TLR5-Derived, TIR-Interacting Decoy Peptides to Inhibit TLR Signaling

Area of Science:

• Immunology
• Molecular Biology
• Drug Discovery

Background:

• Toll-like receptor 5 (TLR5) is crucial for immune responses against bacterial pathogens by recognizing flagellin.
• TLR signaling involves TIR domain dimerization and adapter recruitment, a key step for initiating intracellular cascades.
• Targeting TLR signaling pathways offers therapeutic potential for modulating immune responses.

Purpose of the Study:

• To identify and characterize cell-permeable decoy peptides that inhibit TLR5 signaling.
• To investigate the inhibitory potential of identified peptides against other Toll-like receptors (TLRs).
• To explore the evolutionary conservation and binding specificities of these inhibitory peptides.

Main Methods:

• Screening of decoy peptides derived from the TLR5 TIR domain in a HEK-Blue-mTLR5 reporter cell line.
• Assessment of peptide inhibition on TLR-induced cytokine expression in cell-based assays and in vivo mouse models.
• Time-resolved fluorescence spectroscopy used to determine peptide binding specificity to TLR TIR domains and adapter proteins.

Main Results:

• The peptide 5R667, derived from TLR5 TIR helix C″, demonstrated potent inhibition of TLR5 signaling.
• 5R667 exhibited broad inhibitory activity against TLR4, TLR2, and TLR9 signaling, and suppressed LPS-induced systemic cytokine induction in mice.
• 5R667 and TR667 (a homologous peptide from TIRAP) showed multispecific binding to TIR domains of TLR5, MyD88, and TIRAP, targeting an evolutionarily conserved motif.

Conclusions:

• A conserved adapter recruitment motif within TLR5 TIR domains can be targeted by cell-permeable decoy peptides.
• 5R667 and similar peptides represent effective inhibitors of TLR5 and other TLRs, acting as potential pan-specific TLR inhibitors.
• These findings offer a novel strategy for developing therapeutics to modulate inflammatory and immune responses.