Systematic discovery and functional interrogation of SARS-CoV-2 viral RNA-host protein interactions during infection

Abstract

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of a pandemic with growing global mortality. There is an urgent need to understand the molecular pathways required for host infection and anti-viral immunity. Using comprehensive identification of RNA-binding proteins by mass spectrometry (ChIRP-MS), we identified 309 host proteins that bind the SARS-CoV-2 RNA during active infection. Integration of this data with viral ChIRP-MS data from three other positive-sense RNA viruses defined pan-viral and SARS-CoV-2-specific host interactions. Functional interrogation of these factors with a genome-wide CRISPR screen revealed that the vast majority of viral RNA-binding proteins protect the host from virus-induced cell death, and we identified known and novel anti-viral proteins that regulate SARS-CoV-2 pathogenicity. Finally, our RNA-centric approach demonstrated a physical connection between SARS-CoV-2 RNA and host mitochondria, which we validated with functional and electron microscopy data, providing new insights into a more general virus-specific protein logic for mitochondrial interactions. Altogether, these data provide a comprehensive catalogue of SARS-CoV-2 RNA-host protein interactions, which may inform future studies to understand the mechanisms of viral pathogenesis, as well as nominate host pathways that could be targeted for therapeutic benefit.

Highlights

* · ChIRP-MS of SARS-CoV-2 RNA identifies a comprehensive viral RNA-host protein interaction network during infection across two species

* · Comparison to RNA-protein interaction networks with Zika virus, dengue virus, and rhinovirus identify SARS-CoV-2-specific and pan-viral RNA protein complexes and highlights distinct intracellular trafficking pathways

* · Intersection of ChIRP-MS and genome-wide CRISPR screens identify novel SARS-CoV-2-binding proteins with pro- and anti-viral function

* · Viral RNA-RNA and RNA-protein interactions reveal specific SARS-CoV-2-mediated mitochondrial dysfunction during infection

Competing Interest Statement

K.R.P., H.Y.C., and A.T.S. are co-founders of Cartography Biosciences. A.T.S. is a co-founder of Immunai and receives research funding from Arsenal Biosciences, Sonoma Biotherapeutics, and Allogene Therapeutics. H.Y.C. is a co-founder of Accent Therapeutics, Boundless Bio, and an advisor for 10x Genomics, Arsenal Biosciences, and Spring Discovery. Yale University (C.B.W.) has a patent pending related to this work entitled: Compounds and Compositions for Treating, Ameliorating, and/or Preventing SARS-CoV-2 Infection and/or Complications Thereof. Yale University has committed to rapidly executable non-exclusive royalty-free licenses to intellectual property rights for the purpose of making and distributing products to prevent, diagnose and treat COVID-19 infection during the pandemic and for a short period thereafter.

Footnotes

* Lead Contact: raflynn{at}stanford.edu