EGR1 functions as a new host restriction factor for SARS-CoV-2 to inhibit virus replication through the E3 ubiquitin ligase MARCH8

Coronavirus disease 2019, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has led to an unprecedented public health crisis worldwide. Though the host produces interferons (IFNs) and restriction factors to suppress virus infection, SARS-CoV-2 has evolved multiple strategies to inhibit the antiviral responses. Understanding host restriction factors and viral escape mechanisms is conducive to developing effective anti-SARS-CoV-2 drugs. Here, we constructed SARS-CoV-2 nucleocapsid (2N) protein- and green fluorescent protein (GFP)-stably expressing cells that were transfected with polyinosinic-polycytidylic acid (poly(I:C)) to activate IFN responses. The transcriptome analysis showed that poly(I:C)-induced IFN responses were inhibited by the SARS-CoV-2 N protein. Further analysis revealed that 2N inhibited the production of IFN-stimulated genes by suppressing early growth response gene-1 (EGR1) expression, a transcription factor that can regulate multiple cellular processes. The ectopic expression of EGR1 remarkably reduced 2N expression and suppressed SARS-CoV-2 replication. Mechanistically, EGR1 promoted expression of IFN-regulated antiviral protein (IRAV), which interacted with 2N to induce its degradation via the E3 ubiquitin ligase MARCH8 with the cargo receptor NDP52 in a lysosome-dependent pathway. MARCH8 catalyzed the K48-linked polyubiquitination of 2N at the lysine residue 143, and knockout of endogenous MARCH8 reversed IRAV-mediated 2N degradation. Additionally, the overexpression of IRAV or MARCH8 could inhibit SARS-CoV-2 replication. Our findings reveal that EGR1 is a new host restriction factor to inhibit SARS-CoV-2 replication through the E3 ubiquitin ligase MARCH8, which would contribute to understanding the pathogenesis of emerging coronaviruses.