Achievements

Identification and characterization of ubiquitin-modified SARS-CoV-2 viral proteins RuthMabel Boytz, Jadon Layne, Chu-Chieh Hsu, Maudry Laurent-Rolle Protein ubiquitination is a common post-translational modification (PTM) with many diverse effects on cellular proteins, including proteasomal degradation, protein localization, and biochemical activity. The ubiquitination system is also extensively involved in innate immune signaling pathways in response to viral infection. Many viruses have evolved mechanisms to modulate the host ubiquitin system to promote their replication, such as by inhibiting interferon (IFN) production and signaling and increasing the stability or activity of viral proteins involved in replication. SARS-CoV-2 is known to induce strong cytokine responses in patients, indicative of innate immune activation, yet little is known about how SARS-CoV-2 interacts with the host ubiquitination pathways. We screened 21 structural, non-structural, and accessory proteins from SARS-CoV-2 for ubiquitin modification and found that NSP2, NSP5, NSP12, NSP15, and NSP16 get ubiquitinated. We then tested these proteins for the ability to inhibit IFN responses and found that NSP5 inhibits IFN- signaling. NSP12, the viral RNA-dependent RNA polymerase, associated with poly-ubiquitin chains, but this did not appear to target NSP12 for proteasomal degradation. Mass-spectrometry analysis of NSP12 identified several ubiquitinated lysine residues, but mutation of these lysines to arginine failed to reduce the ubiquitination levels of NSP12, suggesting that non-canonical residues may be ubiquitinated. In support of this hypothesis, NSP12 associated with the non-canonical E3 ubiquitin ligase MYCBP2, which preferentially modifies serine and threonine. Overall, our results identified several viral proteins that are ubiquitinated by host cells and demonstrate the need for additional studies to characterize the effects of these modifications on innate immune responses and viral replication.