The exon junction complex coordinates the cotranscriptional inclusion of blocks of neighboring exons

The exon junction complex (EJC) has roles in mRNA export and cytoplasmic quality control. However, the EJC is recruited to pre-mRNA by the spliceosome prior to the completion of splicing. When splicing is cotranscriptional, the EJC is deposited on nascent RNA early during synthesis, raising the question of whether the EJC regulates downstream RNA processing. Here we show, using long-read sequencing, that degron-mediated depletion of EJC component EIF4A3 leads to skipping of neighboring pairs of two or more exons on the same mRNA molecule. These data suggest that the entire “exon block” requires the EJC for inclusion. Introns flanking EJC-dependent exon blocks were longer and spliced after internal introns. In our working model, block exons are first spliced together to form a larger EJC-marked exon that promotes surrounding splicing events. Strikingly, analysis of 480 RNA binding protein knockdowns across two different human cell lines revealed block exons that are dependent on other splicing factors, indicating that coordinated splicing of adjacent exons is a general mechanism, of which the EJC is the dominant regulator. Cell type-specific coordinated splicing of adjacent exon pairs has been observed before. Here we identify the EJC as the main protein factor massively regulating this novel splicing mechanism in trans.