The Translational Landscape of SARS-CoV-2-infected Cells Reveals Suppression of Innate Immune Genes

Puray-Chavez, Maritza; Lee, Nakyung; Tenneti, Kasyap; Wang, Yiqing; Vuong, Hung R.; Liu, Yating; Horani, Amjad; Huang, Tao; Gunsten, Sean P.; Case, James Brett; Yang, Wei; Diamond, Michael S.; Brody, Steven L.; Dougherty, Joseph D.; Kutluay, Sebla B.

doi:10.1128/mbio.00815-22

Cited by 31 publications

(25 citation statements)

References 93 publications

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“…Surprisingly, we and others observed a large increase in eIF2A protein abundance in SARS-CoV-2-infected cells 33,34 , while its mRNA level remained unchanged, suggesting either that eIF2A mRNA was more efficiently translated or that eIF2A protein became more stable upon infection. Interestingly, a recent study has shown a time-dependent increase in eIF2A mRNA translational efficiency after SARS-CoV-2 infection 35 ( Figure S4C ). The same study has also shown that –1 PRF efficiency, as measured by the relative ribosome footprint densities between ORF1A and ORF1B, is significantly increased at later time points during infection.…”

Section: Discussionmentioning

confidence: 79%

“…Another proteomics study using Caco-2 cells has also shown that eIF2A protein abundance is progressively increased after SARS-CoV-2 infection 34 ( Figure S4B ). A third study by profiling ribosome occupancy has shown an increase in eIF2A mRNA translational efficiency after SARS-CoV-2 infection ( Figure S4C ) 35 . Consistent with these studies, we found that eIF2A protein abundance was increased in SARS-CoV-2-infected Vero E6 cells ( Figure 4A, B ), while its mRNA level remained unchanged ( Figure 4C ).…”

Section: Resultsmentioning

confidence: 97%

“…( A-B ) Boxplots reproduced using datasets from Stukalov et al (2021) 33 ( A ) and Bojkova et al (2020) 34 ( B ). ( C ) Changes in the translational efficiency (TE) of eIF2A mRNA using data from Puray-Chavez et al (2022) 35 .…”

Section: Supplementary Materialsmentioning

confidence: 99%

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Genome-wide CRISPR screens identify noncanonical translation factor eIF2A as an enhancer of SARS-CoV-2 programmed –1 ribosomal frameshifting

Wei

Sun

Guo

2023

Preprint

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Many positive-strand RNA viruses, including all known coronaviruses, employ programmed -1 ribosomal frameshifting (-1 PRF) to regulate the translation of polycistronic viral RNAs. However, only a few host factors have been shown to regulate -1 PRF. Through a reporter-based genome-wide CRISPR/Cas9 knockout screen, we identified several host factors that either suppressed or enhanced -1 PRF of SARS-CoV-2. One of these factors is eukaryotic translation initiation factor 2A (eIF2A), which specifically and directly enhanced -1 PRF in vitro and in cells. Consistent with the crucial role of efficient -1 PRF in transcriptase/replicase expression, loss of eIF2A reduced SARS-CoV-2 replication in cells. Transcriptome-wide analysis of eIF2A-interacting RNAs showed that eIF2A primarily interacted with 18S ribosomal RNA near the contacts between the SARS-CoV-2 frameshift-stimulatory element (FSE) and the ribosome. Thus, our results revealed an unexpected role for eIF2A in modulating the translation of specific RNAs independent of its previously described role during initiation.

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Section: Discussionmentioning

confidence: 79%

Section: Resultsmentioning

confidence: 97%

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Genome-wide CRISPR screens identify noncanonical translation factor eIF2A as an enhancer of SARS-CoV-2 programmed –1 ribosomal frameshifting

Wei

Sun

Guo

2023

Preprint

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“…It can be seen from Figure 14 that in more cells, the change of CAI is larger than zero (meaning that the CAI of the experimental group is higher than that of the control group), which indicates that maybe after virus infection, the gene expression of cells changes to be more conducive to virus translational elongation. Additionally, in the dataset of HBEC cells ( Puray-Chavez et al., 2022 ), the CAI values basically increase with the infection time and the CAI values in infected cells are all larger than control cells after 48 hours of infection. This is also strong evidence for the above conclusion.…”

Section: Resultsmentioning

confidence: 97%

“…However, the correlation between CAI and translational efficiency is maintained in cells infected with SARS-CoV-2. Thus, for studying the effect of viral infection on the translational elongation of SARS-CoV-2 mRNAs, we selected the gene expression datasets of the cells in the control state and the virus-infected state ( Daamen et al., 2021 ; Mulay et al., 2021 ; Sun et al., 2021 ; Puray-Chavez et al., 2022 ), and calculated the CAI values of the SARS-CoV-2 genomes in the corresponding cell datasets. The relative changes of CAI values in the control group and experimental group (virus infection) are consistent in different SARS-CoV-2 subtypes, and the relative positions of CAI values corresponding to ORFs are unchanged.…”

Section: Resultsmentioning

confidence: 99%

Utilizing the codon adaptation index to evaluate the susceptibility to HIV-1 and SARS-CoV-2 related coronaviruses in possible target cells in humans

Zhou

Ren

Yau

2023

Front. Cell. Infect. Microbiol.

