TLR9 stability and signaling are regulated by phosphorylation and cell stress

J Leukoc Biol. 2016 Sep;100(3):525-33. doi: 10.1189/jlb.2A0815-337R. Epub 2016 Mar 8.


Innate sensing of pathogens elicits protective immune responses through pattern recognition receptors, including Toll-like receptors. Although signaling by Toll-like receptors is regulated at multiple steps, including localization, trafficking, proteolytic cleavage, and phosphorylation, the significance of post-translational modifications and cellular stress response on Toll-like receptor stability and signaling is still largely unknown. In the present study, we investigated the role of cytoplasmic tyrosine motifs in Toll-like receptor-9 stability, proteolytic cleavage, and signaling. We demonstrated that tyrosine phosphorylation is essential for mouse Toll-like receptor-9 protein stability and signaling. Upon inhibition of tyrosine kinases with piceatannol, Toll-like receptor-9 tyrosine phosphorylation induced by CpG deoxyribonucleic acid was inhibited, which correlated with decreased signaling. Furthermore, inhibition of Src kinases with 1-tert-Butyl-3-(4-chlorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine also inhibited response to CpG deoxyribonucleic acid. Toll-like receptor-9 protein stability was also sensitive to autophagy, the cellular stress response pathway, and infection by a deoxyribonucleic acid virus. Whereas autophagy induced by rapamycin or low serum levels caused a preferential loss of the mature p80 proteolytic cleavage product, infection with herpes simplex virus-1 and induction of cell stress with tunicamycin caused preferential loss of full-length Toll-like receptor-9, which is localized to the endoplasmic reticulum. Our data reveal new information about the stability and signaling of Toll-like receptor-9 and suggest that immune evasion mechanisms may involve targeted loss of innate sensing receptors.

Keywords: CpG DNA; autophagy; inflammation; innate immunity.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Endoplasmic Reticulum Stress*
  • Mice
  • Mice, Knockout
  • Phosphorylation
  • Protein Processing, Post-Translational*
  • Protein Stability
  • Proteolysis
  • Signal Transduction
  • Toll-Like Receptor 9 / chemistry*
  • Toll-Like Receptor 9 / physiology*
  • Tyrosine / metabolism*


  • Tlr9 protein, mouse
  • Toll-Like Receptor 9
  • Tyrosine