Estrogen and DNA damage modulate mRNA levels of genes involved in homologous recombination repair in estrogen-deprived cells

Abstract

Aims: Breast and ovarian cancers are frequently associated with mutations in genes involved in DNA double-strand break (DSB) repair by homologous recombination (HR). Risk factors for breast cancer are often linked to estrogen-related pathways. Here, we studied the crosslink between estrogen and the HR pathway.

Methods: We analyzed, using online annotation tolls, the enrichment of candidate estrogen-upregulated genes among DNA repair pathways. We analyzed how estrogen modulates mRNA levels of HR repair (HRR) genes in estrogen-receptor (ER)-positive cells. The cells were deprived of estrogen, and the mRNA levels of HRR genes were determined using real-time polymerase chain reaction, following estrogen addition as well as DNA damage induction. In addition, we examined the effect of estrogen on DNA repair, by immuno-fluorescence analysis, using the DSB marker phospho-histone H2AX, as an indicator for DSB repair. Finally, we performed a clonogenic survival assay to determine the effect of estrogen on cell survival.

Results: We discovered that genes whose mRNA levels are upregulated by estrogen are strongly associated with the HR pathway. We validated that estrogen upregulates mRNA levels of the HRR genes MRE11, RAD50, and PALB2, which have not been previously shown to be regulated by estrogen. Additionally, we revealed that DNA damage induces an upsurge in mRNAs encoding BRCA1, MRE11, RAD50, PALB2, and CtIP, in ER-positive cells deprived of estrogen. Notably, DSB repair was impaired in ER-positive cells deprived of estrogen, compared to cells exposed to the hormone. We also established that ER-positive cells deprived of estrogen are hypersensitive to DSBs.

Conclusion: These results suggest that exposure of ER-positive cells to estrogen triggers the expression of HRR genes, which is required to meet the increased repair demands due to the proliferating effect induced by estrogen. This may explain the higher chances of developing estrogen-dependent cancers due to mutations in HRR genes.

Keywords

DNA double-strand breaks (DSBs) / DSB repair / homologous recombination repair / estrogen / MRE11 / RAD50 / CtIP

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