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First Author: Brook E. Heaton  

Journal/preprint name: bioRxiv 

Tags: CRISPR/Cas9 screen, kinases, viral nucleocapsid protein, phosphorylation 

Summary 

The authors performed a CRISPR/Cas9 screen in A549 cells overexpressing ACE2 to find new host factors involved in SARS-CoV-2 infection. Even though the screening was sub-optimal, the authors found that the host kinases SRPK1 and SRPK2 are important for SARS-CoV-2 virus production. They show that these two kinases phosphorylate in vitro the viral nucleocapsid protein. Inhibition of SRPK1 and SRPK2 by small molecules inhibitors decrease SARS-CoV-2 and coronavirus 229E viral production in vitro (immortalized cell lines and primary human lung cells).  

Research Highlights

  1.  Host kinases SRPK1 and SRPK2 are important for SARS-CoV-2 virus production 

  1.  SRPK1 and SRPK2 phosphorylates the virus nucleocapsid protein. 

  1. Inhibition of SRPK1 and SRPK2 by small molecule inhibitors decreased viral production of SARS-CoV-2 and coronavirus 229E in vitro. 

Impact for COVID-19 research:  

  • Does this research alter our view of the disease? 

Evidence that the host kinases SRPK1 and SRPK2 are important for SARS-CoV-2 virus production. This is potentially mediated by the phosphorylation by these two kinases of the viral nucleocapsid protein.  

  • Could this research benefit clinicians (disease management etc.)? 

Inhibition of SRPK1 and SRPK2 decrease SARS-CoV-2 virus production in immortalized cells and in primary human lung cells. Small molecules inhibitors of SRPK1 and SRPK2 are available but none of them have been currently approved by the FDA for therapeutic purposes. 

 

Methodologies: 

  • Study Typein vitro 

  • Important cell lines/viral models used: Cell lines: A549 cell line overexpressing ACE2, Calu-3 cell line, hUH7 cell line, and primary type II pneumocytes. Viral models (coronavirus): SARS-CoV-2 and 229E. 

  • Key Techniques: CRISPR/Cas9 screen.  

Limitations: 

  • No direct confirmation that inhibition of SRPK1 and SRPK2 decreases the phosphorylation of the viral nucleocapsid (N) protein (potential kinases redundancy for example). 

  • No direct evidence that dephosphorylation of the viral N protein is responsible for the decreased SARS-CoV-2 virus production.  

  • No in vivo evidence that inhibition of SRPK1 and SRPK2 decrease SARS-CoV-2 virus production (only performed in vitro). 

  • CRISPR/Cas9 screening in A549 cells overexpressing ACE2 is sub-optimal due to limited cytotoxicity of SARS-CoV-2 in this cell line.  

  • Reliant on pharmacologic kinase inhibition that is fairly toxic