Rate, spectrum, and evolutionary dynamics of spontaneous epimutations

Abstract

Stochastic changes in cytosine methylation are a source of heritable epigenetic and phenotypic diversity in plants. Using the model plant Arabidopsis thaliana, we derive robust estimates of the rate at which methylation is spontaneously gained (forward epimutation) or lost (backward epimutation) at individual cytosines and construct a comprehensive picture of the epimutation landscape in this species. We demonstrate that the dynamic interplay between forward and backward epimutations is modulated by genomic context and show that subtle contextual differences have profoundly shaped patterns of methylation diversity in A. thaliana natural populations over evolutionary timescales. Theoretical arguments indicate that the epimutation rates reported here are high enough to rapidly uncouple genetic from epigenetic variation, but low enough for new epialleles to sustain long-term selection responses. Our results provide new insights into methylome evolution and its population-level consequences.epigenetics | epimutation | DNA methylation | evolution | Arabidopsis P lant genomes make extensive use of cytosine methylation to control the expression of transposable elements (TEs) and genes (1). Despite its tight regulation, methylation losses or gains at individual cytosines or clusters of cytosines can emerge spontaneously, in an event termed "epimutation" (2, 3). Many examples of segregating epimutations have been documented in experimental and wild populations of plants and in some cases contribute to heritable variation in phenotypes independently of DNA sequence variation (4, 5). These observations have led to much speculation about the role of DNA methylation in plant evolution (6-8), and its potential in breeding programs (9). In the model plant Arabidopsis thaliana, spontaneous methylation changes at CG dinucleotides accumulate in a rapid but nonlinear fashion over generations (2,3,10), thus pointing to high forward-backward epimutation rates (11). Precise estimates of these rates are necessary to be able to quantify the long-term dynamics of epigenetic variation under laboratory or natural conditions, and to understand the molecular mechanisms that drive methylome evolution (12-14). Here we combine theoretical modeling with high-resolution methylome analysis of multiple independent A. thaliana mutation accumulation (MA) lines (15), including measurements of methylation changes in continuous generations, to obtain robust estimates of forward and backward epimutation rates. ResultsWe joined whole-genome MethylC-seq (16) data from two earlier MA studies (2, 3) with extensive multigenerational MethylC-seq measurements from three additional MA lines (Fig. 1A and SI Appendix, Tables S1-S6). The first of these new MA lines (MA1 3) was propagated for 30 generations and includes measurements for 13 (nearly) consecutive generations (Fig. 1A). The other two MA lines (MA2 3) were propagated for 17 generations and were measured every four generations on average (Fig. 1A). These new data therefore allowed us to track epimutation...

Methylome evolution in plants

“…An intriguing observation, however, is that heritable alterations in DNA methylation states can also emerge spontaneously and independently of genetic mutations [8, 57, 76–78, 109–113]. The most comprehensive demonstration of this phenomenon has come from the analysis of multi-generational methylome data from A. thaliana mutation accumulation lines (MA-lines) [76–78, 112]. Such lines are derived from a single founder plant (of the Columbia accession) and independently propagated for a large number of generations [114].…”

“…Such lines are derived from a single founder plant (of the Columbia accession) and independently propagated for a large number of generations [114]. Detailed comparisons of the methylomes of MA-lines have been instrumental in providing the first estimates of the rate at which epimutations occur in plant genomes [76–78]. Efforts are now underway to try to understand the extent to which spontaneous epimutations contribute to methylome diversity in natural populations over short timescales up to thousands of generations.

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“…The meiotic inheritance of epimutations, however, appears to be mainly restricted to CG dinucleotides [76–78], probably as a result of context-specific methylation resetting during gametogenesis and early development [115]. Estimates in MA-lines indicate that the rate of forward epimutations (i.e., stochastic gains of methylation) is about 2.56 × 10 −4 per CG site per haploid genome per generation, while the rate of backward epimutations (i.e., stochastic loss of methylation) is about 6.3 × 10 −4 [78]. Hence, methylation loss is globally about 2.5 times as likely as methylation gain.…”

“…Even after only 30 generations of independent selfing, the methylomes of early-generation and late-generation MA-lines can be clearly distinguished. As the methylation status of individual CG sites is simultaneously subject to both forward and backward epimutations, methylome divergence does not increase linearly over time [72, 78, 117] but saturates rather quickly to some equilibrium divergence value (Fig. 3).…”

“…3). On the basis of estimates from Van der Graaf et al [78], only about 4000 generations would be needed in a hypothetical mutation accumulation experiment for methylome divergence to be within 99% of this value. This insight leads to the evolutionary prediction that epimutational processes should dominate methylome diversity in the early stages of lineage divergence but only marginally at later stages.…”