Intragenic Heterochromatin-dependent RNA Polyadenylation mediated by a Protein Complex in Arabidopsis [RIP-seq]

In eukaryotic cells, the constitutive heterochromatins, characterized by enrichments of DNA cytosine methylation and repressive histone modifications, are formed in both intergenic regions and gene body regions. The ASI1-AIPP1-EDM2 (AAE) complex has been shown to participates in the RNA polyadenylation (poly A) regulation of intronic heterochromatin-containing genes. While, how AAE complex components coordinately interacts with the chromatin and RNA remains largely unknown. Here, based on multi-omics analysis, we proved that ASI1 and EDM2 mostly target the common genomic regions and display high preference to intragenic heterochromatin within the transposable element (TE)-overlapping genes. Besides interplay with chromatin, ASI1 also displays high binding affinity to RNA transcripts from intragenic heterochromatin regions. Evidence from poly (A)-specific sequencing proved that AAE complex controls the choice of poly (A) sites not only in genes with intronic heterochromatin but also in heterochromatic TE-overlapping genes. Importantly, we revealed that AAE complex also participates in the alternative splicing of TE-overlapping gene RPP4. Moreover, evidences from gene expression and poly (A) sequencing revealed that RRM protein FPA functions antagonistically with AAE complex to control polyadenylation of histone demethylase gene IBM1 in locus-dependent manner. Taken together, our results provide fundamental evidences that AAE complex interacts with both the chromatin and RNA in the intragenic heterochromatin to regulate poly (A) site choice, and a bidirectional regulation mechanism of intragenic heterochromatin-dependent RNA processing exists in Arabidopsis. Our finding gained further insights into the molecular interaction pattern of AAE complex with targets and will help to better understand the complicated interplay between epigenetic modification-shaped chromatin and RNA processing. PAT-seq:2 samples examined and each sample has two replicates, Col-0, ASI1;ASI1 ChIP-seq:2 samples examined and each sample has two replicates and 1 input,Col-0,ASI1; EDM2 ChIP-seq:2 samples examined and each sample has two replicates and 1 input,Col-0,EDM2; ASI1 RIP-seq:2 samples examined, Col-0 and ASI1, each sample has two replicates and 1 input.