De novo induction of a DNA–histone H3K9 methylation loop on synthetic human repetitive DNA in cultured tobacco cells

• Published in: The Plant journal : for cell and molecular biology 2023-05, Vol.114 (3), p.668-682
• Main Authors: Otake, Koichiro, Kugou, Kazuto, Robertlee, Jekson, Ohzeki, Jun‐ichirou, Okazaki, Koei, Hanano, Shigeru, Takahashi, Seiji, Shibata, Daisuke, Masumoto, Hiroshi
• Format: Article
• Language: English
• Subjects: Arabidopsis - genetics; Arabidopsis - metabolism; Arabidopsis Proteins - genetics; Arabidopsis Proteins - metabolism; Bright plating; BY‐2; Cell culture; CENP‐B; Centromere - metabolism; Chromatin - genetics; Chromatin - metabolism; chromatin manipulation; Chromatin remodeling; Deoxyribonucleic acid; DNA; DNA - metabolism; DNA methylation; DNA Methylation - genetics; DNA methyltransferase; DRM1; Effectors; Epigenetics; H3K9 di‐methylation; Histone methyltransferase; Histones; Histones - genetics; Histones - metabolism; Humans; Nicotiana - genetics; Nicotiana - metabolism; Plant cells; Progeny; SUVH9; synthetic α‐satellite DNA; Tethering; tetR; Tobacco
• ISSN: 0960-7412; 1365-313X
• DOI: 10.1111/tpj.16164

SUMMARY Genetic modifications in plants are crucial tools for fundamental and applied research. Transgene expression usually varies among independent lines or their progeny and is associated with the chromatin structure of the insertion site. Strategies based on understanding how to manipulate the epigenetic state of the inserted gene cassette would help to ensure transgene expression. Here, we report a strategy for chromatin manipulation by the artificial tethering of epigenetic effectors to a synthetic human centromeric repetitive DNA (alphoid DNA) platform in plant Bright‐Yellow‐2 (BY‐2) culture cells. By tethering DNA‐methyltransferase (Nicotiana tabacum DRM1), we effectively induced DNA methylation and histone methylation (H3K9me2) on the alphoid DNA platform. Tethering of the Arabidopsis SUVH9, which has been reported to lack histone methyltransferase activity, also induced a similar epigenetic state on the alphoid DNA in BY‐2 cells, presumably by activating the RNA‐dependent DNA methylation (RdDM) pathway. Our results emphasize that the interplay between DNA and histone methylation mechanisms is intrinsic to plant cells. We also found that once epigenetic modification states were induced by the tethering of either DRM1 or SUVH9, the modification was maintained even when the direct tethering of the effector was inhibited. Our system enables the analysis of more diverse epigenetic effectors and will help to elucidate the chromatin assembly mechanisms of plant cells. Significance Statement By tethering DNA‐methyltransferase (Nicotiana tabacum DRM1), we effectively induced DNA methylation and histone methylation (H3K9me2) on a synthetic human centromeric repetitive DNA (alphoid DNA) platform in tobacco BY‐2 culture cells. The tethering of Arabidopsis SUVH9, which has been reported to lack histone methyltransferase activity in vitro, also induced a similar epigenetic state on the alphoid DNA. Our results emphasize that the interplay between DNA and histone methylation mechanisms is intrinsic to plant cells.