Dual control of ROS1-mediated active DNA demethylation by DNA damage-binding protein 2 (DDB2).

Dolores Córdoba‐Cañero,Valérie Cognat,Jean Molinier,Teresa Roldán Arjona,Rafael R Ariza

doi:10.1111/tpj.13753

Abstract

By controlling gene expression, DNA methylation contributes to key regulatory processes during plant development. Genomic methylation patterns are dynamic and must be properly maintained and/or re-established upon DNA replication and active removal, and therefore require sophisticated control mechanisms. Here we identify direct interplay between the DNA repair factor DNA damage-binding protein 2 (DDB2) and the ROS1-mediated active DNA demethylation pathway in Arabidopsis thaliana. We show that DDB2 forms a complex with ROS1 and AGO4 and that they act at the ROS1 locus to modulate levels of DNA methylation and therefore ROS1 expression. We found that DDB2 represses enzymatic activity of ROS1. DNA demethylation intermediates generated by ROS1 are processed by the DNA 3'-phosphatase ZDP and the apurinic/apyrimidinic endonuclease APE1L, and we also show that DDB2 interacts with both enzymes and stimulates their activities. Taken together, our results indicate that DDB2 acts as a critical regulator of ROS1-mediated active DNA demethylation.

Highlights

As sessile organisms, plants are exposed to the deleterious effects of environmental stress, which can affect integrity of the genome and epigenome by inducing DNA damage and chromatin alterations (Pecinka and Mittelsten Scheid, 2012)
Upregulation of repressor of silencing 1 (ROS1) was observed in plants defective for the expression of the global genome repair (GGR) factor DNA damage-binding protein 2 (DDB2) (Schalk et al, 2016), suggesting that this DNA repair protein acts, directly or indirectly, as a repressor of ROS1 expression
DDB2 interconnects with the RNA-directed DNA methylation (RdDM) pathway via its interaction with AGO4 and represents a material of choice for deciphering the underlying mechanisms of control of ROS1 expression

Summary

Introduction

Plants are exposed to the deleterious effects of environmental stress, which can affect integrity of the genome and epigenome by inducing DNA damage and chromatin alterations (Pecinka and Mittelsten Scheid, 2012). DNA methylation (5-methyl cytosine, 5-meC), a key component of the plant epigenome, contributes to the stable silencing of transposable elements (TEs) as well as to the regulation of gene expression (Law and Jacobsen, 2010). In Arabidopsis, DNA methylation is maintained by four main DNA methyltransferases: methyltransferase 1 (MET1), chromomethylase 3 (CMT3), chromomethylase 2 (CMT2) and domains rearranged methyltransferase 2 (DRM2) acting in the CG, CHG and CHH sequence contexts (where H = A, T or C), respectively (Law and Jacobsen, 2010; Zemach et al, 2013). Cytosines can be methylated de novo through RNA-directed DNA methylation (RdDM) involving different classes of small RNA (Cuerda-Gil and Slotkin, 2016). RdDM transcriptionally represses TEs and is involved in heterochromatin formation (Matzke and Mosher, 2014)

Results

Discussion

Conclusion