The role of DDM1 and DNMTs in DNA Methylation and genome regulation in Physcomitrella patens

Cytosine methylation, is established and maintained by DNA methyltransferases (DNMTs) as well as by chromatin remodelers as DDM1.

In the basal land plant Physcomitrella patens, de-novo methylation at CG and CHH is mediated by DNA Methyltransferase3-b (PpDNMT3b), and CHG by CHROMOMETHYLASE ( PpCMT), which is independent of RdDM pathway. Maintenance of methylation at CG, CHG, and CHH sites is mediated by MET, CMT, and DNMT3b respectively. Whereas in flowering plants, DDM1 mediates methylation in heterochromatin, necessary for silencing transposons and for proper development, in early terrestrial plants its function remains elusive. Here we show that ddm1 knockout in P. patens, disrupts largely any of its sequence contexts. Symmetric CG and CHG sequences were affected more than asymmetric CHH sites. Furthermore, despite their separate targeting mechanisms, CG (MET) and CHG (CMT), methylation was similarly depleted by about 75%. CHH (DNMT3) methylation was overall reduced by about 25%, with an evident hyper-methylation activity within lowly-methylated euchromatic transposon sequences. Despite the strong hypomethylation effect, only a few transposons were transcriptionally activated in Ppddm1. Yet, Ppddm1 develops normally. We concluded that: a) DNA methylation is strongly dependent on DDM1 in a non-flowering plant, b) PpDDM1 is required for plant-DNMT3 (CHH) methylases, though to a lower extent than for MET1 and CMT enzymes, c) distinct and separate methylation pathways (e.g. MET1-CG and CMT-CHG), can be equally regulated by the chromatin, and that DDM1 plays a role in it, d) the biological significance of DDM1 in terms of transposon regulation and plant development, is species dependent