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Effect of DNA damage and chromatin damage on transcription in HT1080 cells with wild type p53 and p53 inhibited by dominant negative mutant

ABSTRACT: DNA-targeting drugs may damage DNA or chromatin. Many anti-cancer drugs do both, making difficult understanding their mechanism of actions. Using molecules causing DNA breaks without altering nucleosome structure (bleomycin) or destabilizing nucleosomes without damaging DNA (curaxin), we investigated consequences of DNA or chromatin damage in normal and tumor cells. As expected, DNA damage caused p53-dependent growth arrest followed by senescence. Chromatin damage caused higher p53 accumulation than DNA damage, but growth arrest was p53-independent and did not result in senescence. Chromatin damage activated transcription of multiple genes, including classical p53 targets, in a p53-independent manner. These activated genes, though they were not highly expressed, have chromatin organization around transcription start sites (TSS) characteristic for the most highly expressed genes and the highest level of paused RNA polymerase. We proposed that nucleosomes around TSS of these genes are the most sensitive to chromatin damage, and therefore nucleosome loss upon curaxin treatment enables transcription without assistance of sequence-specific transcriptional factors. We confirmed this by showing greater loss of nucleosomes around TSS of these genes upon curaxin treatment and activation of specific p53 reporter in p53-null cells by chromatin damaging agents, but not DNA damaging agents.

ORGANISM(S): Homo sapiens

PROVIDER: GSE223327 | GEO | 2023/09/18

REPOSITORIES: GEO

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Project description:Small molecule curaxin CBL0137 has broad anti-cancer activity in different preclinical models. It interferes with histone-DNA interactions via binding to DNA without causing DNA damage. It resposents first in class "chromatin damaging" agent without genotoxic properties. Its effect on the transcription in human tumor cells was evaluated. DNA-targeting small molecules are widely used for anticancer therapy based on their ability to induce cell death, presumably via DNA damage. DNA in the eukaryotic cell is packed into chromatin, a highly-ordered complex of DNA, histones, and non-histone proteins. These agents perturb chromatin organization. However, the mechanisms, consequences, and impact of the alterations of chromatin structure in relation to their anti-cancer activity is unclear because it is difficult to separate DNA damage and chromatin damage in cells. We recently demonstrated that curaxins, small molecules with broad anticancer activity, bind DNA without causing detectable DNA damage by interfering with histone/DNA interactions and destabilizing the nucleosome. Chromatin unfolding caused by curaxins is sensed by histone chaperone FACT. FACT binds unfolded nucleosomes, which leads to chromatin trapping or c-trapping. In this study, we investigated whether other DNA-targeting small molecules disturb chromatin and cause c-trapping. We found that only compounds directly binding DNA induce chromatin damage and c-trapping. Chromatin damage may occur in the absence of DNA damage and is dependent on the mechanism of compound binding to DNA and its ability to bind chromatinized DNA in cells. We show that FACT is sensitive to a plethora of nucleosomes perturbations induced by DNA-binding small molecules, including displacement of the linker histone, eviction of core histones, and accumulation of negative supercoiling. Most importantly, the cytotoxicity of DNA-binding small molecules correlates with their ability to cause chromatin damage , but not DNA damage.

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Effect of DNA damage and chromatin damage on transcription in HT1080 cells with wild type p53 and p53 inhibited by dominant negative mutant

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