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Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method
Published on: May 2, 2025
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PARP1 auto-modification promotes faithful Okazaki fragment processing and limits replication fork speed
Jonas D Elsborg1, Sebastian H N Munk2, Alba Adelantado-Rubio2
1Department of Cellular and Molecular Medicine, Novo Nordisk Foundation Center for Protein Research, Proteomics Program, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
Molecular Cell
|October 3, 2025
Summary
Poly(ADP-ribose) polymerase inhibitors target cancer by exploiting synthetic lethality. A new study reveals PARP1 auto-modification, distinct from its enzymatic activity, is crucial for DNA repair and preventing replication stress.
Area of Science:
- Molecular Biology
- Genetics
- Cancer Research
Background:
- Poly(ADP-ribose) polymerase (PARP) inhibitors are effective cancer treatments for tumors with homologous recombination deficiencies, utilizing synthetic lethality.
- PARP1 is a key enzyme in DNA break response, performing auto-modification and modifying other proteins to aid repair.
- Dissecting the roles of PARP1 auto-modification versus its catalytic activity is challenging due to current inhibitor mechanisms.
Purpose of the Study:
- To differentiate the functional roles of PARP1 auto-modification and catalytic activity in DNA repair and cellular response.
- To investigate the impact of PARP1 auto-modification on DNA replication and stress response.
- To explore novel synthetic lethality interactions involving PARP1 auto-modification.
Main Methods:
- Proteomics and site-directed mutagenesis were employed to create a PARP1 mutant with impaired auto-modification but retained catalytic activity.
- The functional consequences of this separation-of-function mutant were analyzed in response to DNA damage.
- Synthetic lethality was assessed by combining loss of PARP1 auto-modification with inhibition of other cellular processes, such as FEN1 activity.
Main Results:
- A PARP1 mutant deficient in auto-modification, but catalytically active, was generated.
- PARP1 auto-modification was found to impede DNA replication fork progression but is not essential for recruiting repair factors.
- Auto-modification facilitates PARP1 release from DNA breaks, prevents replication stress, and is implicated in Okazaki fragment processing, showing synthetic lethality with FEN1 inhibition.
Conclusions:
- PARP1 auto-modification has distinct roles separate from its catalytic activity, influencing replication fork dynamics and protein release at DNA breaks.
- Trapping PARP1 at DNA break sites hinders repair factor access, contributing to PARP inhibitor cytotoxicity.
- Targeting PARP1 auto-modification presents a potential new avenue for cancer therapy, particularly in combination strategies.
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