PT  - JOURNAL ARTICLE
AU  - Evangelos D. Karousis
AU  - Lukas-Adrian Gurzeler
AU  - Giuditta Annibaldis
AU  - René Dreos
AU  - Oliver Mühlemann
TI  - Human NMD ensues independently of stable ribosome stalling
AID  - 10.1101/2019.12.11.872861
DP  - 2019 Jan 01
TA  - bioRxiv
PG  - 2019.12.11.872861
4099  - http://biorxiv.org/content/early/2019/12/12/2019.12.11.872861.short
4100  - http://biorxiv.org/content/early/2019/12/12/2019.12.11.872861.full
AB  - Nonsense-mediated mRNA decay (NMD) is a translation-dependent RNA degradation pathway that is important for the elimination of faulty and the regulation of normal mRNAs. The molecular details of the early steps in NMD are not fully understood but previous work suggests that NMD activation occurs as a consequence of ribosome stalling at the termination codon (TC). To test this hypothesis, we established an in vitro translation-coupled toeprinting assay based on lysates from human cells that allows monitoring of ribosome occupancy at the TC of reporter mRNAs. In contrast to the prevailing NMD model, our in vitro system revealed similar ribosomal occupancy at the stop codons of NMD-sensitive and NMD-insensitive reporter mRNAs. Moreover, ribosome profiling revealed a similar density of ribosomes at the TC of endogenous NMD-sensitive and NMD-insensitive mRNAs in vivo. Together, these data show that NMD activation is not accompanied by stable stalling of ribosomes at TCs.