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Comprehensive identification of possible target cells for viruses is crucial for understanding the pathological mechanism of virosis. The susceptibility of cells to viruses depends on many factors. Besides the existence of receptors at the cell surface, effective expression of viral genes is also pivotal for viral infection. The regulation of viral gene expression is a multilevel process including transcription, translational initiation and translational elongation. At the translational elongation level, the translational efficiency of viral mRNAs mainly depends on the match between their codon composition and cellular translational machinery (usually referred to as codon adaptation). Thus, codon adaptation for viral ORFs in different cell types may be related to their susceptibility to viruses. In this study, we selected the codon adaptation index (CAI) which is a common codon adaptation-based indicator for assessing the translational efficiency at the translational elongation level to evaluate the susceptibility to two-pandemic viruses (HIV-1 and SARS-CoV-2) of different human cell types. Compared with previous studies that evaluated the infectivity of viruses based on codon adaptation, the main advantage of our study is that our analysis is refined to the cell-type level. At first, we verified the positive correlation between CAI and translational efficiency and strengthened the rationality of our research method. Then we calculated CAI for ORFs of two viruses in various human cell types. We found that compared to high-expression endogenous genes, the CAIs of viral ORFs are relatively low. This phenomenon implied that two kinds of viruses have not been well adapted to translational regulatory machinery in human cells. Also, we indicated that presumptive susceptibility to viruses according to CAI is usually consistent with the results of experimental research. However, there are still some exceptions. Finally, we found that two viruses have different effects on cellular translational mechanisms. HIV-1 decouples CAI and translational efficiency of endogenous genes in host cells and SARS-CoV-2 exhibits increased CAI for its ORFs in infected cells. Our results implied that at least in cases of HIV-1 and SARS-CoV-2, CAI can be regarded as an auxiliary index to assess cells’ susceptibility to viruses but cannot be used as the only evidence to identify viral target cells.

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Optimization of 5′UTR to evade SARS-CoV-2 Nonstructural protein 1-directed inhibition of protein synthesis in cells

Chen

Chang

et al. 2023

Appl Microbiol Biotechnol

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Maximizing the expression level of therapeutic proteins in cells is the general goal for DNA/mRNA therapies. It is particularly challenging to achieve efficient protein expression in the cellular contexts with inhibited translation machineries, such as in the presence of cellular Nonstructural protein 1 (Nsp1) of coronaviruses (CoVs) that has been reported to inhibit overall protein synthesis of host genes and exogenously delivered mRNAs/DNAs. In this study, we thoroughly examined the sequence and structure contexts of viral and non-viral 5′UTRs that determine the protein expression levels of exogenously delivered DNAs and mRNAs in cells expressing SARS-CoV-2 Nsp1. It was found that high 5′-proximal A/U content promotes an escape from Nsp1-directed inhibition of protein synthesis and results in selective protein expression. Furthermore, 5′-proximal Cs were found to significantly enhance the protein expression in an Nsp1-dependent manner, while Gs located at a specific window close to the 5′-end counteract such enhancement. The distinct protein expression levels resulted from different 5′UTRs were found correlated to Nsp1-induced mRNA degradations. These findings ultimately enabled rational designs for optimized 5′UTRs that lead to strong expression of exogenous proteins regardless of the translationally repressive Nsp1. On the other hand, we have also identified several 5′-proximal sequences derived from host genes that are capable of mediating the escapes. These results provided novel perspectives to the optimizations of 5′UTRs for DNA/mRNA therapies and/or vaccinations, as well as shedding light on the potential host escapees from Nsp1-directed translational shutoffs. Key points • The 5′-proximal SL1 and 5a/b derived from SARS-CoV-2 genomic RNA promote exogenous protein synthesis in cells expressing Nsp1 comparing with non-specific 5′UTRs. • Specific 5′-proximal sequence contexts are the key determinants of the escapes from Nsp1-directed translational repression and thereby enhance protein expressions. • Systematic mutagenesis identified optimized 5′UTRs that strongly enhance protein expression and promote resistance to Nsp1-induced translational repression and RNA degradation. Supplementary Information The online version contains supplementary material available at 10.1007/s00253-023-12442-2.

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The Translational Landscape of SARS-CoV-2-infected Cells Reveals Suppression of Innate Immune Genes

Abstract: SARS-CoV-2 utilizes a number of strategies to modulate host responses to ensure efficient propagation. Here, we used ribosome profiling in SARS-CoV-2-infected cells to gain a deeper understanding of the translationally regulated events in infected cells.

Cited by 31 publications

References 93 publications

Genome-wide CRISPR screens identify noncanonical translation factor eIF2A as an enhancer of SARS-CoV-2 programmed –1 ribosomal frameshifting

Genome-wide CRISPR screens identify noncanonical translation factor eIF2A as an enhancer of SARS-CoV-2 programmed –1 ribosomal frameshifting

Utilizing the codon adaptation index to evaluate the susceptibility to HIV-1 and SARS-CoV-2 related coronaviruses in possible target cells in humans

Optimization of 5′UTR to evade SARS-CoV-2 Nonstructural protein 1-directed inhibition of protein synthesis in cells

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