Research Information System
5th International Molecular Immunology & Immunogenetics Congress, İzmir, Turkey, 20 - 22 October 2022, pp.11, (Summary Text)
Infection with SARS-CoV-2 inhibits early type I IFN response but activates inflammasome signaling and pro-inflammatory cytokine production. To this end, in addition to structural protein Nucleocapsid, different accessory and non-structural proteins’ effect on immunomodulation was assessed by using lentiviral gene transduction system. For stable protein expression in THP1-Dual cell line, plasmids for the production of lentiviral vectors that provide packaging and envelope protein expression were combined with SARS-CoV-2 protein plasmids. Cell lines stably expressing, NSP9, NSP10, ORF3a, ORF8, Nucleocapsid, and their mock transduced control were then exposed to virus-mimetic pattern recognition receptor ligands or to recombinant IFN-β to measure type I IFN and type-I-dependent ISG15 production, respectively. Furthermore, to evaluate the impact of SARS-CoV-2-related protein expression on NLRP3, NLRC4, AIM2, or non-canonical inflammasome activation were stimulated with corresponding activating ligands. Cellular IL-1β cytokine production and lactate dehydrogenase enzyme (LDH) release were analyzed as indicators of inflammasome activation and pyroptosis, respectively. RESULTS Our results indicate that ORF3a and ORF8 accessory proteins significantly antagonized type I IFN production and ORF3a downregulated ISG15 production. Specifically, ORF8 expression downmodulated IFN production 3- and 2-fold in response to RIG-I and TLR7/8 agonists, respectively. Moreover, NSP9 and N were found to antagonize type I IFN production only when compared to WT, but not to MOCK. Results related to inflammasome activation indicated showed that ORF3a accessory protein significantly increased NLRP3 and NLRC4 inflammasome-mediated IL-1β production (5- to 10-fold) and LDH release. study suggests that ORF3a and ORF8 accessory proteins can be categorized as type I IFN antagonists. Furthermore, ORF3a accessory protein facilitated NLRP3 and NLRC4 inflammasome-mediated IL-1β production and LDH release, indicating that the ORF3a viroprotein could contribute to the increased inflammatory response in the host.