An epigenetic cytosine oxidative layer in Silene latifolia
DNA methylation and its oxidative derivatives are key components of epigenetic regulation, yet their roles in plant repetitive DNA remain poorly understood. Here, we investigate the genomic distribution of oxidized cytosine modifications: 5-hydroxy-methyl-cytosine (5hmC), 5-formyl-cytosine (5fC) and 5-carboxy-cytosine (5caC) within the repeat-rich genome of Silene latifolia, a dioecious plant with heteromorphic sex chromosomes. Using low-coverage oxi-mC sequencing combined with RepeatExplorer2 clustering, ChIP-seq Mapper analysis, genome mapping, and immunoFISH validation, we identify widespread association of oxi-mCs across major repeat classes, including LTR retrotransposons, DNA transposons, satellite repeats, and ribosomal DNA (rDNA). While transposable elements largely reflect genomic abundance, satellite DNA shows selective and context-dependent enrichment, particularly the centromeric repeat STAR-C, which exhibits association with 5caC in specific chromosome-biased clusters rather than across centromeres globally. In contrast, 45S rDNA arrays display strong and consistent enrichment of oxi-mCs, especially 5fC and 5caC, in transcriptionally active nucleolar organizing regions, supported by both sequencing and cytological evidence. These findings suggest that oxidative cytosine modifications are not randomly distributed but are linked to chromatin state and transcriptional activity, particularly in rDNA loci. Overall, our results reveal cytosine oxidation as an additional layer of epigenetic regulation in plant repetitive DNA.
DNA methylation and its oxidative derivatives are key components of epigenetic regulation, yet their roles in plant repetitive DNA remain poorly understood. Here, we investigate the genomic distribution of oxidized cytosine modifications: 5-hydroxy-methyl-cytosine (5hmC), 5-formyl-cytosine (5fC) and 5-carboxy-cytosine (5caC) within the repeat-rich genome of Silene latifolia, a dioecious plant with heteromorphic sex chromosomes. Using low-coverage oxi-mC sequencing combined with RepeatExplorer2 clustering, ChIP-seq Mapper analysis, genome mapping, and immunoFISH validation, we identify widespread association of oxi-mCs across major repeat classes, including LTR retrotransposons, DNA transposons, satellite repeats, and ribosomal DNA (rDNA). While transposable elements largely reflect genomic abundance, satellite DNA shows selective and context-dependent enrichment, particularly the centromeric repeat STAR-C, which exhibits association with 5caC in specific chromosome-biased clusters rather than across centromeres globally. In contrast, 45S rDNA arrays display strong and consistent enrichment of oxi-mCs, especially 5fC and 5caC, in transcriptionally active nucleolar organizing regions, supported by both sequencing and cytological evidence. These findings suggest that oxidative cytosine modifications are not randomly distributed but are linked to chromatin state and transcriptional activity, particularly in rDNA loci. Overall, our results reveal cytosine oxidation as an additional layer of epigenetic regulation in plant repetitive DNA.
Institute of Biophysics Academy of Sciences of the Czech Republic