Readthrough of stop codons under limiting ABCE1 concentration involves frameshifting and inhibits nonsense-mediated mRNA decay

To gain insight into the mechanistic link between translation termination and nonsense-mediated mRNA decay (NMD), we depleted the ribosome recycling factor ABCE1 in human cells, resulting in an upregulation of NMD-sensitive mRNAs. Suppression of NMD on these mRNAs occurs prior to their SMG6-mediated endonucleolytic cleavage. ABCE1 depletion caused ribosome stalling at termination codons (TCs) and increased ribosome occupancy in 3' UTRs, implying enhanced TC readthrough. ABCE1 knockdown indeed increased the rate of readthrough and continuation of translation in different reading frames, providing a possible explanation for the observed NMD inhibition, since enhanced readthrough displaces NMD activating proteins from the 3' UTR. Our results indicate that stalling at TCs triggers ribosome collisions and activates ribosome quality control. Collectively, we show that improper translation termination can lead to readthrough of the TC, presumably due to ribosome collisions pushing the stalled ribosomes into the 3' UTR, where it can resume translation in-frame as well as out-of-frame

Resurgence of Ebola virus in 2021 in Guinea suggests a new paradigm for outbreaks

These authors contributed equally: Alpha K. Keita, Fara R. Koundouno, Martin Faye, Ariane Düx, Julia Hinzmann.International audienc

Ribosome profiling in mammalian cells to reveal the role of NMD factors in translation termination

Eukaryotes have acquired numerous ways to control the integrity and quality of mRNA. Nonsense-mediated mRNA decay (NMD) is one of these conserved pathways that contribute to mRNA surveillance. It is thought NMD is activated when a ribosome stalls at a termination codon (TC) in a context in which the presence/absence of specific messenger ribonucleoproteins (mRNPs) prevents correct and efficient termination [1-2]. In the past, NMD was believed to degrade exclusively aberrant mRNAs with premature TCs and hence avoid production of potentially deleterious truncated proteins. However, with the advent of transcriptome-wide profiling methods, it became clear that NMD regulates the abundance of 5-10% of all mRNAs, the vast majority of them encoding perfectly functional full-length proteins [3-6]. Unfortunately, the different approaches used in the above-mentioned studies yielded a highly divergent set of apparent endogenous NMD-targeted mRNAs, which so far precluded the identification of a bona fide NMD target set of mRNAs. Recently, Ingolia and Weissman have developed a technique termed ribosome profiling, which enables to determine a transcriptome-wide snapshot of ribosome occupancy in vivo by the analysis of ribosome-protected mRNA fragments (RPFs) obtained from mild nuclease digestion and identified by deep sequencing [7-8]. Using ribosome profiling, we want to determine the ribosome occupancy at TCs relative to the occupancy within the coding sequence of the same mRNA as a surrogate for translation termination efficiency. With this approach, we aim at testing in mammalian cells whether prolonged stalling of ribosomes at TCs can indeed reliably identify transcriptome-wide the corresponding mRNAs as NMD substrates. Moreover, comparing ribosome profiles from cells depleted of NMD factors or other factors with putative roles in translation termination may give insights into the exact role of these factors in the efficiency and fidelity of translation termination

Human NMD ensues independently of stable ribosome stalling

Nonsense-mediated mRNA decay (NMD) was thought to ensue when ribosomes fail to terminate translation properly. However, the authors observe similar ribosome occupancy at stop codons of NMD sensitive and insensitive mRNAs, showing that human NMD is not activated by stable ribosome stalling as previously suggested

A prospective, multi-site, cohort study to estimate incidence of infection and disease due to Lassa fever virus in West African countries (the Enable Lassa research programme)-Study protocol.

BackgroundLassa fever (LF), a haemorrhagic illness caused by the Lassa fever virus (LASV), is endemic in West Africa and causes 5000 fatalities every year. The true prevalence and incidence rates of LF are unknown as infections are often asymptomatic, clinical presentations are varied, and surveillance systems are not robust. The aim of the Enable Lassa research programme is to estimate the incidences of LASV infection and LF disease in five West African countries. The core protocol described here harmonises key study components, such as eligibility criteria, case definitions, outcome measures, and laboratory tests, which will maximise the comparability of data for between-country analyses.MethodWe are conducting a prospective cohort study in Benin, Guinea, Liberia, Nigeria (three sites), and Sierra Leone from 2020 to 2023, with 24 months of follow-up. Each site will assess the incidence of LASV infection, LF disease, or both. When both incidences are assessed the LASV cohort (nmin = 1000 per site) will be drawn from the LF cohort (nmin = 5000 per site). During recruitment participants will complete questionnaires on household composition, socioeconomic status, demographic characteristics, and LF history, and blood samples will be collected to determine IgG LASV serostatus. LF disease cohort participants will be contacted biweekly to identify acute febrile cases, from whom blood samples will be drawn to test for active LASV infection using RT-PCR. Symptom and treatment data will be abstracted from medical records of LF cases. LF survivors will be followed up after four months to assess sequelae, specifically sensorineural hearing loss. LASV infection cohort participants will be asked for a blood sample every six months to assess LASV serostatus (IgG and IgM).DiscussionData on LASV infection and LF disease incidence in West Africa from this research programme will determine the feasibility of future Phase IIb or III clinical trials for LF vaccine candidates