Periodic Reporting for period 2 - DNAREPAIRMAN (Regulated Assembly of Molecular Machines for DNA REPAIR: a Molecular Analysis training Network)
Période du rapport: 2019-01-01 au 2020-12-31
Our focus on the process of DNA repair arises from its essential role in maintaining genome stability. In this process multiple proteins collaborate as large molecular machines to detect and repair very rare errors that occur in DNA. These evolutionarily-conserved processes are of fundamental importance, and there is an increasing awareness that targeting of DNA repair pathways could also be useful in the development of personalized anti-cancer therapies and new antimicrobials.
This Training Network uniquely brings together biologists, chemists, and physicists around a common research question: What are the mechanistic details that underlie the molecular organization of DNA repair?
The objectives of DNARepairMan are
- To unravel the mechanistic details of two DNA repair pathways and how they interact with DNA replication and transcription
- To develop new reagents, assays, technology and software to analyze fundamental chemical and physical principles
- collaborate with an Art Academy to explore new ways and means to visualize research projects and explain the importance of fundamental research to society.
The protein that recognizes the replication error, MutS, has to recognize this error, bind to it and only then recruit the next protein in the repair cascade. During this process MutS changes shape multiple times. Using a high resolution technique called cryo-electron microscopy, the consortium has taken snapshots of these different states, which give unprecedented detail and explain how MutS does its important job.
Understanding which proteins collaborate at which reactions steps and how they do this: During the DNA repair process, many proteins collaborate to 1) find copy errors or damage in the DNA, 2) recruit partner proteins that can remove the error or damage, and 3) recruit proteins that can restore the original information. In order to understand the molecular mechanism of these processes, the consortium has designed new reagents, assays and technologies. These are used to study the mechanism of the repair processes in detail.
The training program: The training program is an essential and central component of DNARepairMan. The consortium has organized four scientific workshops, which have been attended by all the fellows as well as several researchers from outside the consortium. The workshops entailed ‘Protein Structure Analysis’, ‘Advanced single molecule analysis’, ‘Fluorescence and cross linking approaches’ and 'Hybrid structural biology. Fellows have also received entrepreneurship coaching sessions to prepare them for a putative career as entrepreneur, a lecture on Research Ethics, as well as other transferable skills and a career event.
DNARepairMan is a truly interactive consortium; all the young researchers have already spend prolonged research visits (secondments) in each other’s laboratories to learn new techniques and to collect data.
A unique component of DNARepairMan is the active collaboration with the Art Academy in Rotterdam (de Willem de Kooning Akademy). Through an intensive program including the DNArepairMan fellows, scientists, students and teachers from the Art Academy and the Erasmus MC, the consortium is exploring new ways and means to communicate scientific topics and results as well as the importance of fundamental science in general to society. This has resulted in several short animations about the scientific projects and the importance of science in general.
• DNA molecules carrying specific errors or DNA damage combined with fluorescent labels that allow readout of mechanisms of communication and achieving directional DNA repair
• Purified protein molecules that carry fluorescent labels that allow quantitative analysis with interaction partners.
• Purified protein molecules with functional groups that allow cross linking interaction analysis.
• Purified protein variants that carry changes in structure to analyze functional importance
• Single molecule assays that allow detailed identification of binding partners and kinetics of reaction steps
• Microfluidic flow cells that allow controlled reagent addition during single molecule analysis
• New fluorescent assays to quantify protein interactions
• New biochemical assays to quantify DNA repair with purified components.
The abovementioned investments in beyond-the-state-of-the-art developments and extensive training of the young DNARepairMan researchers have lead to 1) important new understanding of the molecular mechanism underlying DNA repair with impact on human health, and 2) a new generation of innovative young scientists which has been trained in cutting-edge biophysics and with affinity for the academic, industrial and creative sector.