Investigating the Proviral and Immunogenic Effects of Sars-Cov-2 Accessory Protein 7A

SARS-CoV-2 and related sarbecoviruses encode a set of accessory proteins (3a, 3b, 6, 7a, 7b, 8, 9b, and 10) that control host responses to infection and promote virus growth. Of these accessory proteins, 7a is set apart by its intracellular localization near coronavirus (CoV) budding sites and its incorporation into secreted virions. Initial studies utilizing a virus-like particle (VLP) system identified significant proviral effects mediated by 7a, but the system lacked the context of an actual CoV infection. To investigate 7a functions during CoV infections, we constructed recombinant Mouse Hepatitis Viruses (rMHV strain A59) that express 7a genes. Comparative infections revealed that 7a increased viral replication and viral output in immortalized murine cell cultures and in primary bone marrow-derived macrophages (BMDMs). This proviral effect was independent of a previously reported 7a-mediated interferon antagonizing activity. 7a is a type I transmembrane protein with a short cytoplasmic tail that operates in subcellular trafficking and signal transduction. To further elucidate tail functions, we generated a set of rA59 viruses expressing substitutions in the tail di-lysine motifs. Several substitutions reduced 7a proviral activities; notably the K119A change eliminated 7a support of virus yield. 7a expression was robustly proinflammatory in BMDMs, as measured cytokine arrays. Cytoplasmic tail substitutions tempered these proinflammatory responses, implying connections with proviral activities. SARS-CoV-2 infected macrophages have been implicated in inflammatory COVID-19 and these findings point to 7a cytoplasmic tails as potential contributors to cytokine-mediated disease. This study shows that SARS-CoV-2 accessory protein 7a promotes infection of a phylogenetically distinct embecovirus and in doing so elicits proinflammatory and potentially disease-relevant host responses. The findings localize and highlight a specific proviral component in a sarbecovirus accessory protein.