Adopting orphan receptors: zebrafish Tlr4 homologs mediate responses to group IX/X transition metals

Aaron P D Fox ¹, Tracy S Lee ², Sakina N Mithaiwala ¹

⁰Department of Biological Sciences, Faculty of Science, University of Alberta, Edmonton, AB, Canada.

Frontiers in Immunology +

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December 12, 2025

View abstract on PubMed

Summary

Toll-like receptor 4 (TLR4) homologs in zebrafish are essential for mediating toxic responses to transition metals like nickel, cobalt, and platinum. This research clarifies the function of TLR4 in zebrafish and suggests a conserved role in detecting metals across species.

Area Of Science

Immunology
Toxicology
Developmental Biology

Background

Toll-like receptor 4 (TLR4) is a key immune receptor recognizing lipopolysaccharide (LPS), crucial for infection defense.
The precise function of TLR4 homologs in zebrafish and other fish species remains unclear, with conflicting data regarding LPS effects.
Previous studies suggest zebrafish Tlr4 may mediate ototoxicity from platinum-based chemotherapy, hinting at broader ligand detection capabilities.

Purpose Of The Study

To investigate the hypothesis that zebrafish Tlr4 detects group IX/X transition metals.
To explore conserved roles of Tlr4 with human TLR4 in mediating responses to metals, specifically nickel.
To resolve the function of TLR4 homologs in the zebrafish model for disease research.

Main Methods

Utilized the larval zebrafish lateral line model to assess ototoxicity.
Administered varying concentrations of nickel (Ni), cobalt (Co), and platinum (Pt) to evaluate dose-dependent effects.
Employed CRISPR technology for Tlr4 knockdown to determine its necessity in metal-induced toxicity.
Expressed zebrafish Tlr4 in a human cell line to assess its sufficiency in mediating inflammatory responses to metals.

Main Results

Demonstrated dose-dependent ototoxicity of Ni, Co, and Pt at (sub-)micromolar levels in zebrafish larvae.
Showed that Tlr4 knockdown significantly reduced the ototoxic effects of these metals by approximately 50%, highlighting TLR4's requirement.
Confirmed that zebrafish Tlr4 is sufficient to mediate inflammatory responses to transition metals when expressed heterologously in human cells.

Conclusions

Zebrafish TLR4 homologs are both necessary and sufficient for mediating responses to transition metals.
These findings support the conserved function of TLR4 homologs in detecting transition metals, potentially dating back to the last common ancestor of fish and mammals.
This study clarifies the role of TLR4 in the zebrafish model, particularly concerning metal-induced toxicities.

Keywords:

MD-2 ecotoxicology inflammation innate immunity toll-like receptor 4