SARS-CoV2 Immunity and Cell Biology

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Principal Investigator: Jonathan W Yewdell
Organization: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
Fiscal Year: 2020
Award: $490,071
Funding agency: National Institute of Allergy and Infectious Diseases

We have generated Sleeping Beauty transposase vectors that drive expression of the SARS-CoV-2 receptor ACE2 and or protease TMPRSS2 in host cells and use these vectors to create permanent transfected lines of A549, 293FT, and BHK-21 cells.
We have generated a recombinant vesicular stomatitis virus that expresses a reporter fluorescent protein and SARS-CoV-2 spike protein in place of the normal viral receptor.  This virus allows us to perform highly accurate neutralization assays using anti-spike mAbs or polyclonal sera.  
We have generated several hundred mouse mAbs specific for SARS-CoV-2 spike protein and are in the process of defining their epitopes and biological activity.  For this effort, we are generating library in which each residue in spike is replaced with alanine.  The library will be used express spike via transfection or by an equivalent recombinant vesicular stomatitis library.
We are generating mAbs specific for SARS-CoV-2 nucleocapsid protein.
We have characterized the binding of dozens of chemokines to a number of SARS-CoV-2 proteins and have discovered a number of chemokines that bind with high affinity as determined by bio-layer interferometry.
We have created a system for generating recombinant seasonal CoVs or recombinant SARS-CoV-2.  We are creating seasonal CoVs that express SARS-CoV-2 spike in place of native spike to enable a more realistic neutralization assay.  We are also generating recombinant CoVs that express MHC class I peptides to enable studying the effects of CoV infection on the MHC class I antigen processing pathway.

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