Abstract
Flowering plants utilize small RNA (sRNA) molecules to guide DNA methyltransferases to genomic sequences. This RNA-directed DNA methylation (RdDM) pathway preferentially targets euchromatic transposable elements. However, RdDM is thought to be recruited by methylation of histone H3 at lysine 9 (H3K9me), a hallmark of heterochromatin. How RdDM is targeted to euchromatin despite an affinity for H3K9me is unclear. Here, we show that loss of histone H1 enhances heterochromatic RdDM, preferentially at nucleosome linker DNA. Surprisingly, this does not require SHH1, the RdDM component that binds H3K9me. Furthermore, H3K9me is dispensable for RdDM, as is CG DNA methylation. Instead, we find that non-CG methylation is specifically associated with sRNA biogenesis, and without H1 sRNA production quantitatively expands to non-CG-methylated loci. Our results demonstrate that H1 enforces the separation of euchromatic and heterochromatic DNA methylation pathways by excluding the sRNA-generating branch of RdDM from non-CG-methylated heterochromatin.
Highlights
Transposable elements (TEs) and their remnants comprise a substantial fraction of eukaryotic genomes and generally must be kept silent to ensure genome integrity and function (Bourque et al, 2018)
We show that linker histone H1 impedes RNA-directed DNA methylation (RdDM) activity in GC-rich heterochromatin, thereby restricting RdDM to AT-rich euchromatic TE
Small RNA biogenesis is associated with H3K9me and non-CG methylation Because H3K9me is thought to recruit polymerase II derivative (Pol IV) activity (Law et al, 2013; Zhang et al, 2013), we investigated how small RNAs (sRNA) distribution changes in relation to H3K9me1/2 in h1 plants
Summary
Transposable elements (TEs) and their remnants comprise a substantial fraction of eukaryotic genomes and generally must be kept silent to ensure genome integrity and function (Bourque et al, 2018). In contrast to the SUVH/CMT feedback loop that predominates in heterochromatin, RNA-directed DNA methylation (RdDM) catalyzes cytosine methylation within euchromatic TEs (Zemach et al, 2013; Zhong et al, 2012). A more paradoxical feedback loop is thought to involve recruitment of Pol IV to H3K9me (Erdmann and Picard, 2020; Matzke and Mosher, 2014; Raju et al, 2019; Wendte and Pikaard, 2017) This hypothesis emerged from the observation that Pol IV-mediated sRNA production at many loci requires SHH1/DTF1, a protein that binds H3K9me and monomethylated H3K9me (H3K9me1) in vitro (Law et al., 2013; Zhang et al, 2013). The affinity of H1 for GC-rich heterochromatin (Choi et al, 2020) focuses RdDM activity on short, AT-rich euchromatic TEs that RdDM is uniquely suited to silence
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