SARS-CoV-2 Nucleocapsid protein attenuates stress granule formation and interacts directly with mRNAs to impair host stress response (iCLIP-Seq)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid (N) protein is highly expressed upon infection and is essential for viral replication, making it a promising target for antiviral drug and vaccine development. Here, starting from a functional proteomics workflow, we catalogued the protein-protein interactions of 28 SARS-CoV-2 proteins in HEK293 cells, including an evolutionarily conserved specific interaction of N with the stress granule resident proteins G3BP1 and G3BP2. N protein localizes to stress granules and sequesters G3BP1 and G3BP2 away from their normal interaction partners, thus attenuating stress granule formation. N also binds directly to host mRNAs, with a preference for 3´ UTRs, and modulates target mRNA stability. We show that SARS-CoV-2 N protein rewires the G3BP1 mRNA-binding profile, thereby suppressing host gene expression changes induced in response to cellular stress. iCLIP-seq of GFP-tagged SARS-CoV-2 N protein in HEK293 cells treated with or withour Sodium Arsenite. iCLIP-seq of G3BP1 in n HEK293 cells treated with or withour Sodium Arsenite.iCLIP-seq of G3BP1 in N-expressing HEK293 cells treated with or withour Sodium Arsenite. Each experiment was performed in biological duplicate. Control libraries for size-match inputs (SMI) were prepared in parallel