Widespread Exon Junction Complex Footprints in the RNA Degradome Mark mRNA Degradation before Steady State Translation

Abstract

Exon junction complexes (EJCs) are deposited on mRNAs during splicing and displaced by ribosomes during the pioneer round of translation. Nonsense-mediated mRNA decay (NMD) degrades EJC-bound mRNA, but the lack of suitable methodology has prevented the identification of other degradation pathways. Here, we show that the RNA degradomes of Arabidopsis (Arabidopsis thaliana), rice (Oryza sativa), worm (Caenorhabditis elegans), and human (Homo sapiens) cells exhibit an enrichment of 59 monophosphate (59P) ends of degradation intermediates that map to the canonical EJC region. Inhibition of 59 to 39 exoribonuclease activity and overexpression of an EJC disassembly factor in Arabidopsis reduced the accumulation of these 59P ends, supporting the notion that they are in vivo EJC footprints. Hundreds of Arabidopsis NMD targets possess evident EJC footprints, validating their degradation during the pioneer round of translation. In addition to premature termination codons, plant microRNAs can also direct the degradation of EJC-bound mRNAs. However, the production of EJC footprints from NMD but not microRNA targets requires the NMD factor SUPPRESSOR WITH MORPHOLOGICAL EFFECT ON GENITALIA PROTEIN7. Together, our results demonstrating in vivo EJC footprinting in Arabidopsis unravel the composition of the RNA degradome and provide a new avenue for studying NMD and other mechanisms targeting EJC-bound mRNAs for degradation before steady state translation.

Improved computational analysis of ribosome dynamics from 5’P degradome data using fivepeseq

“…Although initially considered independent events, multiple evidence has now demonstrated that translation and mRNA decay are interconnected processes and that co-translational mRNA degradation is a general phenomenon (2)(3)(4)(5). The interaction between the translation and decay machinery occurs so close that the positions of 5'P co-translational mRNA degradation intermediates can be used as a proxy for ribosome dynamics, as we and others have shown in yeast (6)(7)(8) and plants (4,9,10). In particular, XRN1driven 5'-3' mRNA degradation is linked to the movement of the last translating ribosome allowing to obtain an in vivo footprint of the ribosome position.…”

Telling Footprints: Exon Junction Complexes Mark Targets of Nonsense- and miRNA-Mediated mRNA Decay

“…Similarly, exon junction complexes (EJCs) deposited upstream of the exon-exon junctions on mRNAs during splicing and removed upon the first round of translation, can protect mRNAs from degradation. In the current issue, Lee et al (2020) show that the RNA degradome contains EJC footprints that mark mRNAs degraded during or before the pioneer round of translation. EJC footprints were prevalent in targets of nonsense-mediated decay (NMD) pathway, but were also present in some mRNAs degraded through miRNA-mediated cleavage.…”

“…EJC footprints were prevalent in targets of nonsense-mediated decay (NMD) pathway, but were also present in some mRNAs degraded through miRNA-mediated cleavage. Lee et al (2020) analyzed RNA degradome data for 5'P end occurrence and found that it was relatively higher in the regions 25-30 nt upstream of the 3' ends of exons, where EJCs are canonically positioned. This pattern was present in Arabidopsis, rice, human, and worm datasets, but not in budding yeast that lacks EJCs, suggesting that the 5'P peaks corresponded to 5' termini of RNA degradation intermediates protected from degradation by EJCs.…”

“…The study by Lee et al (2020) indicates that EJC-footprints mark mRNAs that are degraded during or before the pioneer round of translation, many of them through the NMD pathway. Therefore, novel NMD targets could be identified through EJC footprinting.…”

“…Hanna Hõrak Institute of Technology University of Tartu, Estonia hanna.horak@ut.ee ORCID: 0000-0002-6392-859X Sources of exon junction complex (EJC) footprints. EJCs positied on mRNA during splicing will protect mRNA from degradation by 5'-3' exoribonucleases (XRNs) during nonsense-mediated decay (NMD) caused by premature stop codons (PTCs), or after miRNA-mediated cleavage of mRNA (Modified from Lee et al [2020], Figure 12).…”