PYM1 limits non-canonical Exon Junction Complex occupancy in a gene architecture dependent manner to tune mRNA expression

The Exon Junction Complex (EJC) decorates RNA exon-exon junctions and modulates mRNA fate at multiple post-transcriptional steps until its disassembly during translation. Our investigation of the EJC disassembly factor PYM1 in human embryonic kidney 293 (HEK293) cells show that the EJC-PYM1 interaction is required for translation-independent EJC destabilization but not for translation-dependent disassembly. Surprisingly, PYM1 interaction deficient EJCs are enriched on locations away from canonical EJC binding site, particularly on transcripts with no or few introns. Such non-canonical EJCs are capable of inducing nonsense-mediated mRNA decay when present downstream of stop codons. Suppression of PYM1 in human cells, including by previously reported PYM1-flavivirus capsid protein interaction, stabilizes mRNAs with fewer and longer exons that localize to endoplasmic reticulum associated TIS-granules. In summary, PYM1 limits non-canonical EJC and thereby acts as an EJC specificity factor that is hijacked by flaviviruses to reshape host cell mRNA regulation. Exon Junction Complexes influence many aspects of mRNA fate. Here, the authors show that the disassembly factor PYM1 primarily acts in translation-independent EJC removal, with resistant EJCs accumulating at non-canonical sites and impacting mRNA stability in a gene architecture dependent manner.