Widespread chromatin context-dependencies of DNA double-strand break repair proteins

Vergara, Xabier; Manjón, Anna G.; de Haas, Marcel; Morris, Ben; Schep, Ruben; Leemans, Christ; Friskes, Anoek; Beijersbergen, Roderick L.; Sanders, Mathijs A.; Medema, René H.; van Steensel, Bas

doi:10.1038/s41467-024-49232-x

Cited by 3 publications

References 59 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

Chromatin context-dependent effects of epigenetic drugs on CRISPR-Cas9 editing

Schep,

Trauernicht,

Vergara

et al. 2024

Nucleic Acids Research

View full text Add to dashboard Cite

The efficiency and outcome of CRISPR/Cas9 editing depends on the chromatin state at the cut site. It has been shown that changing the chromatin state can influence both the efficiency and repair outcome, and epigenetic drugs have been used to improve Cas9 editing. However, because the target proteins of these drugs are not homogeneously distributed across the genome, the efficacy of these drugs may be expected to vary from locus to locus. Here, we systematically analyzed this chromatin context-dependency for 160 epigenetic drugs. We used a human cell line with 19 stably integrated reporters to induce a double-stranded break in different chromatin environments. We then measured Cas9 editing efficiency and repair pathway usage by sequencing the mutational signatures. We identified 58 drugs that modulate Cas9 editing efficiency and/or repair outcome dependent on the local chromatin environment. For example, we find a subset of histone deacetylase inhibitors that improve Cas9 editing efficiency throughout all types of heterochromatin (e.g. PCI-24781), while others were only effective in euchromatin and H3K27me3-marked regions (e.g. apicidin). In summary, this study reveals that most epigenetic drugs alter CRISPR editing in a chromatin-dependent manner, and provides a resource to improve Cas9 editing more selectively at the desired location.

show abstract

Chromatin context-dependent effects of epigenetic drugs on CRISPR-Cas9 editing

Schep,

Trauernicht,

Vergara

et al. 2024

Nucleic Acids Research

View full text Add to dashboard Cite

show abstract

DNA double-strand break movement in heterochromatin depends on the histone acetyltransferase dGcn5

Kendek,

Sandron,

Lambooij

et al. 2024

Nucleic Acids Research

View full text Add to dashboard Cite

Cells employ diverse strategies to repair double-strand breaks (DSBs), a dangerous form of DNA damage that threatens genome integrity. Eukaryotic nuclei consist of different chromatin environments, each displaying distinct molecular and biophysical properties that can significantly influence the DSB-repair process. DSBs arising in the compact and silenced heterochromatin domains have been found to move to the heterochromatin periphery in mouse and Drosophila to prevent aberrant recombination events. However, it is poorly understood how chromatin components, such as histone post-translational modifications, contribute to these DSB movements within heterochromatin. Using irradiation as well as locus-specific DSB induction in Drosophila tissues and cultured cells, we find enrichment of histone H3 lysine 9 acetylation (H3K9ac) at DSBs in heterochromatin but not euchromatin. We find this increase is mediated by the histone acetyltransferase dGcn5, which rapidly localizes to heterochromatic DSBs. Moreover, we demonstrate that in the absence of dGcn5, heterochromatic DSBs display impaired recruitment of the SUMO E3 ligase Nse2/Qjt and fail to relocate to the heterochromatin periphery to complete repair. In summary, our results reveal a previously unidentified role for dGcn5 and H3K9ac in heterochromatic DSB repair and underscore the importance of differential chromatin responses at heterochromatic and euchromatic DSBs to promote safe repair.

show abstract

Cell cycle regulation of polymerase theta mediated end-joining underlies its sensitivity to loss of BLM-DNA2

McBrine,

Jensen,

King

2024

Preprint

View full text Add to dashboard Cite

DNA double-strand breaks (DSBs) are predominantly resolved by the error-prone non-homologous end-joining (NHEJ) or high-fidelity homologous recombination (HR) DNA repair pathways. NHEJ involves minimal end-processing and drives direct ligation of the broken ends, while HR first requires a DSB resection step that degrades 5′ strands to generate 3′ ssDNA tails necessary for homology search. When HR fails, an alternative end-joining pathway involving polymerase theta can serve as a backup repair mechanism by annealing and ligating microhomologies present in ssDNA tails on either side of a resected DSB. Resection is the early step that commits a DSB to repair by HR, and is therefore considered a prerequisite for TMEJ as well. It has been proposed that the exonuclease EXO1 and the helicase/nuclease complex of BLM-DNA2 have redundant roles in resection, and therefore may both contribute to TMEJ. Here, we use a sensitive deep-sequencing approach to identify DSB repair outcomes at an endogenous locus in response to genetic and chemical perturbation of the resection machinery. While we observe that both EXO1 and BLM-DNA2 contribute to resection, our findings highlight a predominant role for BLM-DNA2 in facilitating TMEJ. However, careful cell cycle analysis revealed that BLM-DNA primarily contributes to TMEJ by ensuring normal progression to the G2/M transition, required for TMEJ to become activated. We propose that rather than direct, mechanistic roles in resection, the critical functions of BLM and DNA2 in genome replication largely account for their genetic contribution to efficient TMEJ.

show abstract

Widespread chromatin context-dependencies of DNA double-strand break repair proteins

Cited by 3 publications

References 59 publications

Chromatin context-dependent effects of epigenetic drugs on CRISPR-Cas9 editing

Chromatin context-dependent effects of epigenetic drugs on CRISPR-Cas9 editing

DNA double-strand break movement in heterochromatin depends on the histone acetyltransferase dGcn5

Cell cycle regulation of polymerase theta mediated end-joining underlies its sensitivity to loss of BLM-DNA2

Contact Info

Product

Resources

About