Herpes simplex virus (HSV) has evolved multiple strategies to modulate host immune responses. In a screen of HSV open reading frames to identify additional HSV-encoded proteins that affect NF-κB signaling, we identified the viral US3 tegument protein as an inhibitor of NF-κB signaling. We found that the US3 protein is required for inhibition of TLR2 signaling induced by viral infection and that this inhibition occurs at very early times post-infection. Expression of US3 in transfected cells inhibits TLR2 signaling induced by Zymosan, and this inhibition occurs at or downstream of MyD88 and upstream of p65. Polyubiquitination of TRAF6 is critical for its function in TLR2 signaling. Using US3-null and US3 kinase-defective mutant viruses, we demonstrate that HSV US3 reduces TRAF6 polyubiquitination and that the kinase activity of US3 is necessary for this effect. Therefore, US3 is necessary and sufficient for inhibiting TLR2 signaling at or before the stage of TRAF6 ubiquitination.
One of the first critical lines of defense by a host organism against an invading virus is its innate immune system. The earliest events of innate immune responses include sensing of virus components by host pattern recognition receptors (PRRs), which initiate signaling cascades and transcriptional activation of genes encoding type I interferons (IFNs) and proinflammatory cytokines. These signaling pathways are complex mechanisms that engage a variety of cell types (inflammatory cells, dendritic cells and lymphocytes) to control viral infection and are tightly regulated. In addition to type I IFNs, which mediate the early antiviral response to a large extent, cytokines (like IL-1β, IL-6, IL-8, IL-18 and IL-12) induced by innate immune cells are also critical for an effective early antiviral defense. Pathogen sensing triggers a cascade of intracellular signaling events involving a large number of adaptor proteins. Sequential steps of post-translational modifications on these proteins, such as phosphorylation and ubiquitination, result in the translocation of transcription factors such as NF-κB, AP-1, or JNK to the nucleus where they stimulate the transcription of antiviral and pro-inflammatory genes. These events function to curb early events in productive viral infection as well as to program the adaptive immune response. Not surprisingly, viruses have also evolved numerous mechanisms to blunt or evade these protective measures elicited by the host. NF-κB is a major transcriptional activator for pro-inflammatory cytokine genes (Hayden et al.Glycerol Endogenous Metabolite , 2006), and herpes simplex virus (HSV) infection activates the NF-κB signaling pathway by both TLR-dependent and -independent pathways resulting in the induction of cytokines IL-6 and IL-8 (Hilton et al., 1995; Kurt-Jones et al., 2005, 2004).
Upon microbial recognition, TLR2 dimerizes with either TLR1 or TLR6 and recruits MyD88 and Mal/TIRAP through TIR domain interactions. This complex then recruits the IRAKs (IL-1 receptor-associated kinases). IRAK4 is recruited first, becomes activated and phosphorylates IRAK-1, which activates IRAK-2. IRAK activation leads to an interaction with TRAF6 (tumor necrosis factor receptor-associated factor 6) and oligomerization of TRAF6 and its autoubiquitination. TRAF6, an E3 ubiquitin ligase, catalyzes the synthesis of polyubiquitin chains (polyUb) bound to itself and TAK1, thereby activating TAK1. The polyUb chains bind to NEMO, the regulatory component of the IKK complex.2,3,5,6-Tetrafluoroterephthalic acid Biological Activity The resulting complex leads to phosphorylation of IKKα by TAK1, leading to activation of the IKK complex, which phosphorylates the β-subunit of IκB (inhibitor of κB), causing its ubiquitination and degradation, release of NF-κB and its translocation into the nucleus. Nuclear NF-κB binds to κB elements in enhancers and promoters and also to the basal transcriptional machinery to activate transcription (Oliveira-Nascimento et al., 2012; Rathinam and Fitzgerald, 2011). The TLR2 dependence for HSV induction of NF-κB signaling is cell type-specific (Rathinam and Fitzgerald, 2011). We have shown that infection with HSV-1 wild-type (WT) strains KOS and F can activate TLR2 signaling in