RNA-directed DNA methylation prevents rapid and heritable reversal of transposon silencing under heat stress in Zea mays.

In large complex plant genomes, RNA-directed DNA methylation (RdDM) ensures that epigenetic silencing is maintained at the boundary between genes and flanking transposable elements. In maize, RdDM is dependent on Modifer of Paramutation 1 (Mop1), a putative RNA dependent RNA polymerase. Here we show that although RdDM is essential for the maintenance of DNA methylation of a silenced MuDR transposon in maize, a loss of that methylation does not result in a restoration of activity. Instead, heritable maintenance of silencing is maintained by histone modifications. At one terminal inverted repeat (TIR) of this element, heritable silencing is mediated via H3K9 and H3K27 dimethylation, even in the absence of DNA methylation. At the second TIR, heritable silencing is mediated by H3K27 trimethylation, a mark normally associated with somatically inherited gene silencing. We find that a brief exposure of high temperature in a mop1 mutant rapidly reverses both of these modifications in conjunction with a loss of transcriptional silencing. These reversals are heritable, even in mop1 wild type progeny in which methylation is restored at both TIRs. These observations suggest that DNA methylation is neither necessary to maintain silencing, nor is it sufficient to initiate silencing once has been reversed. However, given that heritable reactivation only occurs in a mop1 mutant background, these observations suggest that DNA methylation is required to buffer the effects of environmental stress on transposable elements. Author SummaryMost plant genomes are mostly transposable elements (TEs), most of which are held in check by modifications of both DNA and histones. The bulk of silenced TEs are associated with methylated DNA and histone H3 lysine 9 demethylation (H3K9me2). In contrast, epigenetically silenced genes are often associated with histone lysine 27 trimethylation (H3K27me3). Although stress can affect each of these modifications, plants are generally competent to rapidly reset them following that stress. Here we demonstrate that although DNA methylation is not required to maintain silencing of the MuDR element, it is essential for preventing heat-induced, stable and heritable changes in both H3K9me2 and H3K27me3 at this element, and for concomitant changes in transcriptional activity. These finding suggest that RdDM acts to buffer the effects of heat on silenced transposable elements, and that a loss of DNA methylation under conditions of stress can have profound and long lasting effects on epigenetic silencing in maize.