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Authors:Lan et al 

Journal/ Pre-Print:bioRxiv 

Tags: Bioinformatics, Molecular biology, Virology 

Research Highlights

  1. First in-cell map at single nucleotide resolution of the secondary structures of the SARS-CoV-2 genome 

  1. Uncover new structures in regulatory elements, including the genomic transcription-regulating sequences (TRSs) 

  1. Provide new structures of the frameshift element, which differ from previous in vitro models. There is evidence for heterogeneity of   

Summary

The authors performed the first in-cell map of the secondary structures of the SARS-CoV-2 genome. They used dimethyl sulphate mutational profiling with sequencing (DMS-MaPseq) to find unpaired nucleotides, which were used to constrain in-silico pairing predictions. The 5’UTR structures are similar to previous reports, but new structures were found for the genomic Transcription-Regulating Sequences (TRSs), which mostly lie within stem loops. The structure of the frameshift element (FSE), an important regulatory element of SARS-CoV-2 viral cycle, differed from previous in vitro observations, which the authors argue were an artefact of short length of the refolded viral RNA used for the in vitro studies. Clustering of DMS-MaPseq reads suggested the presence of two distinct FSE structures across different viral copies. 

Impact for SARS-CoV2/COVID19 research efforts   

Understand the virology and/or cell biology of SARS-CoV2/COVID19 

Better understanding of the secondary structures of the SARS-CoV-2 genome, which will be useful for a better design of RNA-based therapeutics. 

Study Type  

  • In silico study / bioinformatics study 

  • In vitro study 

Strengths and limitations of the paper 

Novelty: First in-cell investigation of the secondary structure of the SARS-CoV-2 genome at a single nucleotide resolution. This provides a map of the major secondary structures of the SARS-CoV-2 genome in cells.  

Standing in the field: The major secondary structures at the 5’UTR of the SARS-CoV-2 genome found in this study agree with what has been previously observed in vitro or in other viruses, such as SARS-CoVA comparison of their data with structures computationally predicted by RNAz and Contrafold, found a better agreement with the former. The major difference is observed for the secondary structures of the frameshift element, which differ from previous in vitro studies. However, the authors show that the difference could be explained by the size of the RNA fragment used for determining the secondary structure in vitro, as longer fragment size and a full-length virus show similar secondary structure as the in-cell data.  

Appropriate statistics:Yes 

Viral model used:SARS-CoV-2 

Translatability: No direct one. However, the determination of the secondary structures of the SARS-CoV-2 genome will allow a better design of RNA-based therapeutics. 

Main limitations: 1) Low number of reads, which could result in some secondary structures not observed because they will be below statistical significance. 

2)  The DMS-MaPseq experiment was performed on a population of cells infected with SARS-CoV-2, therefore the secondary structures observed are a mix of viral genomes at different stages of the viral cycle. Further work will be required to determine secondary structures unique to each stage of the virus and their dynamics across the viral cycle.