MDA5

Abstract: In the autoimmune disease systemic lupus erythematosus (SLE), our normal antiviral defenses are inappropriately activated, resulting in over-activity of the type I interferon (IFN) pathway. This increased activity of the type I IFN pathway is an important primary pathogenic factor in the disease. Emerging evidence has implicated the antiviral helicases in this process. The antiviral helicases normally function as nucleic acid receptors in viral immunity. Genetic variations in antiviral helicase genes have been associated with SLE, supporting the idea that helicase pathways are involved in the primary pathogenesis of SLE. Studies have documented functional consequences of these genetic variations within the type I IFN pathway in human cell lines and SLE patients. In this review, we summarize the function of helicases in the anti-viral immune response, and how this response is dysregulated in SLE patients. In particular, we will focus on known functional genetic polymorphisms in the IFIH1 (MDA5) and mitochondrial antiviral signaling protein genes which have been implicated in human SLE. These data provide fascinating evidence for dysregulation of helicase-mediated innate immunity in SLE, and may support novel therapeutic strategies in the disease.

Abstract: Because of their potent regenerative and immunomodulatory properties, mesenchymal stem cells (MSCs) have promising therapeutic benefits in clinical treatment of inflammatory and infectious diseases. Recent studies suggest that many biological activities of MSCs are largely determined by pattern recognition receptors (PRRs), such as Toll-like receptors (TLRs). However, the role of PRRs in regulating the survival of MSCs remains unknown. In the present study, we examined the viability of MSCs after stimulation of distinct PRRs. Activation of TLRs by direct addition with their respective ligands showed no significant effect on the survival of MSCs, whereas transfection with double-stranded RNA (dsRNA) resulted in marked cell death in MSCs. Transfection of dsRNA upregulated cytosolic retinoic acid inducible gene I (RIG-I)-like receptors (RLRs), including RIG-I and melanoma differentiation-associated antigen 5 (MDA5). Moreover, transfection of dsRNA activated downstream transcription factors interferon regulatory factor 3 (IRF3) and nuclear factor κB (NF-κB), as well as induced the expression of interferon-β (IFN-β) and pro-inflammatory cytokine interleukin 6 (IL-6) via RLR signaling. Furthermore, we found that transfection of dsRNA triggered both extrinsic and intrinsic apoptotic responses via RLRs. However, ectopic expression of RIG-I or MDA5 was not sufficient to induce apoptosis of MSCs without dsRNA transfection. Our study also revealed that IκB kinase α/β (IKKα/β) was required for RLR-mediated apoptosis in MSCs, while TANK-binding kinase 1 (TBK1)/IKKɛ served a pro-survival role. Moreover, neither overexpression of B-cell lymphoma 2 (Bcl2) nor neutralizing autocrined IFN-β reduced RLR-mediated apoptosis. In addition, autophagy was induced upon activation of RLRs, however, blocking autophagy did not rescue MSCs from the dsRNA-induced cell death. To the best of our knowledge, this is the first study to explore the role of RLRs in controlling the survival of MSCs, which may provide a clue to understand the pathogenesis of viral infection in MSCs.

Abstract: Coronaviruses generate double-stranded (ds) RNA intermediates during viral replication that can activate host immune sensors. To evade activation of the host pattern recognition receptor MDA5, coronaviruses employ Nsp15, which is a uridine-specific endoribonuclease. Nsp15 is proposed to associate with the coronavirus replication-transcription complex within double-membrane vesicles to cleave these dsRNA intermediates. How Nsp15 recognizes and processes dsRNA is poorly understood because previous structural studies of Nsp15 have been limited to small single-stranded (ss) RNA substrates. Here we present cryo-EM structures of SARS-CoV-2 Nsp15 bound to a 52nt dsRNA. We observed that the Nsp15 hexamer forms a platform for engaging dsRNA across multiple protomers. The structures, along with site-directed mutagenesis and RNA cleavage assays revealed critical insight into dsRNA recognition and processing. To process dsRNA Nsp15 utilizes a base-flipping mechanism to properly orient the uridine within the active site for cleavage. Our findings show that Nsp15 is a distinctive endoribonuclease that can cleave both ss- and dsRNA effectively.