Ufd1-Npl4 recruit Cdc48 for disassembly of ubiquitylated CMG helicase at the end of chromosome replication

Marija Maric; Progya Mukherjee; Michael H. Tatham; Ronald Hay; Karim Labib

doi:10.1016/j.celrep.2017.03.020

Ufd1-Npl4 recruit Cdc48 for disassembly of ubiquitylated CMG helicase at the end of chromosome replication

Marija Maric, Progya Mukherjee, Michael H. Tatham, Ronald Hay, Karim Labib (Lead / Corresponding author)

MRC PPU

Research output: Contribution to journal › Article › peer-review

36 Citations (Scopus)

276 Downloads (Pure)

Abstract

Disassembly of the Cdc45-MCM-GINS 1 (CMG) DNA helicase is the key regulated step during DNA replication termination in eukaryotes, involving ubiquitylation of the Mcm7 helicase subunit, leading to a disassembly process that requires the Cdc48 ‘segregase’. Here we employ a screen to identify partners of budding yeast Cdc48 that are important for disassembly of ubiquitylated CMG helicase at the end of chromosome replication. We demonstrate that the ubiquitin-binding Ufd1-Npl4 complex recruits Cdc48 to ubiquitylated CMG. Ubiquitylation of CMG in yeast cell extracts is dependent upon lysine 29 of Mcm7, which is the only detectable site of ubiquitylation both in vitro and in vivo (though in vivo other sites can be modified when K29 is mutated). Mutation of K29 abrogates in vitro recruitment of Ufd1-Npl4-Cdc48 to the CMG helicase, supporting a model whereby Ufd1-Npl4 recruits Cdc48 to ubiquitylated CMG at the end of chromosome replication, thereby driving the disassembly reaction.

Original language	English
Pages (from-to)	3033-3042
Number of pages	10
Journal	Cell Reports
Volume	18
Issue number	13
DOIs	https://doi.org/10.1016/j.celrep.2017.03.020
Publication status	Published - 28 Mar 2017

Keywords

CMG helicase
DNA replication termination
Cdc48
Ufd1-NpI4
ubiquitylation
ubiquitin
replisome
disassembly

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10.1016/j.celrep.2017.03.020Licence: CC BY

Final Published Version
© 2017 The Author(s). This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
Final published version, 8.8 MBLicence: CC BY

Functional Dissection of the Eukaryotic Replisome (Senior Investigator Award)
Labib, K. (Investigator)
1/04/14 → 29/02/20
Project: Research
Determining the Role and Mechanism of Action of SUMO Targeted Ubiquitin Ligase RNF4 in Maintaining Genome Integrity (Senior Investigator Award)
Hay, R. (Investigator)
1/10/12 → 31/01/20
Project: Research

In vitro reconstitution of the ubiquitylation and disassembly of the eukaryotic replisome
Mukherjee, P. (Author), Labib, K. (Supervisor), 2018
Student thesis: Doctoral Thesis › Doctor of Philosophy

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Labib, Karim
- MRC PPU - Professor of Genome Integrity
Person: Academic

Cite this

@article{7bc944f6571b4a3da2ac8e76150aa474,

title = "Ufd1-Npl4 recruit Cdc48 for disassembly of ubiquitylated CMG helicase at the end of chromosome replication",

abstract = "Disassembly of the Cdc45-MCM-GINS 1 (CMG) DNA helicase is the key regulated step during DNA replication termination in eukaryotes, involving ubiquitylation of the Mcm7 helicase subunit, leading to a disassembly process that requires the Cdc48 {\textquoteleft}segregase{\textquoteright}. Here we employ a screen to identify partners of budding yeast Cdc48 that are important for disassembly of ubiquitylated CMG helicase at the end of chromosome replication. We demonstrate that the ubiquitin-binding Ufd1-Npl4 complex recruits Cdc48 to ubiquitylated CMG. Ubiquitylation of CMG in yeast cell extracts is dependent upon lysine 29 of Mcm7, which is the only detectable site of ubiquitylation both in vitro and in vivo (though in vivo other sites can be modified when K29 is mutated). Mutation of K29 abrogates in vitro recruitment of Ufd1-Npl4-Cdc48 to the CMG helicase, supporting a model whereby Ufd1-Npl4 recruits Cdc48 to ubiquitylated CMG at the end of chromosome replication, thereby driving the disassembly reaction.",

keywords = "CMG helicase, DNA replication termination, Cdc48, Ufd1-NpI4, ubiquitylation, ubiquitin, replisome, disassembly",

