Unravelling the mechanism of eukaryotic helicase activation

Project Information

MechHelicaseActiv8on

Grant agreement ID: 101025825

DOI

10.3030/101025825

Project closed

EC signature date 27 April 2021

Start date 1 September 2022

End date 31 August 2024

Funded under

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Total cost

€ 212 933,76

EU contribution

€ 212 933,76

Coordinated by

THE FRANCIS CRICK INSTITUTE LIMITED
United Kingdom

Periodic Reporting for period 1 - MechHelicaseActiv8on (Unravelling the mechanism of eukaryotic helicase activation)

Reporting period: 2022-09-01 to 2024-08-31

Summary of the context and overall objectives of the project

The project aimed to unravel the mechanism of eukaryotic helicase activation in DNA replication initiation. During DNA replication, helicase is essential for unwinding the DNA double helix, a step critical for cellular division. However, the exact mechanisms by which the helicase is activated to start DNA unwinding, including the transition from double-stranded to single-stranded DNA, remain unclear.
Understanding DNA replication mechanisms is fundamental to advancing knowledge in genome maintenance. Errors in DNA replication can lead to genetic mutations, contributing to cancer and other genetic disorders. By shedding light on helicase activation, this research has the potential to aid in the development of targeted therapies for diseases that result from DNA replication errors.
The main objectives of the project were: (1) to uncover the fundamental mechanisms by which helicase is activated to initiate DNA unwinding, (2) to investigate the specific roles of various protein components within the helicase complex.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

For helicase to get activated during DNA replication initiation, the helicase has to transit from double stranded DNA to single stranded DNA. To find out the trajectory of lagging strand template during helicase activation, I purified all essential DNA replication proteins and confirmed their activity in reconstituted DNA replication assay. I prepared a set of helicase mutants required for testing the helicase activation mechanism.
Mcm10 assists helicase activation by supporting the transition from double stranded DNA to single stranded DNA and increasing replication speed. Together with our collaborators, we combined biochemistry with structural biology to further understand the mechanism of Mcm10 action. We identified distinct roles for the N-terminal (initiates replication) and C-terminal (increases speed) of Mcm10. Mutations in Mcm10 confirmed that its interaction with single stranded DNA is not essential for replication. We disseminated the findings by publishing in Nature Structural Biology with Open Acces.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

The project contributed to our understanding of Mcm10 function as well as provided a separation-of-function mutant that might be useful for further research in the field. The helicase mutants might be further used in testing other processes, including how the helicase bypass DNA lesion. This could lead to further insight on not only the mechanism of DNA replication, but also other processes like DNA repair, which is important for genome stability.