The RNA-binding dynamics of the exon-junction complex underlying splicing-coupled RNA fate determination in Arabidopsis

The exon-junction complex represents a hallmark of protein-RNA packaging that is coupled with co-transcriptional RNA processing and influences the subsequent fate of RNA. Although being extensively studied in animals, the RNA-binding property and the function of EJC in plants remain elusive. Through eCLIP-seq, we found that the RNA-binding core of the EJC, eIF4AIII, is delivered to the -25 nt to -30 nt region upstream of exon-exon junctions in a splicing-dependent manner. eIF4AIII also binds the 3′ end of introns, suggesting an intermediate binding state prior to exon-exon junction deposition. eIF4AIII binding strength correlates with the number of gene exons and is distinct across different genes. Inducible knockdown of eIF4AIII leads to widespread alteration of intron-retention, a small fraction of which is also regulated by HEN2 or UPF3, indicating roles for eIF4AIII in RNA decay and RNA splicing. Interestingly, CRISPR-mediated fragment deletion that introduces an exonic premature termination codon (PTC) leads to NMD independently of eIF4AIII, even when the EJC is deposited downstream of the PTC, indicating that the EJC is not compulsory for NMD in Arabidopsis. Overall, our study uncovers the RNA-binding landscape and novel functions of the exon-junction complex in plants.