author = "Marija Maric and Progya Mukherjee and Tatham, {Michael H.} and Ronald Hay and Karim Labib",

note = "We gratefully acknowledge the support of the Medical Research Council (core grant MC_UU_12016/13), the Wellcome Trust (references 097945/B/11/Z for flow cytometry, 102943/Z/13/Z for an Investigator award to KL and 098391/Z/12/Z for an Investigator award to RTH) and Cancer Research U.K. (reference C434/A13067 for an award to RTH).",

year = "2017",

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doi = "10.1016/j.celrep.2017.03.020",

language = "English",

volume = "18",

pages = "3033--3042",

journal = "Cell Reports",

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T1 - Ufd1-Npl4 recruit Cdc48 for disassembly of ubiquitylated CMG helicase at the end of chromosome replication

AU - Maric, Marija

AU - Mukherjee, Progya

AU - Tatham, Michael H.

AU - Hay, Ronald

AU - Labib, Karim

N1 - We gratefully acknowledge the support of the Medical Research Council (core grant MC_UU_12016/13), the Wellcome Trust (references 097945/B/11/Z for flow cytometry, 102943/Z/13/Z for an Investigator award to KL and 098391/Z/12/Z for an Investigator award to RTH) and Cancer Research U.K. (reference C434/A13067 for an award to RTH).

PY - 2017/3/28

Y1 - 2017/3/28

N2 - Disassembly of the Cdc45-MCM-GINS 1 (CMG) DNA helicase is the key regulated step during DNA replication termination in eukaryotes, involving ubiquitylation of the Mcm7 helicase subunit, leading to a disassembly process that requires the Cdc48 ‘segregase’. Here we employ a screen to identify partners of budding yeast Cdc48 that are important for disassembly of ubiquitylated CMG helicase at the end of chromosome replication. We demonstrate that the ubiquitin-binding Ufd1-Npl4 complex recruits Cdc48 to ubiquitylated CMG. Ubiquitylation of CMG in yeast cell extracts is dependent upon lysine 29 of Mcm7, which is the only detectable site of ubiquitylation both in vitro and in vivo (though in vivo other sites can be modified when K29 is mutated). Mutation of K29 abrogates in vitro recruitment of Ufd1-Npl4-Cdc48 to the CMG helicase, supporting a model whereby Ufd1-Npl4 recruits Cdc48 to ubiquitylated CMG at the end of chromosome replication, thereby driving the disassembly reaction.

AB - Disassembly of the Cdc45-MCM-GINS 1 (CMG) DNA helicase is the key regulated step during DNA replication termination in eukaryotes, involving ubiquitylation of the Mcm7 helicase subunit, leading to a disassembly process that requires the Cdc48 ‘segregase’. Here we employ a screen to identify partners of budding yeast Cdc48 that are important for disassembly of ubiquitylated CMG helicase at the end of chromosome replication. We demonstrate that the ubiquitin-binding Ufd1-Npl4 complex recruits Cdc48 to ubiquitylated CMG. Ubiquitylation of CMG in yeast cell extracts is dependent upon lysine 29 of Mcm7, which is the only detectable site of ubiquitylation both in vitro and in vivo (though in vivo other sites can be modified when K29 is mutated). Mutation of K29 abrogates in vitro recruitment of Ufd1-Npl4-Cdc48 to the CMG helicase, supporting a model whereby Ufd1-Npl4 recruits Cdc48 to ubiquitylated CMG at the end of chromosome replication, thereby driving the disassembly reaction.

KW - CMG helicase

KW - DNA replication termination

KW - Cdc48

KW - Ufd1-NpI4

KW - ubiquitylation

KW - ubiquitin

KW - replisome

KW - disassembly

U2 - 10.1016/j.celrep.2017.03.020

DO - 10.1016/j.celrep.2017.03.020

M3 - Article

C2 - 28355556

SN - 2211-1247

VL - 18

SP - 3033

EP - 3042

JO - Cell Reports

JF - Cell Reports

IS - 13

ER -

Ufd1-Npl4 recruit Cdc48 for disassembly of ubiquitylated CMG helicase at the end of chromosome replication

Abstract

Keywords

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Projects

Functional Dissection of the Eukaryotic Replisome (Senior Investigator Award)

Determining the Role and Mechanism of Action of SUMO Targeted Ubiquitin Ligase RNF4 in Maintaining Genome Integrity (Senior Investigator Award)

Student theses

In vitro reconstitution of the ubiquitylation and disassembly of the eukaryotic replisome

Profiles

Labib, Karim

Cite this