Nucleosomes effectively shield DNA from radiation damage in living cells

Our plain-English summary of Nucleosomes effectively shield DNA from radiation damage in living cells, by F. Brambilla, J.M. Garcia-Manteiga, E. Monteleone, et al (Nucleic Acids Research, 2020).

The study

• Authors: F. Brambilla, J.M. Garcia-Manteiga, E. Monteleone, L. Hoelzen, A. Zocchi, A. Agresti, M.E. Bianchi
• Journal: Nucleic Acids Research
• Published: 2020;48(16):8993-9006
• Cited by: 41 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Eukaryotic DNA is organized in nucleosomes, which package DNA and regulate its accessibility to transcription, replication, recombination and repair. Here, we show that in living cells nucleosomes protect DNA from high-energy radiation and reactive oxygen species. We combined sequence-based methods (ATAC-seq and BLISS) to determine the position of both nucleosomes and double strand breaks (DSBs) in the genome of nucleosome-rich malignant mesothelioma cells, and of the same cells partially depleted of nucleosomes. The results were replicated in the human MCF-7 breast carcinoma cell line. We found that, for each genomic sequence, the probability of DSB formation is directly proportional to the fraction of time it is nucleosome-free; DSBs accumulate distal from the nucleosome dyad axis. Nucleosome free regions and promoters of actively transcribed genes are more sensitive to DSB formation, and consequently to mutation. We argue that this may be true for a variety of chemical and physical DNA damaging agents.

© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.

In plain English

This laboratory study used cultured cells, not people, and did not test asbestos. The authors used a mouse malignant mesothelioma cell line (AB22) as a model because it contained more histone H3, a protein around which DNA wraps to form nucleosomes, than normal mouse mesothelial cells. They lowered nucleosome levels by knocking down a protein called HMGB1, which reduced histone H3 by 10–15%. They then exposed the cells to gamma radiation (0, 3, 5 and 10 Gy) and mapped DNA double strand breaks across the genome. Among irradiated cells, those with fewer nucleosomes had significantly more DNA breaks at every dose. Without radiation, a DNA-break test (comet assay) found no statistically significant difference. Breaks clustered in open, nucleosome-free stretches of DNA and near the start points of highly active genes. On average, the break probability was 0.02 for nucleosomes with 99–100% occupancy, compared with 0.09 for nucleosomes with 20% occupancy. Human MCF-7 breast cancer cells showed a similar inverse relationship. The authors concluded that nucleosomes shield DNA from ionizing radiation and reactive oxygen species, and they argued this may be true for other damaging agents.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory experiment on cultured mouse and human cell lines (genome mapping of nucleosomes and radiation-induced DNA double strand breaks)
• Who was studied: Mouse malignant mesothelioma AB22 cells, unmodified (shCTRL) and HMGB1-depleted (shHMGB1). Comparison cells were primary mesothelial cells from the peritoneal cavity of 6–10 week old male BALB/c mice, human mesothelioma REN and mesothelial LP9 cells, and human MCF-7 breast cancer and MCF10A breast epithelial cells. The work was done in Milan, Italy.
• Exposure: Gamma radiation from a cesium-137 source at 0, 3, 5 and 10 Gy for mesothelioma cells, and 0 or 10 Gy for MCF-7 cells, some of which were pretreated with the antioxidant N-acetyl-L-cysteine. Asbestos was mentioned only as background (mesothelioma described as arising from asbestos exposure). No asbestos exposure was tested.
• Main results:
  • Mouse mesothelioma cells contained significantly more histone H3 than primary mesothelial cells.
  • HMGB1 knockdown in mesothelioma cells reduced histone H3 by 10–15% compared with control cells.
  • HMGB1-depleted mesothelioma cells had about 10% fewer nucleosomes and 4% more nucleosome-free regions than control cells.
  • In the comet assay, HMGB1-depleted mesothelioma cells had significantly more DNA breaks than control cells at every radiation dose.
  • In the comet assay, non-irradiated HMGB1-depleted and control mesothelioma cells were not statistically different.
  • The number of double strand breaks rose with radiation dose and was significantly higher at each dose in HMGB1-depleted cells than in control cells.
  • In both control and HMGB1-depleted mesothelioma cells, double strand breaks were strongly enriched in open chromatin regions relative to the whole genome.
  • In both cell types, double strand breaks were enriched in nucleosome-free regions compared with nucleosome-occupied regions.
  • Double strand breaks were enriched at the start sites of the most transcribed genes, with no clear difference between the two cell types.
  • In mesothelioma cells, the average probability of a double strand break was about 4-fold lower (0.02 versus 0.09) for nucleosomes with 99–100% occupancy than for nucleosomes with 20% occupancy.
  • In irradiated MCF-7 cells, radiation significantly increased double strand breaks, and pretreatment with N-acetyl-L-cysteine largely suppressed this increase. The figure legend reports that the NAC treatment effect alone was not significant (P = 0.28), while the interaction of irradiation and NAC was significant (P = 0.043).
  • The inverse correlation between nucleosome occupancy and break probability was also found in MCF-7 cells. The log-log slopes ranged from minus 0.7 to minus 1.0 in MCF-7 cells and from minus 0.7 to minus 0.8 in mesothelioma cells.
• The authors' conclusion: The authors concluded that nucleosomes effectively shield DNA from ionizing radiation and reactive oxygen species. They argued this may be true for a variety of chemical and physical DNA-damaging agents.
• Limitations the authors state:
  • In MCF-7 cells, nucleosome data came from a public dataset. That dataset identified a higher proportion of highly occupied nucleosomes than in mesothelioma cells, possibly because of DNA size selection. The authors called this a potential confounding factor.
  • MCF-7 experiments were performed on cycling cells, whereas mesothelioma cells were synchronized in G1 to limit breaks from DNA replication.

Topics

Medical terms in this study, explained in our Lexicon: Reactive Oxygen Species (ROS), Mesothelioma, Carcinoma.

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