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The Initiation of Eukaryotic DNA Replication: Mechanism, Regulation and Role in Genome Stability

Objective

In each cell cycle, eukaryotic cells must faithfully replicate large genomes in a relatively short time. This is accomplished by initiating DNA replication from many replication origins distributed along chromosomes. Ensuring that each origin is efficiently activated once and only once per cell cycle is crucial for maintaining the integrity of the genome. Recent evidence indicates that defects in the regulation of origin firing may be important contributors to genome instability in cancer. Strict once per cell cycle DNA replication is achieved by a two-step mechanism. DNA replication origins are first licensed by loading an inactive DNA helicase (Mcm2-7) into pre-replicative complexes (pre-RCs). This can only occur during G1 phase. Initiation then occurs during S phase, triggered by cyclin dependent kinases (CDKs) and Dbf4-dependent kinase (DDK), which promote recruitment of proteins required for helicase activation and replisome assembly. Research proposed herein will lead to a deeper understanding of the mechanism and regulation of DNA replication. We have reconstituted the licensing reaction with purified proteins which will be used to characterise the mechanism of licensing and the mechanism by which licensing is regulated in the cell cycle. We will also use this system to reconstitute events leading to the initiation of DNA replication. We will use genetic and biochemical approaches to characterise the mechanisms by which perturbed licensing causes gross chromosome rearrangements. We will also explore mechanisms involved in regulating the temporal programme of origin firing and how origin firing is regulated in response to DNA damage. Work in budding yeast and mammalian cells will be pursued in parallel to exploit the specific advantages of each system.

Field of science

  • /natural sciences/biological sciences/genetics and heredity/chromosome
  • /natural sciences/biological sciences/biochemistry/biomolecules/proteins
  • /medical and health sciences/clinical medicine/oncology/cancer
  • /natural sciences/biological sciences/genetics and heredity/genome

Call for proposal

ERC-2009-AdG
See other projects for this call

Funding Scheme

ERC-AG - ERC Advanced Grant

Host institution

THE FRANCIS CRICK INSTITUTE LIMITED
Address
1 Midland Road
NW1 1AT London
United Kingdom
Activity type
Research Organisations
EU contribution
€ 2 449 998,99
Principal investigator
John Diffley (Dr.)
Administrative Contact
Heather Joanne Woods (Ms.)

Beneficiaries (2)

THE FRANCIS CRICK INSTITUTE LIMITED
United Kingdom
EU contribution
€ 2 449 998,99
Address
1 Midland Road
NW1 1AT London
Activity type
Research Organisations
Principal investigator
John Diffley (Dr.)
Administrative Contact
Heather Joanne Woods (Ms.)
CANCER RESEARCH UK LBG

Participation ended

United Kingdom
Address
St John Street 407 Angel Building
EC1V 4AD London
Activity type
Research Organisations
Administrative Contact
Holly Elphinstone (Ms.)