 ATPase, Molecular Biology, Genetics, Biochemistry",

author = "Fabrizio Villa and Ryo Fujisawa and Johanna Ainsworth and Kohei Nishimura and Michael Lie-A-Ling and Georges Lacaud and Karim Labib",

note = "Funding Information: We thank Remi Sonneville, Iain Porter and Paul Appleton for invaluable assistance with spinning disk confocal microscopy, Masato Kanemaki for a plasmid expressing mCherry‐PCNA and MRC PPU Reagents and Services ( https://mrcppureagents.dundee.ac.uk ) for antibody production. We are very grateful for financial support from the Medical Research Council (core grant MC_UU_12016/13 to KL), the Wellcome Trust (reference 102943/Z/13/Z for an Investigator award to KL), Cancer Research UK (Programme Grant C578/A24558 and PhD studentship C578/A25669 to KL; Core Funding C5759/A20971 to GL), Biotechnology and Biological Sciences Research Council (reference BB/I001794/1/BB for award to GL), the Japan Society for the Promotion of Science (postdoctoral fellowship and “Kakenhi” grants JP19K06611 and JP20K21423 to KN) and the Naito Foundation (postdoctoral fellowship to KN). Publisher Copyright: {\textcopyright} 2021 The Authors. Published under the terms of the CC BY 4.0 license",

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doi = "10.15252/embr.202052164",

language = "English",

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AU - Villa, Fabrizio

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AU - Ainsworth, Johanna

AU - Nishimura, Kohei

AU - Lie-A-Ling, Michael

AU - Lacaud, Georges

AU - Labib, Karim

N1 - Funding Information: We thank Remi Sonneville, Iain Porter and Paul Appleton for invaluable assistance with spinning disk confocal microscopy, Masato Kanemaki for a plasmid expressing mCherry‐PCNA and MRC PPU Reagents and Services ( https://mrcppureagents.dundee.ac.uk ) for antibody production. We are very grateful for financial support from the Medical Research Council (core grant MC_UU_12016/13 to KL), the Wellcome Trust (reference 102943/Z/13/Z for an Investigator award to KL), Cancer Research UK (Programme Grant C578/A24558 and PhD studentship C578/A25669 to KL; Core Funding C5759/A20971 to GL), Biotechnology and Biological Sciences Research Council (reference BB/I001794/1/BB for award to GL), the Japan Society for the Promotion of Science (postdoctoral fellowship and “Kakenhi” grants JP19K06611 and JP20K21423 to KN) and the Naito Foundation (postdoctoral fellowship to KN). Publisher Copyright: © 2021 The Authors. Published under the terms of the CC BY 4.0 license

PY - 2021/3/3

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N2 - The eukaryotic replisome is disassembled in each cell cycle, dependent upon ubiquitylation of the CMG helicase. Studies of Saccharomyces cerevisiae, Caenorhabditis elegans and Xenopus laevis have revealed surprising evolutionary diversity in the ubiquitin ligases that control CMG ubiquitylation, but regulated disassembly of the mammalian replisome has yet to be explored. Here, we describe a model system for studying the ubiquitylation and chromatin extraction of the mammalian CMG replisome, based on mouse embryonic stem cells. We show that the ubiquitin ligase CUL2LRR1 is required for ubiquitylation of the CMG‐MCM7 subunit during S‐phase, leading to disassembly by the p97 ATPase. Moreover, a second pathway of CMG disassembly is activated during mitosis, dependent upon the TRAIP ubiquitin ligase that is mutated in primordial dwarfism and mis‐regulated in various cancers. These findings indicate that replisome disassembly in diverse metazoa is regulated by a conserved pair of ubiquitin ligases, distinct from those present in other eukaryotes.

AB - The eukaryotic replisome is disassembled in each cell cycle, dependent upon ubiquitylation of the CMG helicase. Studies of Saccharomyces cerevisiae, Caenorhabditis elegans and Xenopus laevis have revealed surprising evolutionary diversity in the ubiquitin ligases that control CMG ubiquitylation, but regulated disassembly of the mammalian replisome has yet to be explored. Here, we describe a model system for studying the ubiquitylation and chromatin extraction of the mammalian CMG replisome, based on mouse embryonic stem cells. We show that the ubiquitin ligase CUL2LRR1 is required for ubiquitylation of the CMG‐MCM7 subunit during S‐phase, leading to disassembly by the p97 ATPase. Moreover, a second pathway of CMG disassembly is activated during mitosis, dependent upon the TRAIP ubiquitin ligase that is mutated in primordial dwarfism and mis‐regulated in various cancers. These findings indicate that replisome disassembly in diverse metazoa is regulated by a conserved pair of ubiquitin ligases, distinct from those present in other eukaryotes.

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KW - DNA replication

KW - TRAIP

KW - p97 ATPase

KW - Molecular Biology

KW - Genetics

KW - Biochemistry

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SN - 1469-221X

VL - 22

JO - EMBO Reports

JF - EMBO Reports

IS - 3

M1 - e52164

ER -

CUL2^LRR1, TRAIP and p97 control CMG helicase disassembly in the mammalian cell cycle

Abstract

Keywords

ASJC Scopus subject areas

UN SDGs

Access to Document

Other files and links

Fingerprint

Novel Genome Integrity Pathways that Regulate DNA Replication Termination in Metazoa

Functional Dissection of the Eukaryotic Replisome (Senior Investigator Award)

Exploring and Exploiting C. Elegans as a Model to Better Understand Apoptosis Induction; Combined Cytological, Genetic and Proteomics Based Approaches

Cite this

CUL2LRR1, TRAIP and p97 control CMG helicase disassembly in the mammalian cell cycle

Abstract

Keywords

ASJC Scopus subject areas

UN SDGs

Access to Document

Other files and links

Fingerprint

Projects

Novel Genome Integrity Pathways that Regulate DNA Replication Termination in Metazoa

Functional Dissection of the Eukaryotic Replisome (Senior Investigator Award)

Exploring and Exploiting C. Elegans as a Model to Better Understand Apoptosis Induction; Combined Cytological, Genetic and Proteomics Based Approaches

Cite this

CUL2^LRR1, TRAIP and p97 control CMG helicase disassembly in the mammalian cell cycle