Abstract
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.
Highlights
The Coronaviridae are a family of enveloped viruses with positive-sense, non-segmented, single-stranded RNA genomes
During CoV replication, the massive production and modification of viral proteins, as well as virion budding-related endoplasmic reticulum (ER) membrane depletion, can lead to overloading of the folding capacity of the ER and ER stress [6]. This activates the unfolded protein response (UPR) which is controlled by three ER-resident transmembrane sensors: inositol-requiring enzyme-1 α (IRE1α), activating transcription factor-6 (ATF6), and PKR-like ER kinase (PERK), each triggering a different branch of the UPR (Fig 1A)
The most significantly enriched pathway associated with transcriptionally up-regulated genes was “Unfolded Protein Response” (R-HSA-381119, p = 1.1×10−10), and pathways denoting the three branches of the UPR (ATF6 branch: R-HSA-381183, PERK branch: R-HSA-380994, IRE1α branch: R-HSA-381070) were significantly enriched (S3 Table)
Summary
The Coronaviridae are a family of enveloped viruses with positive-sense, non-segmented, single-stranded RNA genomes. SARS-CoV-2 is the causative agent of the current COVID-19 pandemic, which has resulted in over 150 million cases and more than 3 million deaths since the end of 2019. Up to 15% of the cases develop a severe pathology [4,5], no specific therapeutic treatment for COVID-19 has been approved to date, highlighting the urgent need to identify new antiviral strategies to combat SARS-CoV-2, besides future CoV zoonoses. During CoV replication, the massive production and modification of viral proteins, as well as virion budding-related endoplasmic reticulum (ER) membrane depletion, can lead to overloading of the folding capacity of the ER and ER stress [6] This activates the unfolded protein response (UPR) which is controlled by three ER-resident transmembrane sensors: inositol-requiring enzyme-1 α (IRE1α), activating transcription factor-6 (ATF6), and PKR-like ER kinase (PERK), each triggering a different branch of the UPR (Fig 1A). Activation of these pathways leads to decreased protein synthesis and increased ER folding capacity, returning the cell to homeostasis [7]
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