mouse macrophages and human cells expressing TLR2 (Kurt-Jones et al., 2005, 2004). Further, while TLR2 is essential for the recognition of HSV and induction of pro-inflammatory cytokines by macrophages, microglial cells and myeloid dendritic cells (Aravalli et al., 2007, 2005; Lima et al., 2010), plasmacytoid dendritic cells (pDCs) can sense HSV in a TLR2-independent fashion (Rasmussen et al., 2007; Sato et al., 2006). Recently, it has also been reported that in response to HSV infection, type I interferon production in inflammatory monocytes is partially dependent on TLR2 (Barbalat et al., 2009). Moreover, TLR2 recognition of HSV in vivo appears to depend on route of inoculation and virus subtype. In the case of HSV-2 infection in mice, although TLR2 appears to be nonessential for the control of viral spread following intraperitoneal or vaginal infection, an effective cytokine response in the brain following natural vaginal infection is dependent on a synergistic role of TLR2 and TLR9 (Sorensen et al., 2008). In the corneal and intraperitoneal infection models in mice, TLR2 sensing of HSV has been shown to mount an excessive immune response that can be detrimental to the host (Kurt-Jones et al., 2004; Sarangi et al., 2007). Interestingly, in humans, two polymorphisms in TLR2 are associated with increased HSV-2 viral shedding and increased lesions (Bochud et al., 2007), supporting a role for TLR2 in the control of virus infection. In addition, work done by Iwasaki and colleagues indicated that TLR2 sensing of HSV-1 is virus strain/clone-dependent (Sato et al., 2006), although the molecular mechanism underlying this phenomenon is not known. It has been recently demonstrated that HSV gB and gH/gL proteins interact with TLR2, but gH/gL alone are capable of triggering NF-κB activation (Leoni et al.PMID:34699587 , 2012). HSV gene products have been shown to regulate NF-κB signaling in a number of ways. HSV infection activates NF-κB signaling, which is essential for optimal viral replication (Amici et al., 2001; Patel et al., 1998). It has been demonstrated that ICP27 is essential for NF-κB induction (Hargett et al., 2006). The virion UL37 protein was shown to activate NF-κB signaling by interacting with and activating TRAF6 (Liu et al., 2008). Infection with UV-inactivated virus and binding of gD to HVEM can also lead to activation of NF-κB (Medici et al., 2003; Sciortino et al., 2008). In contrast, HSV-1 ICP0 inhibited NF-κB signaling by reducing levels of adaptor proteins (van Lint et al., 2010). Therefore, the net induction of NF-κB signaling by HSV is the result of the combined activities of HSV proteins that both activate and inhibit NF-κB signaling.
In this study, in a screen of the HSV open reading frames (ORFs) to identify additional proteins that modulate NF-κB activity, we identified the HSV US3 tegument protein as an inhibitor of the NF-κB signaling pathway. The HSV-1 US3 protein kinase is a virion tegument protein that has been implicated in a variety of processes during productive virus infection. Most notable among them are its roles in virion maturation and egress (Favoreel et al., 2005; Matsuzaki et al., 2005; Mou et al., 2007; Reynolds et al., 2002; Wisner et al., 2009), in preventing virus-induced apoptosis (Benetti and Roizman, 2004; Hata et al., 1999; Leopardi et al., 1997; Munger and Roizman, 2001; Ogg et al., 2004), enhancing histone acetylation (Gu et al., 2005; Poon et al., 2006), suppression of the IFNα response by downregulating the IFNAR-1 receptor (Liang and Roizman, 2008), and more recently, in stimulating mRNA translation (Chuluunbaatar et al., 2010). In this study we show that US3 also down-regulates TLR2-dependent NF-κB signaling at very early times post infection, and we found that this inhibition of TLR2 signaling occurs before or at the stage of TRAF6 ubiquitination.MedChemExpress (MCE) offers a wide range of high-quality research chemicals and biochemicals (novel life-science reagents, reference compounds and natural compounds) for scientific use. We have professionally experienced and friendly staff to meet your needs. We are a competent and trustworthy partner for your research and scientific projects.Related websites: https://www.medchemexpress.com