DNA demethylase ROS1 prevents inheritable DREB1A / CBF3 repression by transgene-induced promoter methylation in the Arab
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DNA demethylase ROS1 prevents inheritable DREB1A/CBF3 repression by transgene‑induced promoter methylation in the Arabidopsis ice1‑1 mutant June‑Sik Kim1 · Satoshi Kidokoro2 · Kazuo Shinozaki1 · Kazuko Yamaguchi‑Shinozaki2,3 Received: 23 April 2020 / Accepted: 23 August 2020 © Springer Nature B.V. 2020
Key message In the ros1-defective mutant, DREB1A repression by the transgene-induced promoter methylation of ice1-1 became inheritable across generations even in the absence of the causative transgene NICE1. Abstract Transgene silencing (TGS) is a widely observed event during plant bioengineering, which is presented as a gradual decrease in ectopic gene expression across generations and occasionally coupled with endogenous gene silencing based on DNA sequence similarity. TGS is known to be established by guided DNA methylation machinery. However, the machinery underlying gene recovery from TGS has not been fully elucidated. We previously reported that in ice1-1 outcross descendants, the expressional repression and recovery of DREB1A/CBF3 were instantly achieved by a newly discovered NICE1 transgene, instead of the formerly proposed ice1-1 mutation in the ICE1 gene. The plants harboring NICE1 produced small RNAs targeting and causing the DREB1A promoter to be hypermethylated and silenced. To analyze the role of the plantspecific active DNA demethylase REPRESSOR OF SILENCING 1 (ROS1) in instant DREB1A recovery, we propagated the NICE1-segregating population upon ros1 dysfunction and evaluated the gene expression and DNA methylation levels of DREB1A through generations. Our results showed that the epigenetic DREB1A repression was substantially sustained in subsequent generations even without NICE1 and stably inherited across generations. Consistent with the gene expression results, only incomplete DNA methylation removal was detected in the same generations. These results indicate that a novel inheritable epiallele emerged by the ros1 dysfunction. Overall, our study reveals the important role of ROS1 in the inheritability of TGS-associated gene repression. Keywords Transgene silencing · DNA methylation · ice1-1 · NICE1 · ROS1 · DREB1A/CBF3
Introduction
Electronic supplementary material The online version of this article (https://doi.org/10.1007/s11103-020-01061-4) contains supplementary material, which is available to authorized users. * Kazuko Yamaguchi‑Shinozaki [email protected]‑tokyo.ac.jp 1
Gene Discovery Research Group, RIKEN Center for Sustainable Resource Science, Tsukuba, Ibaraki 305‑0074, Japan
2
Laboratory of Plant Molecular Physiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Bunkyo‑ku, Tokyo 113‑8657, Japan
3
Research Institute for Agricultural and Life Sciences, Tokyo University of Agriculture, Setagaya‑ku, Tokyo 156‑8502, Japan
Terrestrial plants have evolved complicated stress-responsive gene regulatory networks (GRNs) to survive under harsh environmental conditions, including chilling. Many cold stress-responsive genes have been isolated from
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