A night shift for histone methylation in DNA damage control

Abstract

Plants fine-tune their physiology to the time of day, largely through dynamic shifts in gene expression. While these shifts are generally attributed to transcription factor activity, an additional layer of regulation comes from chromatin modifications. Covalent histone modifications, collectively referred to as the ‘histone code’, affect chromatin structure and recruitment of regulatory proteins and thereby determine transcriptional activity.

Histone marks show distinct links to diurnal and circadian rhythms in plants. In the model plant Arabidopsis thaliana, signatures of Histone 3 acetylation at Lysine residue 9 (H3K9ac) and 27 (H3K27ac) and phosphorylation at Serine residue 28 (H3S28p) vary between day and night (Baerenfaller et al., 2016). Additionally, many components of the circadian clock are regulated at the chromatin level, particularly through histone acetylation (Xiong et al., 2022). Histone acetylation is generally associated with gene activation, while histone methylation can either activate or repress gene expression, depending on the site of modification (Liu et al., 2010). For example, Histone H3 monomethylation at Lysine residue 27 (H3K27me1) is associated with switched off genes: It plays a crucial role in constitutive silencing of transposable elements and contributes to the maintenance of heterochromatin and the low expression of some genes within euchromatin (Jacob et al., 2010; Potok et al., 2022). However, it remained unknown whether H3K27me1 deposition follows diurnal patterns, and how such patterns affect gene function.

Crisanto Gutierrez's lab has substantially advanced our understanding of chromatin dynamics, especially in regard to cell division and genome replication. Recently, the lab turned its focus to exploring the effects of chromatin changes on gene expression and their impact on plant development and environmental sensing. Jorge Fung-Uceda, co-first author of the highlighted study, began his work on chromatin dynamics and the circadian clock during his PhD before joining Gutierrez's lab as a postdoctoral researcher to study H3K27me1's role in gene regulation. He was joined on the project by co-first author María Sol Gomez, who brought in expertise in plant stress responses and environmental perception.

Fung-Uceda et al. observed that H3K27me1 levels fluctuate with the time of day, with higher levels at night than during the day (Figure 1a), and that this difference was more pronounced under short-day conditions than under long-day conditions. H3K27me1 is deposited by the methyl transferases ARABIDOPSIS TRITHORAX-RELATED PROTEIN 5 (ATXR5) and ATXR6 (Jacob et al., 2009). In agreement with increased H3K27me1 levels, transcript levels of ATXR5 peaked during the night, while ATXR6 transcript levels remained low throughout the 24-h period (Figure 1b). Whether these oscillations are controlled by the circadian clock is unclear, but the presence of two circadian clock binding sites in the ATXR5 promoter suggests a possible regulatory link.

A full knock-out of both ATXR5 and ATXR6 is lethal (Potok et al., 2022), but a hypomorphic atxr5 atxr6 double mutant displays reduced leaf and rosette size (Jacob et al., 2009). Fung-Uceda et al. found that this phenotype is specific to short-day conditions, aligning with the larger fluctuations in H3K27me1. While H3K27me1 deposition in the atxr5 atxr6 mutant was significantly reduced both at midday and midnight, more genes were differentially expressed at night. Notably, genes with decreased H3K27me1 levels were largely more highly expressed, whereas most genes with lower expression showed no change in H3K27me1 levels and are likely not direct ATXR5/6 targets. These observations support the role of H3K27me1 as a repressive mark, with a more prominent role at night-time.

Genes more highly expressed in atxr5 atxr6 were enriched for those involved in cell cycle control and DNA damage repair (DDR), and many showed a marked drop in the H3K27me1 signal across their gene body, but not at the transcriptional start site (Figure 1c). Interestingly, many DDR genes also exhibited rhythmic expression patterns in atxr5 atxr6. Expression of core circadian clock genes remained unchanged in atxr5 atxr6, suggesting that DDR gene activation is directly diurnally gated by H3K27me1 deposition. To test whether the responsiveness to DNA damage varied by time of day, wild-type and atxr5 atxr6 plants were treated with bleomycin, a genotoxic compound that causes DNA double strand breaks. In the wild type, bleomycin-induced DDR gene expression was highest at night, while its pattern and magnitude in atxr5 atxr6 remained unaffected by bleomycin treatment.

Taken together, the findings by Fung-Uceda et al. imply that the response to DNA damage varies with time of day, and that this effect is mediated by H3K27me1. The physiological relevance of this gating mechanism is currently unclear. Fung-Uceda et al. propose that H3K27me1 helps synchronise DDR activity with cellular processes such as DNA replication, during which DDR gene function might be crucial to maintain genome integrity. H3K27me1 might thus function as a ‘repressive switch’ that prevents unnecessary induction of DDR genes at times when the risk of DNA damage is low. Moving forward, Crisanto Gutierrez plans to further investigate the link between diurnal control of genome replication and DDR, potentially providing evidence to support the long-standing hypothesis that the cell cycle is diurnally gated to avoid DNA damage during times of high light exposure.