Poster abstracts
Poster number 31 submitted by Justin Brennan
Double-stranded RNA spatial dysregulation and innate immune activation by a SARS-CoV-2 defective viral genome
Justin W. Brennan (Division of Immunobiology, Cincinnati Childrens Hospital), Yan Sun (Department of Microbiology & Immunology, University of Rochester Medical Center)
Abstract:
Defective viral genomes (DVGs) are truncated derivatives of their parental viral genomes that are incapable of productive infection without a co-infecting standard helper virus. SARS-CoV-2 infection generates deletion-type DVGs that arise from non-homologous recombination events during viral genomic replication. The role of DVGs in SARS-CoV-2 pathogenesis, specifically in innate immune activation, is largely unknown. Our group previously identified conserved hotspots (termed A, B, and C) clustering with SARS-CoV-2 deletion DVG recombination sites. The DVG population arising from hotspot B (DVG-B) bears a deletion ablating the ORF7a to the nucleocapsid protein (N) genomic region. To characterize the functions of DVG-B during infection, we generated virus-like particles specifically packaging DVG-B genomes. We observed that DVG-B robustly induced types I and III interferon (IFN) and IFN stimulated gene expression, relative to wild-type SARS-CoV-2 (WT) in A549-ACE2 cells. Despite its greater IFN-induction activity, by immunofluorescence DVG-B infected cells exhibited lower dsRNA levels compared to WT. Interestingly, DVG-B-derived dsRNA localized to discrete puncta throughout the cell, while WT virus derived dsRNA was perinuclear. Compared to WT, DVG-B infection induced significantly greater IRF-3 nuclear translocation than SARS-CoV-2 in a MAVS dependent manner, suggesting more efficient activation of the RIG-I-like receptor (RLR) pathway by DVG-B-derived dsRNA. Interestingly, RIG-I was identified as the primary sensor detecting DVG-B-derived dsRNA. DVG-B infection complemented with N partially restored the perinuclear dsRNA phenotype typical of WT infection but, unexpectedly, enhanced IFN-induction. Overall, these findings suggest that the N protein plays a key role in regulating SARS-CoV-2 dsRNA localization and that a more complex relationship exists between dsRNA localization and RLR activation during SARS-CoV-2 infection.
Keywords: dsRNA, Interferon, RIG-I-like receptors