Mechanisms of DNA methyltransferase 3A1-mediated DNA methylation of nucleosomes

DNA methyltransferase 3A1 (DNMT3A1) plays a crucial role in establishing DNA methylation patterns that regulate gene expression and drive cellular differentiation. In this study, we investigated the biochemical mechanisms underlying DNMT3A1's interactions with nucleosomes, the fundamental units of chromatin. Using radiochemical activity assays, along with fluorescence anisotropy and AlphaLISA binding assays, we demonstrated that DNMT3A1 has multivalent interactions with nucleosomes, binding linker DNA and nucleosome cores. Nanopore-based 5mC sequencing revealed that DNMT3A1 interactions with the nucleosome stimulate methylation of linker DNA up to 24 bp away from the nucleosome core. Additionally, our results indicate that DNMT3A1 binding is restricted to a single nucleosome, suggesting that its activity is not allosterically regulated by unattached nucleosomes. Together, these findings provide new mechanistic insights into how DNMT3A1 engages nucleosomes.