BTK: sensing pathogenic nucleic acids

We are under constant threats from pathogens. A failure to initiate an appropriate immune response will lead to an immunocompromised state. Our innate immune system could sense foreign nucleic acids from parasites, viruses, fungi and bacteria. This is achieved through pattern recognition receptors (PRRs) on innate cells such as macrophages that recognise different pathogen-associated molecular patterns including those found on foreign nucleic acids. Viral double-stranded RNA (dsRNA) in particular are recognised by PRRs such as endosomal Toll-like receptor (TLR)-3 and cytosolic RIG-I-like receptors (RLRs), RIG-I. Long dsRNA binds to TLR3 leading to receptor dimerization and phosphorylation of two tyrosine residues in its intracellular domain, Tyr759 and Tyr858 [1]. TRIF adaptor downstream of TLR3 subsequently recruits TBK1 to activate Interferon regulatory factor (IRF) 3. PI3K is reported to bind to phosphorylated TLR3 at Tyr759 [1] and activate downstream AKT for signaling to IRF3 as well. In addition, RIP1 binds to the C-terminus of TRIF to activate NFκB signaling. Finally, MAP kinases are phosphorylated downstream of TRIF for AP-1 signaling and together with NFκb and IRF3 transcription factors, cooperatively activate Type 1 IFN production, particularly IFN-β, critical for antiviral response [2]. Since TLR3 signaling is complex, we hypothesized that more molecules could be involved. Bruton's tyrosine kinase (BTK) is critical for B cell development and mutations in BTK leads to X-linked agammaglobulinaemia (XLA) in humans and X-linked immunodeficiency in mice [3]. XLA patients were also observed to develop recurrent bacterial and viral infections suggesting a possible role for BTK in innate immunity. Indeed BTK were found to be activated by several TLRs that signal through the adaptor MyD88 [4]. TLR3 signaling is unique as it strictly uses only the TRIF adaptor, and hence presented a good system to examine if BTK plays a role in TRIF-signaling [5]. We found D-galactosamine sensitised BTK knockout mice to survive better than wildtype mice when challenged with poly(I:C). Ex vivo experiments using macrophages from wildtype and btk-/-mice stimulated with naked poly(I:C) revealed that the production of inflammatory cytokines and IFN-β were defective in the absence of BTK. To corroborate this finding, we further infected wildtype and btk-/-macrophages with dengue viruses. Consistent with prior observations of the requirement of BTK in TRIF signaling for Type 1 IFN production, dengue virus infected btk-/-macrophages were found to have defective IFN-β mRNA upregulation and unable to clear dengue virus infection. To gain insight into BTK's role in TLR3 signaling, we …