The translational landscape of SARS-CoV-2 and infected cells

Puray-Chavez, Maritza; Tenneti, Kasyap; Hr, Vuong; N, Lee; Y, Liu; Horani, Amjad; Huang, Tao; Sp, Gunsten; Jb, Case; Yang, Wei; Ms, Diamond; Sl, Brody; Dougherty, Joseph D.; Sb, Kutluay

doi:10.1101/2020.11.03.367516

Cited by 20 publications

(32 citation statements)

References 114 publications

(144 reference statements)

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“…When used in dual luciferase reporters, the longer sequence (3kb) frameshifts at much higher rate than the minimal FSE (~40% compared to ~20% of the minimal sequence). These results underscore a functional role for long range RNA interactions (Ziv et al , 2020) and explain data from recent ribosome profiling studies showing that the ribosomes frameshifts at ~50% in infected cells (Puray-Chavez et al , 2020; Finkel et al , 2021).…”

Section: Discussionsupporting

confidence: 80%

Insights into the secondary structural ensembles of the full SARS-CoV-2 RNA genome in infected cells

Lan

Allan

Malsick

et al. 2020

Preprint

196

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SUMMARYSARS-CoV-2 is a betacoronavirus with a single-stranded, positive-sense, 30-kilobase RNA genome responsible for the ongoing COVID-19 pandemic. Currently, there are no antiviral drugs or vaccines with proven efficacy, and development of these treatments are hampered by our limited understanding of the molecular and structural biology of the virus. Like many other RNA viruses, RNA structures in coronaviruses regulate gene expression and are crucial for viral replication. Although genome and transcriptome data were recently reported, there is to date little experimental data on predicted RNA structures in SARS-CoV-2 and most putative regulatory sequences are uncharacterized. Here we report the secondary structure of the entire SARS-CoV-2 genome in infected cells at single nucleotide resolution using dimethyl sulfate mutational profiling with sequencing (DMS-MaPseq). Our results reveal previously undescribed structures within critical regulatory elements such as the genomic transcription-regulating sequences (TRSs). Contrary to previous studies, our in-cell data show that the structure of the frameshift element, which is a major drug target, is drastically different from prevailing in vitro models. The genomic structure detailed here lays the groundwork for coronavirus RNA biology and will guide the design of SARS-CoV-2 RNA-based therapeutics.

show abstract

Section: Discussionsupporting

confidence: 80%

Insights into the secondary structural ensembles of the full SARS-CoV-2 RNA genome in infected cells

Lan

Allan

Malsick

et al. 2020

Preprint

196

View full text Add to dashboard Cite

show abstract

“…When used in dual luciferase reporters, the longer sequence (3kb) frameshifts at much higher rate than the minimal FSE (~ 40% compared to ~ 20% of the minimal sequence). These results underscore a functional role for long range RNA interactions (Ziv et al, 2020) and explain data from recent ribosome pro ling studies showing that the ribosomes frameshifts at ~ 50% in infected cells (Puray-Chavez et al, 2020;Finkel et al, 2021).…”

Section: Discussionsupporting

confidence: 79%

Insights into the secondary structural ensembles of the full SARS-CoV-2 RNA genome in infected cells

Rouskin

Lan

Allan

et al. 2021

Preprint

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SARS-CoV-2 is a betacoronavirus with a single-stranded, positive-sense, 30-kilobase RNA genome responsible for the ongoing COVID-19 pandemic. Currently, there are no antiviral drugs with proven efficacy, and development of these treatments are hampered by our limited understanding of the molecular and structural biology of the virus. Like many other RNA viruses, RNA structures in coronaviruses regulate gene expression and are crucial for viral replication. Although genome and transcriptome data were recently reported, there is to date little experimental data on native RNA structures in SARS-CoV-2 and most putative regulatory sequences are functionally uncharacterized. Here we report secondary structure ensembles of the entire SARS-CoV-2 genome in infected cells at single nucleotide resolution using dimethyl sulfate mutational profiling with sequencing (DMS-MaPseq) and the algorithm ‘detection of RNA folding ensembles using expectation–maximization’ clustering (DREEM). Our results reveal previously undescribed alternative RNA conformations across the genome, including structures of the frameshift stimulating element (FSE), a major drug target, that are drastically different from prevailing in vitro population average models. Importantly, we find that this structural ensemble promotes frameshifting rates (~40%) similar to in vivo ribosome profiling studies and much higher than the canonical minimal FSE (~20%). Overall, our result highlight the value of studying RNA folding in its native, dynamic and cellular context. The genomic structures detailed here lays the groundwork for coronavirus RNA biology and will guide the design of SARS-CoV-2 RNA-based therapeutics.

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“…These results were consistent in Huh7 cells also, validating this mechanism across cell types (Figure S4A). Recent reports have demonstrated a global decay of host mRNA possibly driven by Nsp1 during SARS-CoV-2 infection (6, 18). We investigated the association of the loss of 4EBP1 and ULK1 upon infection with a potential degradation of their transcripts using quantitative RT-PCR and surprisingly detected significantly elevated levels of their transcripts in the infected cells indicating the involvement of post-transcriptional regulations (Figure 5 B and C).…”

Section: Resultsmentioning

confidence: 99%

“…Nsp1 is reported to interfere with host translation through its interaction with 40S ribosomes (6, 14-17). Reports also indicate that translation efficiency of viral mRNAs are not higher than the host mRNAs, but SARS-CoV-2 mediated preferential destruction of host mRNAs lead to their reduced translation events (6, 18). However, the molecular mechanisms remain much elusive.…”

Section: Introductionmentioning

confidence: 99%

Suppression of Global Protein Translation in SARS-CoV-2 Infection

Parthasarathy

Gupta

et al. 2021

Preprint

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The relationship of SARS-CoV-2 with the host translation remains largely unexplored. Using polysome profiling of SARS-CoV-2 infected Caco2 cells, we here demonstrate that the virus induces a strong suppression of global translation by 48 hours of infection. Heavy polysome fractions displayed substantial depletion in the infected cells, indicating the loss of major translational activities in them. Further assessment of the major pathways regulating translation in multiple permissive cell lines revealed strong eIF4E dephosphorylation accompanied by Mnk1 depletion and ERK1/2 dephosphorylations. p38MAPK showed consistent activation and its inhibition lowered viral titers, indicating its importance in viral survival. No significant change was noticed in eIF2 α phosphorylation. mTORC1 pathway showed the most profound inhibition, indicating its potential contribution to the suppression of global translation associated with the infection. Pharmacological activation of mTORC1 caused a drop in viral titers while inhibition resulted in higher viral RNA levels, confirming a critical role of mTORC1 in regulating viral replication. Surprisingly, the infection did not cause a general suppression of 5′-TOP translation, as evident from the continued expression of ribosomal proteins. Our results collectively indicate that the differential suppression of mTORC1 might allow SARS-CoV-2 to hijack translational machinery in its favor and specifically target a set of host mRNAs.

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The translational landscape of SARS-CoV-2 and infected cells

Cited by 20 publications

References 114 publications

Insights into the secondary structural ensembles of the full SARS-CoV-2 RNA genome in infected cells

Insights into the secondary structural ensembles of the full SARS-CoV-2 RNA genome in infected cells

Insights into the secondary structural ensembles of the full SARS-CoV-2 RNA genome in infected cells

Suppression of Global Protein Translation in SARS-CoV-2 Infection

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