Profiling Selective Packaging of Host RNA and Viral RNA Modification in SARS-CoV-2 Viral Preparations.

Noah Peña,Adolfo Garcia-Sastre,Hala Alshammary,Randy A Albrecht,Mateusz Halucha,Tao Pan,Christopher Katanski,Christopher Watkins,Viviana Simon,Matthew M Hernandez,Wen-Chun Liu,Wen Zhang

doi:10.3389/fcell.2022.768356

Abstract

Viruses package host RNAs in their virions which are associated with a range of functions in the viral life cycle. Previous transcriptomic profiling of host RNA packaging mostly focused on retroviruses. Which host RNAs are packaged in other viruses at the transcriptome level has not been thoroughly examined. Here we perform proof-of-concept studies using both small RNA and large RNA sequencing of six different SARS-CoV-2 viral isolates grown on VeroE6 cells to profile host RNAs present in cell free viral preparations and to explore SARS-CoV-2 genomic RNA modifications. We find selective enrichment of specific host transfer RNAs (tRNAs), tRNA fragments and signal recognition particle (SRP) RNA in SARS-CoV-2 viral preparations. Different viral preparations contain the same set of host RNAs, suggesting a common mechanism of packaging. We estimate that a single SARS-CoV-2 particle likely contains up to one SRP RNA and four tRNA molecules. We identify tRNA modification differences between the tRNAs present in viral preparations and those in the uninfected VeroE6 host cells. Furthermore, we find uncharacterized candidate modifications in the SARS-CoV-2 genomic RNA. Our results reveal an under-studied aspect of viral-host interactions that may be explored for viral therapeutics.

Highlights

Viral assembly is a critical stage in the viral life cycle that produces mature virus containing the viral genome and proteins needed to infect another host target cell
A substantial proportion of reads mapped to viral genomic RNA as expected, and transfer RNAs (tRNAs) and signal recognition particle (SRP) RNA are present at almost high proportions, followed by a small amount of rRNA (Figure 1B)
Our results show a ~150-fold enrichment of the SRP RNA over tRNA in the viral preparation samples vs. the cell samples, which suggests that we eliminated most if not all of the cellular debris. These results indicate that SARS-CoV-2 virions package tRNA and SRP RNA in significant proportions

Summary

Introduction

Viral assembly is a critical stage in the viral life cycle that produces mature virus containing the viral genome and proteins needed to infect another host target cell. As early as 1980s it was shown that certain viruses package host RNAs into their virions. Host transfer RNA (tRNA) is a major cellular RNA family, which is packaged in virions (Isaac and Keene, 1981; Jiang et al, 1993). TRNAs are the most abundant RNA in copy numbers in cells, and their small size and stable structure make them good targets for interacting with viral RNA and viral proteins. The best studied viral packaging of host RNAs has been described for retroviruses. Retroviruses require a specific host tRNA as reverse transcriptase primers in the cDNA synthesis of the viral genomic RNA upon infection. HIV-1 uses tRNALys(TTT) from the host cell since it has a fully

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Conclusion