ABSTRACT
DNA methylation is an important epigenetic mark implicated in selective rRNA gene expression, but the DNA methylation readers and effectors remain largely unknown. Here, we report a protein complex that reads DNA methylation to regulate variant-specific 45S ribosomal RNA (rRNA) gene expression in Arabidopsis (Arabidopsis thaliana). The complex, consisting of METHYL-CpG-BINDING DOMAIN PROTEIN5 (MBD5), MBD6, ALPHA-CRYSTALLIN DOMAIN PROTEIN15.5 (ACD15.5), and ACD21.4, directly binds to 45S rDNA. While MBD5 and MBD6 function redundantly, ACD15.5 and ACD21.4 are indispensable for variant-specific rRNA gene expression. These 4 proteins undergo phase separation in vitro and in vivo and are interdependent for their phase separation. The α-crystallin domain of ACD15.5 and ACD21.4, which is essential for their function, enables phase separation of the complex, likely by mediating multivalent protein interactions. The effector MICRORCHIDIA6 directly interacts with ACD15.5 and ACD21.4, but not with MBD5 and MBD6, and is recruited to 45S rDNA by the MBD-ACD complex to regulate variant-specific 45S rRNA expression. Our study reveals a pathway in Arabidopsis through which certain 45S rRNA gene variants are silenced, while others are activated.
Subject(s)
Arabidopsis Proteins , Arabidopsis , alpha-Crystallins , Arabidopsis/genetics , Arabidopsis/metabolism , Genes, rRNA , DNA Methylation/genetics , RNA, Ribosomal/genetics , RNA, Ribosomal/metabolism , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , DNA, Ribosomal/genetics , DNA, Ribosomal/metabolism , alpha-Crystallins/genetics , alpha-Crystallins/metabolismABSTRACT
The R-loop, composed of a DNA-RNA hybrid and the displaced single-stranded DNA, regulates diverse cellular processes. However, how cellular R-loops are recognized remains poorly understood. Here, we report the discovery of the evolutionally conserved ALBA proteins (AtALBA1 and AtALBA2) functioning as the genic R-loop readers in Arabidopsis. While AtALBA1 binds to the DNA-RNA hybrid, AtALBA2 associates with single-stranded DNA in the R-loops in vitro. In vivo, these two proteins interact and colocalize in the nucleus, where they preferentially bind to genic regions with active epigenetic marks in an R-loop-dependent manner. Depletion of AtALBA1 or AtALBA2 results in hypersensitivity of plants to DNA damaging agents. The formation of DNA breaks in alba mutants originates from unprotected R-loops. Our results reveal that the AtALBA1 and AtALBA2 protein complex is the genic R-loop reader crucial for genome stability in Arabidopsis.