Periodic Reporting for period 4 - DPC_REPAIR (Mechanism of DNA-protein cross-link repair in S phase)
Periodo di rendicontazione: 2021-07-01 al 2022-06-30
We are addressing how a specific type of DNA lesion known as DNA-protein crosslink is repaired during DNA replication.
This is highly relevant to society because DNA-protein crosslinks are highly cytotoxic lesions known to trigger cancer in humans. Moreover, DNA-protein crosslinks are the byproduct of most chemotherapeutic drugs used in the clinic to treat cancer.
Thus, if we can understand how DNA-protein crosslinks are repaired, we will be able to develop new treatments to fight cancer in the futur.
This is highly relevant to society because DNA-protein crosslinks are highly cytotoxic lesions known to trigger cancer in humans. Moreover, DNA-protein crosslinks are the byproduct of most chemotherapeutic drugs used in the clinic to treat cancer.
Thus, if we can understand how DNA-protein crosslinks are repaired, we will be able to develop new treatments to fight cancer in the futur.
During the 5 years of this project, we have studied how DNA protein crosslinks are repaired during DNA replication but also how they are sensed and recognised in the absence of DNA replication.
Particularly, we have delineated the molecular function of SPRTN in DPC repair and provided the molecular events that activate and target the protease to DNA-protein crosslinks. Contrary to what was anticipated, SPRTN-mediated DPC degradation does not require DPC ubiquitylation. Instead the protease is activated when a nascent strand is extended to the lesion which appears to be the essential event that targets the protease behind replication forks (to maintain the replisome away from protease activity).
These findings were published in Molecular Cell and were made open source:
https://www.cell.com/molecular-cell/fulltext/S1097-2765(18)30994-8?_returnURL=https%3A%2F%2Flinkinghub.elsevier.com%2Fretrieve%2Fpii%2FS1097276518309948%3Fshowall%3Dtrue
We also participated in a study that showed how DPCs can be bypassed by the CMG helicase when encountered during DNA replication on the leading strand template (the strand CMG translocates on) (published in Cell and made open source). This highlighted the remarkable capacity of the replisome to bypass/traverse large protein roadblocks:
https://www.sciencedirect.com/science/article/pii/S0092867418314521?via%3Dihub
Conversely, DPCs encountered on the lagging strand only partially block CMG translocation when these DPCs interact with duplex DNA (published in Cell Reports and made open source).
https://www.sciencedirect.com/science/article/pii/S2211124719301226?via%3Dihub
More recently, we showed how the ubiquitin ligase RFWD3 ubiquitylates DPCs on single strand DNA and is a key regulator of DNA damage bypass by translesion DNA synthesis (published in Molecular Cell and made open access):
https://www.sciencedirect.com/science/article/pii/S1097276520308297?via%3Dihub
We also shown how DPCs are targeted to degradation in the absence of DNA replication via the SUMO-RNF4 pathway (published in EMBO and made open access):
https://www.sciencedirect.com/science/article/pii/S1097276520308297?via%3Dihub
and unravelled a previously unknown function of the protein SCAI in DNA crosslink repair (published in EMBO reports 2022, open access):
https://www.embopress.org/doi/full/10.15252/embr.202153639
Thanks to the ERC funding we also made publically available a nice review on DPC repair mechanisms that summarises our findings and contributions to the field (published in DNA repair and made open access)
https://www.sciencedirect.com/science/article/pii/S1568786420301737?via%3Dihub
We are currently about to submit a new story about the bypass of DPCs by translesion DNA synthesis. This project has received partial funding from my ERC grant and our results will be made publically available once published.
Our findings were also made available for laymen public in several occasions:
https://www.cpr.ku.dk/cpr-news/2020/researchers-discover-protein-function-that-could-improve-chemotherapy-in-the-future/
https://www.cpr.ku.dk/cpr-news/2018/frog-eggs-help-researchers-understands-repair-of-dna-damages/
Particularly, we have delineated the molecular function of SPRTN in DPC repair and provided the molecular events that activate and target the protease to DNA-protein crosslinks. Contrary to what was anticipated, SPRTN-mediated DPC degradation does not require DPC ubiquitylation. Instead the protease is activated when a nascent strand is extended to the lesion which appears to be the essential event that targets the protease behind replication forks (to maintain the replisome away from protease activity).
These findings were published in Molecular Cell and were made open source:
https://www.cell.com/molecular-cell/fulltext/S1097-2765(18)30994-8?_returnURL=https%3A%2F%2Flinkinghub.elsevier.com%2Fretrieve%2Fpii%2FS1097276518309948%3Fshowall%3Dtrue
We also participated in a study that showed how DPCs can be bypassed by the CMG helicase when encountered during DNA replication on the leading strand template (the strand CMG translocates on) (published in Cell and made open source). This highlighted the remarkable capacity of the replisome to bypass/traverse large protein roadblocks:
https://www.sciencedirect.com/science/article/pii/S0092867418314521?via%3Dihub
Conversely, DPCs encountered on the lagging strand only partially block CMG translocation when these DPCs interact with duplex DNA (published in Cell Reports and made open source).
https://www.sciencedirect.com/science/article/pii/S2211124719301226?via%3Dihub
More recently, we showed how the ubiquitin ligase RFWD3 ubiquitylates DPCs on single strand DNA and is a key regulator of DNA damage bypass by translesion DNA synthesis (published in Molecular Cell and made open access):
https://www.sciencedirect.com/science/article/pii/S1097276520308297?via%3Dihub
We also shown how DPCs are targeted to degradation in the absence of DNA replication via the SUMO-RNF4 pathway (published in EMBO and made open access):
https://www.sciencedirect.com/science/article/pii/S1097276520308297?via%3Dihub
and unravelled a previously unknown function of the protein SCAI in DNA crosslink repair (published in EMBO reports 2022, open access):
https://www.embopress.org/doi/full/10.15252/embr.202153639
Thanks to the ERC funding we also made publically available a nice review on DPC repair mechanisms that summarises our findings and contributions to the field (published in DNA repair and made open access)
https://www.sciencedirect.com/science/article/pii/S1568786420301737?via%3Dihub
We are currently about to submit a new story about the bypass of DPCs by translesion DNA synthesis. This project has received partial funding from my ERC grant and our results will be made publically available once published.
Our findings were also made available for laymen public in several occasions:
https://www.cpr.ku.dk/cpr-news/2020/researchers-discover-protein-function-that-could-improve-chemotherapy-in-the-future/
https://www.cpr.ku.dk/cpr-news/2018/frog-eggs-help-researchers-understands-repair-of-dna-damages/
Our findings provide new and important mechanistic insights on how DNA-protein crosslinks are repaired which go well beyond the state of the art of the field. Moreover, they have identified new factors that participate in the repair reaction. These factors provide new attractive targets for drug development that could be used to sensitise cells to chemotherapeutic drugs.