Arabidopsis AGDP1 links H3K9me2 to DNA methylation in heterochromatin

Heterochromatin is a tightly packed form of DNA, which is associated with histone 3 lysine 9 methylation (H3K9me). Here, we identify an H3K9me2 binding protein, Agenet domain (AGD)-containing protein 1 (AGDP1), in Arabidopsis thaliana. Our biochemical studies revealed that the AGDP1 can specifically recognize the H3K9me2 marks by the three pairs of tandem-AGDs. We determined the crystal structure of the AGD12 of Raphanus sativus AGDP1 in complex with an H3K9me2 peptide. In the complex, the histone peptide adopts a unique helical conformation. AGD12 employs a newly defined interacting interface to specifically recognize the H3K4me0 and H3K9me2 marks. In addition, we found that AGDP1 is required for transcriptional gene silencing, non-CG DNA methylation, and H3K9 dimethylation. Our ChIP-seq data showed that AGDP1 is enriched in TE regions and associated with the heterochromatin marks. Our findings suggest that, as a heterochromatin binding protein, AGDP1 links H3K9me2 to DNA methylation in heterochromatin regions. MethylC-Seq: 2 samples examined, Col-0 and agdp1-1; mRNA-Seq: 3 samples examined, Col-0 (5 replicates), agdp1-1 (5 replicates), and suvh4/suvh5/suvh6 (suvh456) triple mutant; ChiP-Seq: proAGDP1::3xFlag-gAGDP1 transgenic plants and wild-type control were used for ChIP-seq with two biological replicates; ChIP-Seq: H3K9me2 and H3 in Col-0 and agdp1 mutant (each with two replicates).