Overview

The study focuses on the enzyme p300, a lysine acetyltransferase (KAT), and its role in modifying RPA through acetylation. RPA is a heterotrimeric protein complex that binds to single-stranded DNA and plays a central role in DNA replication, recombination, and repair. The researchers aimed to determine how acetylation of RPA by p300 influences its function and, by extension, the cell’s choice of DNA repair pathways. This is particularly relevant in the context of cancer biology, where DNA repair mechanisms are often dysregulated.

Key findings

The researchers discovered that p300 acetylates specific lysine residues on the RPA1 subunit, particularly lysine 163 (K163). This modification alters the biochemical properties of RPA, affecting its interaction with other proteins involved in DNA repair. Notably, acetylation at K163 was shown to influence the recruitment of RPA to sites of DNA damage and modulate its binding to single-stranded DNA.

Using mass spectrometry and immunoblotting techniques, the team confirmed the presence of acetylated lysines on RPA1. They also demonstrated that acetylation by p300 promotes a shift in DNA repair pathway choice, favoring homologous recombination over non-homologous end joining. This finding adds a new layer of regulation to the already complex network of DNA repair mechanisms.

Abcam’s anti-RPA1 antibody was used extensively in immunoblotting experiments to detect total RPA1 levels across different experimental conditions. This antibody enabled the researchers to monitor changes in RPA1 expression and assess the impact of p300-mediated acetylation on the protein’s stability and function.

Implications

By showing that p300-mediated acetylation of RPA influences DNA repair pathway choice, the study suggests a potential mechanism by which cells regulate genome stability. This could be particularly relevant in cancer cells, where mutations in DNA repair genes are common.

Moreover, targeting the acetylation process or modulating p300 activity could offer new therapeutic strategies. For example, inhibiting p300 in tumors that rely heavily on homologous recombination might sensitize them to DNA-damaging agents or PARP inhibitors.

Future work

The study opens up several avenues for future research. One area of interest is exploring how other post-translational modifications of RPA interact with acetylation to fine-tune its function. Additionally, further studies could investigate whether similar regulatory mechanisms exist in other DNA-binding proteins.

The authors also suggest examining the role of p300-mediated acetylation in different cell types and under various stress conditions. This could help determine whether the observed effects are universal or context-dependent. Ongoing research in this area may also explore the therapeutic potential of modulating RPA acetylation in cancer treatment.

References

Ononye, O. et al. Biochemical impact of p300-mediated acetylation of replication protein A: implications for DNA metabolic pathway choice.  J. Biol. Chem.  301, 110250 (2025). https://doi.org/10.1016/j.jbc.2025.110250