Exon-junction complex association with stalled ribosomes and slow translation-independent disassembly

• Published in: Nature communications Vol. 15; no. 1; p. 4209
• Main Authors: Bensaude, Olivier, Barbosa, Isabelle, Morillo, Lucia, Dikstein, Rivka, Le Hir, Hervé
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 17.05.2024; Nature Publishing Group; Nature Portfolio
• Subjects: 49/91; 631/337/1645/1769; 631/337/1645/1792; 631/45/500; 96/10; 96/109; Codons; Complex formation; Cytoplasm; Dismantling; Exons - genetics; Gene sequencing; HEK293 Cells; HeLa Cells; Humanities and Social Sciences; Humans; Life Sciences; multidisciplinary; Protein Biosynthesis; Ribonucleic acid; Ribonucleoproteins - genetics; Ribonucleoproteins - metabolism; Ribosomes; Ribosomes - metabolism; RNA; RNA Splicing; RNA, Messenger - genetics; RNA, Messenger - metabolism; Science; Science (multidisciplinary); Translation
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-024-48371-5

Exon junction complexes are deposited at exon-exon junctions during splicing. They are primarily known to activate non-sense mediated degradation of transcripts harbouring premature stop codons before the last intron. According to a popular model, exon-junction complexes accompany mRNAs to the cytoplasm where the first translating ribosome pushes them out. However, they are also removed by uncharacterized, translation-independent mechanisms. Little is known about kinetic and transcript specificity of these processes. Here we tag core subunits of exon-junction complexes with complementary split nanoluciferase fragments to obtain sensitive and quantitative assays for complex formation. Unexpectedly, exon-junction complexes form large stable mRNPs containing stalled ribosomes. Complex assembly and disassembly rates are determined after an arrest in transcription and/or translation. 85% of newly deposited exon-junction complexes are disassembled by a translation-dependent mechanism. However as this process is much faster than the translation-independent one, only 30% of the exon-junction complexes present in cells at steady state require translation for disassembly. Deep RNA sequencing shows a bias of exon-junction complex bound transcripts towards microtubule and centrosome coding ones and demonstrate that the lifetimes of exon-junction complexes are transcript-specific. This study provides a dynamic vision of exon-junction complexes and uncovers their unexpected stable association with ribosomes. Bensaude et al. use a split luciferase approach to show that exon-junction complex assembly and disassembly occur faster when they are translation-dependent than when they are translation-independent; and they uncover an association with ribosomes.