Toll-like receptors (TLRs) are the basic signaling receptors of the innate immune
system. They are activated by molecules associated with pathogens or injured host
cells and tissue. TLR3 has been shown to respond to double stranded (ds) RNA, a replication
intermediary for many viruses. Here we present evidence that heterologous RNA released
from or associated with necrotic cells or generated by in vitro transcription also
stimulates TLR3 and induces immune activation. To assess RNA-mediated TLR3 activation,
human embryonic kidney 293 cells stably expressing TLR3 and containing a nuclear factor-κB-dependent
luciferase reporter were generated. Exposing these cells to in vitro transcribed RNA
resulted in a TLR3-dependent induction of luciferase activity and interleukin-8 secretion.
Treatment with in vitro transcribed mRNA activated nuclear factor-κB via TLR3 through
a process that was dose-dependent and involved tyrosine phosphorylation. Furthermore,
in vitro transcribed natural or 2′-fluoro-substituted mRNA induced the expression
of TLR3, interferon regulatory factor-1, tumor necrosis factor-α, and interleukin-1
receptor-associated kinase-M mRNA in human dendritic cells (DCs). DCs responded to
mRNA treatment by expressing activation markers, and this maturation was inhibited
by antagonistic TLR3-specific antibody. Endogenous RNA released from or associated
with necrotic cells also stimulated DCs, leading to interferon-α secretion, which
could be abolished by pretreatment of necrotic cells with RNase. These results demonstrate
that RNA, likely through secondary structure, is a potent host-derived activator of
TLR3. This finding has potential physiologic relevance because RNA escaping from damaged
tissue or contained within endocytosed cells could serve as an endogenous ligand for
TLR3 that induces or otherwise modulates immune responses.
