HOIL1 Is Essential for the Induction of Type I and III Interferons by MDA5 and Regulates Persistent Murine Norovirus Infection

Abstract

The linear ubiquitin chain assembly complex (LUBAC), composed of heme-oxidized IRP2 ubiquitin ligase 1 (HOIL1), HOIL1-interacting protein (HOIP), and SHANK-associated RH domain-interacting protein (SHARPIN), is a crucial regulator of multiple immune signaling pathways. In humans, HOIL1 or HOIP deficiency is associated with an immune disorder involving autoinflammation, immunodeficiency, and inflammatory bowel disease (IBD)-like symptoms. During viral infection, LUBAC is reported to inhibit the induction of interferon (IFN) by the cytosolic RNA sensor retinoic acid-inducible gene I (RIG-I). Surprisingly, we found that HOIL1 is essential for the induction of both type I and type III IFNs, as well as the phosphorylation of IFN regulatory factor 3 (IRF3), during murine norovirus (MNoV) infection in cultured dendritic cells. The RIG-I-like receptor, melanoma differentiation-associated protein 5 (MDA5), is also required for IFN induction and IRF3 phosphorylation during MNoV infection. Furthermore, HOIL1 and MDA5 were required for IFN induction after Theiler's murine encephalomyelitis virus infection and poly(I·C) transfection, but not Sendai virus or vesicular stomatitis virus infection, indicating that HOIL1 and LUBAC are required selectively for MDA5 signaling. Moreover, mice exhibited defective control of acute and persistent murine norovirus infection and defective regulation of MNoV persistence by the microbiome as also observed previously for mice deficient in interferon lambda (IFN-λ) receptor, signal transducer and activator of transcription factor 1 (STAT1), and IRF3. These data indicate that LUBAC plays a critical role in IFN induction to control RNA viruses sensed by MDA5. Human noroviruses are a leading cause of gastroenteritis throughout the world but are challenging to study and Murine norovirus (MNoV) provides a tractable genetic and small-animal model to study norovirus biology and immune responses. Interferons are critical mediators of antiviral immunity, but excessive expression can dysregulate the immune system. IFN-λ plays an important role at mucosal surfaces, including the gastrointestinal tract, and both IFN-λ and commensal enteric bacteria are important modulators of persistent MNoV infection. LUBAC, of which HOIL1 is a component, is reported to inhibit type I IFN induction after RIG-I stimulation. We show, in contrast, that HOIL1 is critical for type I and III IFN induction during infection with MNoV, a virus that preferentially activates MDA5. Moreover, HOIL1 regulates MNoV infection These data reveal distinct functions for LUBAC in these closely related signaling pathways and in modulation of IFN expression.

Molecular bases for HOIPINs-mediated inhibition of LUBAC and innate immune responses

“…Various PAMPs activate the Toll-like receptors (TLRs)mediated NF-κB and IFN antiviral pathways 1 . Although LUBAC activity reportedly down-regulate the type I IFN production pathway 32,33 , recent studies suggested that the LUBAC is necessary for the TLR-mediated interferon regulatory factor 3 (IRF3) activation [34][35][36] . Among the PAMPs, HOIPIN-1 suppressed the NF-κB activation induced by CpG, a ligand for TLR9 (ref.…”

“…Although the effects of LUBAC on the IFN antiviral pathway have been controversial, several recent reports suggested that the LUBAC activity is crucial for the IRF3 activation [34][35][36] . Moreover, OTULIN, a linear ubiquitin-specific deubiquitinase, was revealed to down-regulate the IFN induction pathway 52 , suggesting that linear ubiquitination seems to regulate antiviral pathway.…”

Norovirus Replication in Human Intestinal Epithelial Cells Is Restricted by the Interferon-Induced JAK/STAT Signaling Pathway and RNA Polymerase II-Mediated Transcriptional Responses

“…The mechanism by which HuNoV is sensed by the infected cells is not currently known; however, data from MNV suggest a clear role for Mda5-mediated sensing in the restriction of norovirus replication both in cell culture and in vivo (61). The sensing of MNV RNA occurs in a process that requires the HOIL1 component of the linear ubiquitin chain assembly complex (LUBAC) complex (62). Other components of the RNA sensing pathways, including mitochondrial antiviral-signaling protein (MAVS), IRF3, and IRF7, have been implicated in the innate response to MNV (61,62), but the role that they play in sensing of HuNoV RNA is unknown.…”

“…The sensing of MNV RNA occurs in a process that requires the HOIL1 component of the linear ubiquitin chain assembly complex (LUBAC) complex (62). Other components of the RNA sensing pathways, including mitochondrial antiviral-signaling protein (MAVS), IRF3, and IRF7, have been implicated in the innate response to MNV (61,62), but the role that they play in sensing of HuNoV RNA is unknown. In addition to targeting STAT1 for degradation (63), the PIV5 V protein is known to also inhibit the activity of Mda5 (35).…”

Vero Cells as a Mammalian Cell Substrate for Human Norovirus

“…RAD51AP1 facilitates homologous recombination of host DNA [107], while SEC61G functions in membrane protein translocation and incorporation into the endoplasmic reticulum [108]. The interactions between HuNoV and these genes are speculative; however IRF3 [109,110], IRF7 [110,111], MDA5 [112][113][114][115], RIG-I [35,111,113,114], TLR3 [112], and TLR7 [114,116,117] have been previously implicated in NoV responses. Of note, TLR2 enhances cytokine production upon the detection of rotavirus [118].…”