Coronavirus infection induces the unfolded protein response (UPR), a cellular signalling pathway composed of three branches, triggered by unfolded proteins in the endoplasmic reticulum (ER) due to high ER load. We have used RNA sequencing and ribosome profiling to investigate holistically the transcriptional and translational response to cellular infection by murine hepatitis virus (MHV), often used as a model for the Betacoronavirus genus to which the recently emerged SARS-CoV-2 also belongs. We found the UPR to be amongst the most significantly up-regulated pathways in response to MHV infection. To confirm and extend these observations, we show experimentally the induction of all three branches of the UPR in both MHV- and SARS-CoV-2-infected cells. Over-expression of the SARS-CoV-2 ORF8 or S proteins alone is itself sufficient to induce the UPR. Remarkably, pharmacological inhibition of the UPR greatly reduced the replication of both MHV and SARS-CoV-2, revealing the importance of this pathway for successful coronavirus replication. This was particularly striking when both IRE1α and ATF6 branches of the UPR were inhibited, reducing SARS-CoV-2 virion release (~1,000-fold). Together, these data highlight the UPR as a promising antiviral target to combat coronavirus infection.

Coronaviruses (CoVs) are enveloped, positive-sense RNA viruses with an unusually large RNA genome and a unique replication strategy. They cause important diseases in mammals and birds ranging from enteritis in cows and pigs and upper respiratory disease in chickens, to potentially lethal human respiratory infections. Here, we apply ribosome profiling and parallel RNA sequencing to analyse global changes in host cell transcriptome and translatome upon infection with mouse hepatitis virus strain A59 (MHV-A59), a model murine coronavirus in the same genus as the human pathogens severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East respiratory syndrome coronavirus (MERS-CoV). Amongst differentially-regulated cellular genes, we observed up-regulation of all arms of the unfolded protein response (UPR), including translational activation of transcription factors ATF4, ATF5 and Chop. Polysome profiling of infected-cells revealed an accumulation of empty 80S ribosomes, consistent with increased phosphorylation of eIF2α leading to translational shut-off via inhibited initiation. Ribosomal footprints on phosphorylated-eIF2α-resistant mRNAs revealed unambiguous upstream open reading frame (uORF) occupancy consistent with host maintenance of the UPR. Unexpectedly, an inhibitor of PERK that blocks the UPR and relieves translation inhibition was found to attenuate virus growth suggesting that MHV may subvert the UPR to its own advantage. This study sheds new light on the complex interactions between MHV and host during infection and provides new potential targets for antiviral intervention.

The ability of cells to mount an interferon response to virus infections depends on intracellular nucleic acid sensing pattern recognition receptors (PRRs). RIG-I is an intracellular PRR that binds short double stranded viral RNAs to trigger MAVS-dependent signalling. The RIG-I/MAVS signalling complex requires the coordinated activity of multiple kinases and E3 ubiquitin ligases to activate the transcription factors that drive type I and type III interferon production from infected cells. The linear ubiquitin chain assembly complex (LUBAC) regulates the activity of multiple receptor signalling pathways in both ligase-dependent and -independent ways. Here, we show that the three proteins that constitute LUBAC have separate functions in regulating RIG-I signalling. Both HOIP, the E3 ligase capable of generating M1-ubiquitin chains, and LUBAC accessory protein HOIL-1 are required for viral RNA sensing by RIG-I. The third LUBAC component, SHARPIN, is not required for RIG-I signalling. These data cement the role of LUBAC as a positive regulator of RIG-I signalling and as an important component of antiviral innate immune responses.

Campylobacter is the most common cause of bacterial gastroenteritis worldwide, and diarrhoeal disease is a major cause of child morbidity, growth faltering and mortality in low- and middle-income countries. Despite evidence of high incidence and differences in disease epidemiology, there is limited genomic data from studies in developing countries. In this study, we aimed to quantify the extent of gene sharing in local and global populations. We characterized the genetic diversity and accessory-genome content of a collection of Campylobacter isolates from the Cairo metropolitan area, Egypt. In total, 112 Campylobacter isolates were collected from broiler carcasses (n=31), milk and dairy products (n=24), and patients suffering from gastroenteritis (n=57). Among the most common sequence types (STs), we identified the globally disseminated host generalist ST-21 clonal complex (CC21) and the poultry specialists CC206, CC464 and CC48. Notably, CC45 and the cattle-specialist CC42 were under-represented, with a total absence of CC61. Core- and accessory-genome sharing was compared among isolates from Egypt and a comparable collection from the UK (Oxford). Lineage-specific accessory-genome sharing was significantly higher among isolates from the same country, particularly CC21, which demonstrated greater local geographical clustering. In contrast, no geographical clustering was noted in either the core or accessory genome of CC828, suggesting a highly admixed population. A greater proportion of Campylobacter coli isolates were multidrug resistant compared to Campylobacter jejuni . Our results suggest that there is more horizontal transfer of accessory genes between strains in Egypt. This has strong implications for controlling the spread of antimicrobial resistance among this important pathogen.