A Novel Targeted RIG-I Receptor 5'Triphosphate Double Strain RNA-Based Adjuvant Significantly Improves the Immunogenicity of the SARS-CoV-2 Delta-Omicron Chimeric RBD-Dimer Recombinant Protein Vaccine.

The rapid mutation and spread of SARS-CoV-2 variants recently, especially through the emerging variants Omicron BA5, BF7, XBB and BQ1, necessitate the development of universal vaccines to provide broad spectrum protection against variants. For the SARS-CoV-2 universal recombinant protein vaccines, an effective approach is necessary to design broad-spectrum antigens and combine them with novel adjuvants that can induce high immunogenicity. In this study, we designed a novel targeted retinoic acid-inducible gene-I (RIG-I) receptor 5'triphosphate double strain RNA (5'PPP dsRNA)-based vaccine adjuvant (named AT149) and combined it with the SARS-CoV-2 Delta and Omicron chimeric RBD-dimer recombinant protein (D-O RBD) to immunize mice. The results showed that AT149 activated the P65 NF-κB signaling pathway, which subsequently activated the interferon signal pathway by targeting the RIG-I receptor. The D-O RBD + AT149 and D-O RBD + aluminum hydroxide adjuvant (Al) + AT149 groups showed elevated levels of neutralizing antibodies against the authentic Delta variant, and Omicron subvariants, BA1, BA5, and BF7, pseudovirus BQ1.1, and XBB compared with D-O RBD + Al and D-O RBD + Al + CpG7909/Poly (I:C) groups at 14 d after the second immunization, respectively. In addition, D-O RBD + AT149 and D-O RBD + Al + AT149 groups presented higher levels of the T-cell-secreted IFN-γ immune response. Overall, we designed a novel targeted RIG-I receptor 5'PPP dsRNA-based vaccine adjuvant to significantly improve the immunogenicity and broad spectrum of the SARS-CoV-2 recombinant protein vaccine.

DEAD-box RNA helicase 21 negatively regulates cytosolic RNA-mediated innate immune signaling.

DEAD-box RNA helicase 21 (DDX21), also known as RHII/Gu, is an ATP-dependent RNA helicase. In addition to playing a vital role in regulating cellular RNA splicing, transcription, and translation, accumulated evidence has suggested that DDX21 is also involved in the regulation of innate immunity. However, whether DDX21 induces or antagonizes type I interferon (IFN-I) production has not been clear and most studies have been performed through ectopic overexpression or RNA interference-mediated knockdown. In this study, we generated DDX21 knockout cell lines and found that knockout of DDX21 enhanced Sendai virus (SeV)-induced IFN-ß production and IFN-stimulated gene (ISG) expression, suggesting that DDX21 is a negative regulator of IFN-ß. Mechanistically, DDX21 competes with retinoic acid-inducible gene I (RIG-I) for binding to double-stranded RNA (dsRNA), thereby attenuating RIG-I-mediated IFN-ß production. We also identified that the 217-784 amino acid region of DDX21 is essential for binding dsRNA and associated with its ability to antagonize IFN production. Taken together, our results clearly demonstrated that DDX21 negatively regulates IFN-ß production and functions to maintain immune homeostasis.

A Comparative Analysis of Coronavirus Nucleocapsid (N) Proteins Reveals the SADS-CoV N Protein Antagonizes IFN-ß Production by Inducing Ubiquitination of RIG-I.

Coronaviruses (CoVs) are a known global threat, and most recently the ongoing COVID-19 pandemic has claimed more than 2 million human lives. Delays and interference with IFN responses are closely associated with the severity of disease caused by CoV infection. As the most abundant viral protein in infected cells just after the entry step, the CoV nucleocapsid (N) protein likely plays a key role in IFN interruption. We have conducted a comprehensive comparative analysis and report herein that the N proteins of representative human and animal CoVs from four different genera [swine acute diarrhea syndrome CoV (SADS-CoV), porcine epidemic diarrhea virus (PEDV), severe acute respiratory syndrome CoV (SARS-CoV), SARS-CoV-2, Middle East respiratory syndrome CoV (MERS-CoV), infectious bronchitis virus (IBV) and porcine deltacoronavirus (PDCoV)] suppress IFN responses by multiple strategies. In particular, we found that the N protein of SADS-CoV interacted with RIG-I independent of its RNA binding activity, mediating K27-, K48- and K63-linked ubiquitination of RIG-I and its subsequent proteasome-dependent degradation, thus inhibiting the host IFN response. These data provide insight into the interaction between CoVs and host, and offer new clues for the development of therapies against these important viruses.