Differential methylation of the circular DNA in geminiviral minichromosomes

作者：Deuschle Kathrin; Kepp Gabi; Jeske Holger^*

来源：Virology, 2016, 499: 243-258.

DOI：10.1016/j.virol.2016.09.024

• Geminiviridae
• Begomovirus
• Abutilon mosaic virus
• tomato yellow leaf curl Sardinia virus
• Minichromosome purification
• DNA methylation
• Nicotiana benthamiana
• Ligation-mediated RCA

Geminiviral minichromosomes were purified to explore epigenetic modifications. The levels of methylation in their covalently closed circular DNA were examined with the help of methylation-dependent restriction (MdR). DNA with 12 superhelical turns was preferentially modified, indicating minichromosomes with 12 nucleosomes leaving an open gap. MdR digestion yielded a specific product of genomic length, which was cloned and Sanger-sequenced, or amplified following ligation-mediated rolling circle amplification and deep-sequenced (circomics). The conventional approach revealed a single cleavage product indicating specific methylations at the borders of the common region. The circomics approach identified considerably more MdR sites in a preferential distance to each other of similar to 200 nts, which is the DNA length in a nucleosome. They accumulated in regions of nucleosome-free gaps, but scattered also along the genomic components. These results may hint at a function in specific gene regulation, as well as in virus resistance.

• 出版日期2016-12