Lysine Methyltransferase Inhibitors Impair H4K20me2 and 53BP1 Foci in Response to DNA Damage in Sarcomas, a Synthetic Lethality Strategy.

Ignacio Campillo-Marcos,Eva Monte-Serrano,Raúl García-González,Pedro A Lazo,Elena Navarro-Carrasco

doi:10.3389/fcell.2021.715126

Abstract

BackgroundChromatin is dynamically remodeled to adapt to all DNA-related processes, including DNA damage responses (DDR). This adaptation requires DNA and histone epigenetic modifications, which are mediated by several types of enzymes; among them are lysine methyltransferases (KMTs).MethodsKMT inhibitors, chaetocin and tazemetostat (TZM), were used to study their role in the DDR induced by ionizing radiation or doxorubicin in two human sarcoma cells lines. The effect of these KMT inhibitors was tested by the analysis of chromatin epigenetic modifications, H4K16ac and H4K20me2. DDR was monitored by the formation of γH2AX, MDC1, NBS1 and 53BP1 foci, and the induction of apoptosis.ResultsChaetocin and tazemetostat treatments caused a significant increase of H4K16 acetylation, associated with chromatin relaxation, and increased DNA damage, detected by the labeling of free DNA-ends. These inhibitors significantly reduced H4K20 dimethylation levels in response to DNA damage and impaired the recruitment of 53BP1, but not of MDC1 and NBS1, at DNA damaged sites. This modification of epigenetic marks prevents DNA repair by the NHEJ pathway and leads to cell death.ConclusionKMT inhibitors could function as sensitizers to DNA damage-based therapies and be used in novel synthetic lethality strategies for sarcoma treatment.

Highlights

Chromatin structure is dynamically remodeled by DNA and histone epigenetic modifications to control and coordinate all DNA-based processes such as transcription, replication, recombination or DNA repair (Becker and Workman, 2013; Hauer and Gasser, 2017)
We have studied the molecular base by which KMT inhibitors, chaetocin and tazemetostat, impair DNA damage responses (DDR) (32, 52) by mimicking a DNA repair defect, which would allow their use as DNA damage sensitizers (53, 54) and become candidates for novel synthetic lethality strategies in sarcomas cells treated with either ionizing radiation (IR) (Lee et al, 2013) or doxorubicin (Maurel et al, 2009; D’Ambrosio et al, 2020)
No differences in 53BP1 protein levels were detected by western blot after chaetocin and/or IR treatments (Figure 1D), which rules out a reduction of endogenous 53BP1 protein as the cause of foci loss

Summary

Introduction

Chromatin structure is dynamically remodeled by DNA and histone epigenetic modifications to control and coordinate all DNA-based processes such as transcription, replication, recombination or DNA repair (Becker and Workman, 2013; Hauer and Gasser, 2017). In addition to its role as KMT inhibitor (Greiner et al, 2005), chaetocin is a competitive inhibitor of the thioredoxin reductase, whose function is to compensate the deleterious effect of reactive oxygen species (ROS) (Tibodeau et al, 2009) Other exogenous agents such as ionizing radiation facilitate ROS production (Jackson and Bartek, 2009; Ciccia and Elledge, 2010), causing single- and double-strand breaks (SSBs and DSBs, respectively) (Ciccia and Elledge, 2010), and their accumulation can lead to cell death (Ciccia and Elledge, 2010). This adaptation requires DNA and histone epigenetic modifications, which are mediated by several types of enzymes; among them are lysine methyltransferases (KMTs)

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Discussion

Conclusion