Structural insights into the activation of RIG-I, a nanosensor for viral RNAs.

Animal cells use a group of innate immune sensors to detect viral invasion. The RIG-I-like receptor (RLR) family of RNA helicases is largely responsible for detecting replicating viral RNAs in the cytosol of most, if not all, cells infected with RNA viruses (Rehwinkel & Reis e Sousa, 2010). Two RLRs, RIG-I and MDA5, recognize different types of viral RNA, and thus launch antiviral immune responses against different families of RNA virus. RLRs belong to the helicase superfamily 2 (SF2), and have conserved motifs in their two SF2 domains (Hel1 and Hel2), as well as RNA-dependent ATPase activity (FairmanWilliams et al, 2010). The binding of viral RNA to the carboxy-terminal domain (CTD) of RLRs has been proposed to trigger a conformational change that exposes the aminoterminal caspase activation and recruitment domains (CARDs), which then activate the mitochondrial adaptor protein MAVS to induce type-I interferons and other antiviral molecules. Four recent studies, including one published in a recent issue of EMBO reports by the Hopfner group, provide a detailed view of how RIG-I binds to RNA and the conformational changes that lead to its activation (Fig 1; Civril et al, 2011; Jiang et al, 2011; Kowalinski et al, 2011; Luo et al, 2011). Although using different fragments of the RNA sensor, all four groups determined the crystal structures of RIG-I helicase domains. In addition, Kowalinski and colleagues obtained the structure of fulllength RIG-I from duck in the ligand-free state. Comparison of the full-length RIG-I structure in the absence of RNA with the helicase structures in RNA-bound states reveals clear snapshots of the RNA-induced conformational rearrangement. However, before we discuss these beautiful structures, it is important to note that the RNA ligands able to activate RIG-I to induce type I interferons usually contain 5’-triphosphate (5’-ppp), a signature present in replicating viral RNAs but not in host cellular RNAs, which normally contain 5’-modifications— such as a 5’-cap in mRNA (Rehwinkel & Reis e Sousa, 2010). The RNAs used in the structural studies are short double-stranded RNAs (dsRNAs) lacking 5’-ppp, and probably would not induce type I interferons. In addition, we still lack a crystal structure of RNA-bound full-length RIG-I. Therefore, the recent structures of RNA-bound RIG-I helicase might not represent its active form. Furthermore, even after bound to 5’-ppp dsRNA, RIG-I remains inactive until its N-terminal CARDs bind to Lys 63-linked polyubiquitin chains that are not anchored to any cellular protein (Zeng et al, 2010). Thus, to distinguish the dsRNA-bound RIG-I from fully active RIG-I, we refer to the conformation of dsRNA-bound RIG-I as the ‘competent state’. In the absence of RNA, full-length RIG-I adopts an autoinhibited conformation (Kowalinski et al, 2011). Two CARDs, joined head-to-tail, form one rigid unit that is attached to a unique insertion domain in Hel2 (Hel2i) through an extensive interaction interface between Hel2i and the second CARD (C2). The C2–Hel2i interaction on the two apposing surfaces is mediated by salt bridges and hydrophobic inter action patches and is therefore energetically stable. Neither the structure of the CARDs nor of the Hel2i seems to change upon interaction. This interface has significant overlap with the dsRNA–Hel2i interaction interface, suggesting that during the RNA-triggered transition, extra energy is required to disrupt the C2–Hel2i interface and enable a dsRNA to enter its binding site. On the other hand, the ubiquitin E3 ligase TRIM25, which synthesizes Lys 63 polyubiquitin chains and is required for RIG-I activation, has been shown to interact with the first CARD (C1) from the same side where Hel2i contacts the C2 domain, suggesting that Hel2i could exert steric hindrance to polyubiquitination. Furthermore, Structural insights into the activation of RIG-I, a nanosensor for viral RNAs