Toll-like Receptor 8 Stability Is Regulated by Ring Finger 216 in Response to Circulating MicroRNAs.

John Evankovich, Travis Lear, Courtney Baldwin, Yanwen Chen, Virginia White, John Villandre, James Londino, Yuan Liu, Bryan McVerry, Georgios D Kitsios, Rama K Mallampalli, Bill B Chen

American journal of respiratory cell and molecular biology 2020 Feb

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TLR8 (Toll-like receptor 8) is an intracellular pattern recognition receptor that senses RNA in endosomes to initiate innate immune signaling through NF-κB, and mechanisms regulating TLR8 protein abundance are not completely understood. Protein degradation is a cellular process controlling protein concentrations, accomplished largely through ubiquitin transfer directed by E3 ligase proteins to substrates. In the present study, we show that TLR8 has a short half-life in THP-1 monocytes (∼1 h) and that TLR8 is ubiquitinated and degraded in the proteasome. Treatment with the TLR8 agonist R848 causes rapid depletion of TLR8 concentrations at early time points, an effect blocked by proteasomal inhibition. We show a novel role for RNF216 (ring finger protein 216), an E3 ligase that targets TLR8 for ubiquitination and degradation. RNF216 overexpression reduces TLR8 concentrations, whereas RNF216 knockdown stabilizes TLR8. We describe a potential role for TLR8 activation by circulating RNA ligands in humans with acute respiratory distress syndrome (ARDS): Plasma and extracted RNA fractions from subjects with ARDS activated TLR8 in vitro. MicroRNA (miRNA) expression profiling revealed several circulating miRNAs from subjects with ARDS. miRNA mimics promoted TLR8 proteasomal degradation in THP-1 cells. These data show that TLR8 proteasomal disposal through RNF216 in response to RNA ligands regulates TLR8 cellular concentrations and may have implications for innate immune signaling. In addition, TLR8 activation by circulating RNA ligands may be a previously underrecognized stimulus contributing to excessive innate immune signaling characteristic of ARDS.

Citation

John Evankovich, Travis Lear, Courtney Baldwin, Yanwen Chen, Virginia White, John Villandre, James Londino, Yuan Liu, Bryan McVerry, Georgios D Kitsios, Rama K Mallampalli, Bill B Chen. Toll-like Receptor 8 Stability Is Regulated by Ring Finger 216 in Response to Circulating MicroRNAs. American journal of respiratory cell and molecular biology. 2020 Feb;62(2):157-167