Significance: Changes in the methylation status of cytosine nucleotides are a source of heritable epigenetic and phenotypic diversity in plants. Here we derive robust estimates of the rate at which cytosine methylation is spontaneously gained (forward epimutation) or lost (backward epimutation) in the genome of the model plant Arabidopsis thaliana . We show that the forward–backward dynamics of selectively neutral epimutations have a major impact on methylome evolution and shape genome-wide patterns of methylation diversity among natural populations in this species. The epimutation rates presented here can serve as reference values in future empirical and theoretical population epigenetic studies in plants.

Epigenetics has emerged as an important research field for crop improvement under the on-going climatic changes. Heritable epigenetic changes can arise independently of DNA sequence alterations and have been associated with altered gene expression and transmitted phenotypic variation. By modulating plant development and physiological responses to environmental conditions, epigenetic diversity—naturally, genetically, chemically, or environmentally induced—can help optimise crop traits in an era challenged by global climate change. Beyond DNA sequence variation, the epigenetic modifications may contribute to breeding by providing useful markers and allowing the use of epigenome diversity to predict plant performance and increase final crop production. Given the difficulties in transferring the knowledge of the epigenetic mechanisms from model plants to crops, various strategies have emerged. Among those strategies are modelling frameworks dedicated to predicting epigenetically controlled-adaptive traits, the use of epigenetics for in vitro regeneration to accelerate crop breeding, and changes of specific epigenetic marks that modulate gene expression of traits of interest. The key challenge that agriculture faces in the 21st century is to increase crop production by speeding up the breeding of resilient crop species. Therefore, epigenetics provides fundamental molecular information with potential direct applications in crop enhancement, tolerance, and adaptation within the context of climate change.

Despite major progress in dissecting the molecular pathways that control DNA methylation patterns in plants, little is known about the mechanisms that shape plant methylomes over evolutionary time. Drawing on recent intra- and interspecific epigenomic studies, we show that methylome evolution over long timescales is largely a byproduct of genomic changes. By contrast, methylome evolution over short timescales appears to be driven mainly by spontaneous epimutational events. We argue that novel methods based on analyses of the methylation site frequency spectrum (mSFS) of natural populations can provide deeper insights into the evolutionary forces that act at each timescale.

Contents Summary 1253 I. Introduction 1253 II. What is the rate and molecular spectrum of spontaneous epimutations? 1254 III. Do spontaneous epimutations have phenotypic consequences? 1257 IV. Conclusion and discussion 1258 Acknowledgements 1258 References 1258 Summary: Heritable gains or losses of cytosine methylation can arise stochastically in plant genomes independently of DNA sequence changes. These so‐called ‘spontaneous epimutations’ appear to be a byproduct of imperfect DNA methylation maintenance and epigenome reinforcement events that occur in specialized cell types. There is continued interest in the plant epigenetics community in trying to understand the broader implications of these stochastic events, as some have been shown to induce heritable gene expression changes, shape patterns of methylation diversity within and among plant populations, and appear to be responsive to multi‐generational environmental stressors. In this paper we synthesized our current knowledge of the molecular basis and functional consequences of spontaneous epimutations in plants, discuss technical and conceptual challenges, and highlight emerging research directions.

Background: Plants can transmit somatic mutations and epimutations to offspring, which in turn can affect fitness. Knowledge of the rate at which these variations arise is necessary to understand how plant development contributes to local adaption in an ecoevolutionary context, particularly in long-lived perennials.Results: Here, we generate a new high-quality reference genome from the oldest branch of a wildPopulus trichocarpatree with two dominant stems which have been evolving independently for 330 years. By sampling multiple, age-estimated branches of this tree, we use a multi-omics approach to quantify age-related somatic changes at the genetic, epigenetic, and transcriptional level. We show that the per-year somatic mutation and epimutation rates are lower than in annuals and that transcriptional variation is mainly independent of age divergence and cytosine methylation. Furthermore, a detailed analysis of the somatic epimutation spectrum indicates that transgenerationally heritable epimutations originate mainly from DNA methylation maintenance errors during mitotic rather than during meiotic cell divisions.Conclusion: Taken together, our study provides unprecedented insights into the origin of nucleotide and functional variation in a long-lived perennial plant.

Despite the importance and wide exploitation of heterosis in commercial crop breeding, the molecular mechanisms behind this phenomenon are not completely understood. Recent studies have implicated changes in DNA methylation and small RNAs in hybrid performance; however, it remains unclear whether epigenetic changes are a cause or a consequence of heterosis. Here, we analyze a large panel of over 500 Arabidopsis (Arabidopsis thaliana) epigenetic hybrid plants (epiHybrids), which we derived from near-isogenic but epigenetically divergent parents. This proof-of-principle experimental system allowed us to quantify the contribution of parental methylation differences to heterosis. We measured traits such as leaf area, growth rate, flowering time, main stem branching, rosette branching, and final plant height and observed several strong positive and negative heterotic phenotypes among the epiHybrids. Using an epigenetic quantitative trait locus mapping approach, we were able to identify specific differentially methylated regions in the parental genomes that are associated with hybrid performance. Sequencing of methylomes, transcriptomes, and genomes of selected parent-epiHybrid combinations further showed that these parental differentially methylated regions most likely mediate the remodeling of methylation and transcriptional states at specific loci in the hybrids. Taken together, our data suggest that locus-specific epigenetic divergence between the parental lines can directly or indirectly trigger heterosis in Arabidopsis hybrids independent of genetic changes. These results add to a growing body of evidence that points to epigenetic factors as one of the key determinants of hybrid performance.