SARS-CoV-2–associated ssRNAs activate inflammation and immunity via TLR7/8
Valentina Salvi, … , Silvano Sozzani, Daniela Bosisio
Valentina Salvi, … , Silvano Sozzani, Daniela Bosisio
Published August 6, 2021
Citation Information: JCI Insight. 2021;6(18):e150542. https://doi.org/10.1172/jci.insight.150542.
View: Text | PDF
Research Article Immunology

SARS-CoV-2–associated ssRNAs activate inflammation and immunity via TLR7/8

Abstract

The inflammatory and IFN pathways of innate immunity play a key role in the resistance and pathogenesis of coronavirus disease 2019 (COVID-19). Innate sensors and SARS-CoV-2–associated molecular patterns (SAMPs) remain to be completely defined. Here, we identified single-stranded RNA (ssRNA) fragments from the SARS-CoV-2 genome as direct activators of endosomal TLR7/8 and MyD88 pathway. The same sequences induced human DC activation in terms of phenotype and function, such as IFN and cytokine production and Th1 polarization. A bioinformatic scan of the viral genome identified several hundreds of fragments potentially activating TLR7/8, suggesting that products of virus endosomal processing potently activate the IFN and inflammatory responses downstream of these receptors. In vivo, SAMPs induced MyD88-dependent lung inflammation characterized by accumulation of proinflammatory and cytotoxic mediators and immune cell infiltration, as well as splenic DC phenotypical maturation. These results identified TLR7/8 as a crucial cellular sensor of ssRNAs encoded by SARS-CoV-2 involved in host resistance and the disease pathogenesis of COVID-19.

Authors

Valentina Salvi, Hoang Oanh Nguyen, Francesca Sozio, Tiziana Schioppa, Carolina Gaudenzi, Mattia Laffranchi, Patrizia Scapini, Mauro Passari, Ilaria Barbazza, Laura Tiberio, Nicola Tamassia, Cecilia Garlanda, Annalisa Del Prete, Marco A. Cassatella, Alberto Mantovani, Silvano Sozzani, Daniela Bosisio

×

Figure 4

SAMPs activate the TLR8/MyD88/NF-κB axis in moDCs.

Options: View larger image (or click on image) Download as PowerPoint
SAMPs activate the TLR8/MyD88/NF-κB axis in moDCs. (A) HEK293 cells stab...
(A) HEK293 cells stably transfected with human TLR7, TLR8, or luciferase alone (-) were stimulated with SCV2-RNA for 24 hours. NF-κB activation was evaluated as luciferase activity. Data are expressed as mean ± SEM (n = 3); *P < 0.05 versus (-) by 1-way ANOVA with Dunnett’s post hoc test. (B–E) moDCs were stimulated with SCV2-RNA as indicated (B) or pretreated with CU-CPT9a (1 μM) and then stimulated with SCV2-RNA or LPS for 1 hour (E). Nuclear extracts were blotted against NF-κB p65 and lamin B. One representative donor and densitometry of 3 donors are shown. *P < 0.05 versus untreated by 1-way ANOVA with Dunnett’s post hoc test; #P < 0.05 versus “SCV2-RNA” by paired Student’s t test. (C, left panel) moDCs were transfected with indicated siRNAs and target gene expression evaluated by qPCR. Results depict percentage of target gene expression (mean ± SEM n = 4). (C, right panel) moDCs transfected with indicated siRNAs were stimulated with SCV2-RNA for 24 hours and IL-6 production evaluated by ELISA. Data are expressed as percentage of production (n = 4); *P < 0.05 versus “ctr siRNA” by 1-way ANOVA with Dunnett’s post hoc test. (D) moDCs were transfected with indicated siRNAs and the expression of TLR8 was evaluated by qPCR (left y axis, white bars). IL-6 production upon SCV2-RNA stimulation was evaluated by ELISA (right y axis, gray bars). Data (percentage of expression/production) represent the mean ± SEM (n = 3); *P < 0.05 versus respective “ctr” by paired Student’s t test. (F) moDCs were pretreated with CU-CPT9a, then stimulated as indicated for 24 hours and IL-6 production evaluated by ELISA. Data are expressed as percentage of production for each individual stimulation (n = 3); *P < 0.05 versus respective “0” by 1-way ANOVA with Dunnett’s post hoc test.