Human TLR3 Dual Reporter HEK 293 Cells

In humans, four Toll-Like Receptor (TLR) family members TLR3, TLR7, TLR8, and TLR9 are specialized in sensing viral-derived components and are mainly found in the endosome. Among these, TLR3 recognizes double-stranded (ds)RNA, a hallmark of viral replication, and triggers antiviral immune responses [1]. TLR3 is expressed in myeloid dendritic cells, macrophages, as well as non-immune cells [2].

TLR3 activation upon viral infection involves several steps, including translocation of TLR3 from the ER (endoplasmic reticulum) via the Golgi to the endosome, proteolytic cleavage and dimerization of TLR3, and finally receptor-ligand binding [3]. In order to start the signaling cascade, activated TLR3 recruits the adaptor protein TRIF (TIR domain-containing adapter-inducing interferon-β). TRIF binds to TRAF3 (TNF receptor-associated factor 3), which then recruits TBK1 (TANK-binding kinase 1) and IKKε (IκB kinase ε), thus activating the transcription factor IRF3 (interferon regulatory factor 3) and stimulating the production of type I IFNs (interferons). Additionally, TRIF interacts with TRAF6 and RIP1 (kinase receptor-interacting protein 1). RIP1 in turn binds to TAK1 (transforming growth factor β-activated kinase 1) and IKK. TAK1 phosphorylates IKKα and IKKβ, leading to the phosphorylation of IκB, the NF-κB inhibitor. Ultimately, this leads to the release and translocation of NF-κB into the nucleus and the induction of pro-inflammatory cytokines [2,4].

Given its important role in dsRNA recognition, TLR3 signaling has been intensively studied. Various TLR3-agonists, such as the synthetic dsRNA analog Poly(I:C) are being used in vaccine development and cancer therapy [4]. Yet, recent studies have indicated that TLR3 may act as a double-edged sword by showing both protective and damaging functions in the context of some human viral infections [3,5]. Moreover, rare mutations in TLR3 have been associated with viral susceptibility; specifically, infections with HSV-1 (herpes simplex virus 1), influenza, and SARS-Co-V2 have been linked to pathogenic germline variants in TLR3 pathway genes [2]. Understanding the TRIF-dependent TLR3 pathway may be essential for the establishment of specific therapeutic approaches to diminish TLR3-driven disease and exploit its protective functions [3].

References

1. Manuela Sironi, et al., 2012. A Common Polymorphism in TLR3 Confers Natural Resistance to HIV-1 Infection. J Immunol 15; 188 (2): 818–823. 
2. Aluri, J, et al., 2021. Toll-Like Receptor Signaling in the Establishment and Function of the Immune System. Cells, 10, 1374.
3. Chen Y, et al., 2021.  Toll-like receptor 3 (TLR3) regulation mechanisms and roles in antiviral innate immune responses. J Zhejiang Univ Sci B.;22(8):609-632.
4. Komal A, et al., 2021. TLR3 agonists: RGC100, ARNAX, and poly-IC: a comparative review. Immunol Res. 69(4):312-322. 
5. Perales-Linares R, Navas-Martin S. 2013. Toll-like receptor 3 in viral pathogenesis: friend or foe? Immunology.;140(2):153-67.