eccDNA and structural variants analysis reveals massive genome instability in Arabidopsis epigenetic mutants

The epigenome controls transposable element (TE) mobility and mutants affected in the epigenetic machinery display active TEs associated with extrachromosomal circular DNA (eccDNA). However, the interplay between eccDNA and genome stability is poorly understood. Here we show that Arabidopsis plants combining mutations in chromatin remodeling, post-transcriptional gene silencing and RNA-directed DNA methylation have a high eccDNA load associated with integration of truncated TEs and genome instability. We analyzed the eccDNA and genome sequence of ddm1 rdr6 pol4 (Decrease DNA methylation 1, RNA dependent RNA polymerase 6, RNA polymerase 4) triple mutant plants and uncovered TE mobility of full length and truncated TEs. Additionally, TE movement was associated with gene movement of a disease resistance cluster named RPP5. We further discovered a large 2 Mbp inversion and show that this inversion is also present in ddm1 single mutant plants, probably since its isolation 30 years ago. Finally, long read sequencing allowed the detection of two independent ~55 kb duplications in ddm1 siblings. Our results highlight the role of the epigenome in protecting the genome not only against TE mobility but also against chaotic genome rearrangements and eccDNA-driven gene chimerism. We propose that DDM1, on top of its indirect role on DNA methylation and TE control, has a role in protecting the genome against genomic rearrangements.