Conformational rearrangements of RIG-I receptor on formation of a multiprotein:dsRNA assembly

Abstract

The retinoic acid inducible gene-I (RIG-I)-like family of receptors is positioned at the front line of our innate cellular defence system. RIG-I detects and binds to foreign duplex RNA in the cytoplasm of both immune and non-immune cells, and initiates the induction of type I interferons and pro-inflammatory cytokines. The mechanism of RIG-I activation by double-stranded RNA (dsRNA) involves a molecular rearrangement proposed to expose the N-terminal pair of caspase activation recruitment domains, enabling an interaction with interferon-beta promoter stimulator 1 (IPS-1) and thereby initiating downstream signalling. dsRNA is particularly stimulatory when longer than 20 bp, potentially through allowing binding of more than one RIG-I molecule. Here, we characterize full-length RIG-I and RIG-I subdomains combined with a stimulatory 29mer dsRNA using multi-angle light scattering and size-exclusion chromatography–coupled small-angle X-ray scattering, to build up a molecular model of RIG-I before and after the formation of a 2:1 protein:dsRNA assembly. We report the small-angle X-ray scattering–derived solution structure of the human apo-RIG-I and observe that on binding of RIG-I to dsRNA in a 2:1 ratio, the complex becomes highly extended and flexible. Hence, here we present the first model of the fully activated oligomeric RIG-I.

Master sensors of pathogenic RNA – RIG-I like receptors

“…Based on multi-angle light scattering and sizeexclusion chromatography-coupled small-angle X-ray scattering of RIG-I/29mer dsRNA complexes, Beckham e al. suggested that binding of a first RIG-I molecule changes the RNA structure, thus constructively influencing the binding of a second RIG-I molecule (Beckham et al 2013).…”

Defining the functional determinants for RNA surveillance by RIG‐I

“…Multimerization of RIG-I on RNA has been observed using a variety of in vitro techniques [22][23][24]. However, the fact that several proteins can bind one RNA molecule does not mean that they are all functional, it does not mean that the proteins interact with each other and it does not imply that they bind cooperatively to RNA.…”

Ex vivo cytokine production in peripheral blood mononuclear cells after their stimulation with dsRNA of natural origin

“…The production of almost all of the studied cytokines appears to be dependent on the concentration of Larifan used, with the efficacy of induction or suppression decreasing when higher concentrations are used. This “bell‐shaped” response type is well known in the context of dsRNA‐induced IFN production and can be explained by the activation mechanism that involves dimerization and multimerization with dsRNA molecules for most of the dsRNA detecting receptors . It is of interest that, among molecules suppressed by Larifan, there clearly appeared different concentration dependencies, with 200 µg/mL of Larifan being the most effective concentration for suppressing IL‐16 and 400 µg/mL for suppressing IP‐10 and SDF1‐a+b.…”