Abstract
The RLRs play critical roles in sensing and fighting viral infections especially RNA virus infections. Despite the extensive studies on RLRs in humans and mice, there is a lack of systemic investigation of livestock animal RLRs. In this study, we characterized the porcine RLR members RIG-I, MDA5 and LGP2. Compared with their human counterparts, porcine RIG-I and MDA5 exhibited similar signaling activity to distinct dsRNA and viruses, via similar and cooperative recognitions. Porcine LGP2, without signaling activity, was found to positively regulate porcine RIG-I and MDA5 in transfected porcine alveolar macrophages (PAMs), gene knockout PAMs and PK-15 cells. Mechanistically, LGP2 interacts with RIG-I and MDA5 upon cell activation, and promotes the binding of dsRNA ligand by MDA5 as well as RIG-I. Accordingly, porcine LGP2 exerted broad antiviral functions. Intriguingly, we found that porcine LGP2 mutants with defects in ATPase and/or dsRNA binding present constitutive activity which are likely through RIG-I and MDA5. Our work provided significant insights into porcine innate immunity, species specificity and immune biology.
Highlights
The innate immune system, as the first line of host defense, relies on the pattern recognition receptors (PRRs) to sense various danger signals from pathogens via detecting the pathogen associated molecular patterns (PAMPs) [1]
We isolated and cloned porcine RIG-I-like receptors (RLRs) RIG-I, MDA5 and LGP2, which were deposited into GenBank with the accession numbers
In terms of signaling activity, we found that both porcine RIG-I and MDA5 exhibited constitutive signaling activity in ISRE and NF-kB promoter assay, with MDA5 harboring higher constitutive activity (Figures 1B, D)
Summary
The innate immune system, as the first line of host defense, relies on the pattern recognition receptors (PRRs) to sense various danger signals from pathogens via detecting the pathogen associated molecular patterns (PAMPs) [1]. The prototypic RIG-I as well as MDA5 were discovered more than 15 years ago in overexpression studies to induce type I interferons (IFNs) [4, 5]. Both RIG-I and MDA5 have similar domain architecture: N-terminal signal active two tandem caspase recruitment domain (CARDs), middle DExD/Hbox helicase domain and C-terminal repressor domain (CTD) or regulatory domain (RD) [6, 7]. Upon recognition of dsRNA, RIG-I and MDA5 activate downstream common adaptor MAVS, which recruit TRAF3/TBK1/IKKe and TRAF6/IKKs to activate transcription factors IRF3 and NF-kB, driving IFN and proinflammatory cytokine expression, respectively [10]
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