RNA Polymerase III Detects Cytosolic DNA and Induces Type I Interferons through the RIG-I Pathway

Abstract

SummaryType-I interferons (IFNs) are important for antiviral and autoimmune responses. Retinoic acidinduced gene I (RIG-I) and mitochondrial antiviral signaling (MAVS) proteins mediate IFN production in response to cytosolic double-stranded RNA or single-stranded RNA containing 5′-triphosphate (5′-ppp). Cytosolic B-form double-stranded DNA, such as poly(dA-dT)·poly(dA-dT) [poly(dA-dT)], can also induce IFN-β, but the underlying mechanism is unknown. Here we show that the cytosolic poly(dA-dT) DNA is converted into 5′-ppp RNA to induce IFN-β through the RIG-I pathway. Biochemical purification led to the identification of DNA-dependent RNA polymerase III (Pol-III) as the enzyme responsible for synthesizing 5′-ppp RNA from the poly(dA-dT) template. Inhibition of RNA Pol-III prevents IFN-β induction by transfection of DNA or infection with DNA viruses. Furthermore, Pol-III inhibition abrogates IFN-β induction by the intracellular bacterium Legionella pneumophila and promotes the bacterial growth. These results suggest that RNA Pol-III is a cytosolic DNA sensor involved in innate immune responses.

Type I interferonopathies—an expanding disease spectrum of immunodysregulation

“…While MDA5 binds long double-stranded (dsRNA), RIG-I ligands are rather short and characterized by a 5′-triphosphate or a 5′-diphosphate moiety with blunt-end base pairing at the 5′-end [6][7][8]. RIG-I can also recognize viral DNA via the RNA polymerase III pathway, which transcribes DNA into 5′-triphosphate RNA [9,10]. The central mechanism of cytosolic DNA sensing involves the nucleotidyl transferase cyclic GMP-AMP synthase (cGAS), which catalyzes the synthesis of the second messenger cyclic GMP-AMP (cGAMP) following binding of cGAS to dsDNA or single-stranded DNA (ssDNA) [11,12].…”

Identification of a retinoic acid-inducible gene I from Japanese eel (Anguilla japonica) and expression analysis in vivo and in vitro

“…Kong et al first reported that RIG-I À/À mice was found to be more susceptible to infection with Escherichia coli and depletion of RIG-I by RNAi or gene targeting inhibited the LPS-induced phagocytosis of bacteria [20]. Further studies have showed that RIG-I may recognize secreted bacterial nucleic acids in cytoplasm and function in antibacterial response [18,19]. In channel catfish, the most striking up-regulation of RIG-I gene expression of the liver was observed after the Gram negative bacterium Edwardsiella ictaluri infection [23].…”

“…Furthermore, several DNA viruses have now been shown to activate this pathway such as herpes-simplex virus-1 [14], Adenovirus, Epstein-Barr virus [15], hepatitis B virus [16] and Vaccinia virus [17]. Surprisingly, both of the intracellular gram negative and positive bacterium including Legionella pnuemophila and listeria monocytogenes were shown to activate type I IFN responses through RIG-I signaling [18,19]. Moreover, the bacterial lipopolysaccharide has been found to induce the expression of RIG-I in both macrophages and vascular endothelial cells [20,21].…”

Activation and regulation of pathogen sensor RIG-I

“…RNA polymerase III was found to be an additional source for ligands of RIG-I. Pol III can transcribe AT-rich DNA into 5 0 -ppp RNA intermediates that can activate RIG-I [59,60]. Notably, using a specific inhibitor for RNA polymerase III, ML60218, both studies show that Epstein-Barr virus encoded non-coding RNAs (EBER) require RNA polymerase III for interferon induction that is also dependent on RIG-I.…”

“…Notably, using a specific inhibitor for RNA polymerase III, ML60218, both studies show that Epstein-Barr virus encoded non-coding RNAs (EBER) require RNA polymerase III for interferon induction that is also dependent on RIG-I. Additionally, infection with intracellular bacteria Legionella pneumophila and Shigella flexneri also required RNA polymerase III and RIG-I for IFN activation [60,61]. These results also provide a great example of a functional collaboration between DNA sensing and RNA sensing pathways in activating antimicrobial responses in cells.…”