Publication Date:
2014-05-09
Description:
Efficient duplication of the genome requires the concerted action of helicase and DNA polymerases at replication forks to avoid stalling of the replication machinery and consequent genomic instability. In eukaryotes, the physical coupling between helicase and DNA polymerases remains poorly understood. Here we define the molecular mechanism by which the yeast Ctf4 protein links the Cdc45-MCM-GINS (CMG) DNA helicase to DNA polymerase alpha (Pol alpha) within the replisome. We use X-ray crystallography and electron microscopy to show that Ctf4 self-associates in a constitutive disk-shaped trimer. Trimerization depends on a beta-propeller domain in the carboxy-terminal half of the protein, which is fused to a helical extension that protrudes from one face of the trimeric disk. Critically, Pol alpha and the CMG helicase share a common mechanism of interaction with Ctf4. We show that the amino-terminal tails of the catalytic subunit of Pol alpha and the Sld5 subunit of GINS contain a conserved Ctf4-binding motif that docks onto the exposed helical extension of a Ctf4 protomer within the trimer. Accordingly, we demonstrate that one Ctf4 trimer can support binding of up to three partner proteins, including the simultaneous association with both Pol alpha and GINS. Our findings indicate that Ctf4 can couple two molecules of Pol alpha to one CMG helicase within the replisome, providing a new model for lagging-strand synthesis in eukaryotes that resembles the emerging model for the simpler replisome of Escherichia coli. The ability of Ctf4 to act as a platform for multivalent interactions illustrates a mechanism for the concurrent recruitment of factors that act together at the fork.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4059944/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉 〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4059944/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Simon, Aline C -- Zhou, Jin C -- Perera, Rajika L -- van Deursen, Frederick -- Evrin, Cecile -- Ivanova, Marina E -- Kilkenny, Mairi L -- Renault, Ludovic -- Kjaer, Svend -- Matak-Vinkovic, Dijana -- Labib, Karim -- Costa, Alessandro -- Pellegrini, Luca -- 084279/Wellcome Trust/United Kingdom -- Wellcome Trust/United Kingdom -- Medical Research Council/United Kingdom -- England -- Nature. 2014 Jun 12;510(7504):293-7. doi: 10.1038/nature13234. Epub 2014 May 4.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK [2]. ; 1] Clare Hall Laboratories, Cancer Research UK London Research Institute, London EN6 3LD, UK [2]. ; 1] Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK [2] Imperial College, South Kensington, London SW7 2AZ, UK (R.L.P.); Cancer Research UK London Research Institute, London WC2A 3LY, UK (M.E.I.). ; Cancer Research UK Manchester Institute, University of Manchester, Manchester M20 4BX, UK. ; MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dundee DD1 5EH, UK. ; Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK. ; Clare Hall Laboratories, Cancer Research UK London Research Institute, London EN6 3LD, UK. ; Protein purification, Cancer Research UK London Research Institute, London WC2A 3LY, UK. ; Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, UK.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24805245" target="_blank"〉PubMed〈/a〉
Keywords:
Amino Acid Motifs
;
Amino Acid Sequence
;
Catalytic Domain
;
Conserved Sequence
;
Crystallography, X-Ray
;
DNA Helicases/chemistry/*metabolism/ultrastructure
;
DNA Polymerase I/chemistry/*metabolism/ultrastructure
;
*DNA Replication
;
DNA-Binding Proteins/*chemistry/*metabolism/ultrastructure
;
DNA-Directed DNA Polymerase/*chemistry/*metabolism
;
Microscopy, Electron
;
Minichromosome Maintenance Proteins/chemistry/metabolism
;
Models, Molecular
;
Molecular Sequence Data
;
Multienzyme Complexes/*chemistry/*metabolism
;
Nuclear Proteins/chemistry/metabolism
;
Protein Binding
;
*Protein Multimerization
;
Protein Structure, Quaternary
;
Protein Subunits/chemistry/metabolism
;
Saccharomyces cerevisiae/*chemistry/ultrastructure
;
Saccharomyces cerevisiae Proteins/*chemistry/*metabolism/ultrastructure
Print ISSN:
0028-0836
Electronic ISSN:
1476-4687
Topics:
Biology
,
Chemistry and Pharmacology
,
Medicine
,
Natural Sciences in General
,
Physics
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