Yeh, D.W., Liu, Y.L., Lo, Y.C., Yuh, C.H., Yu, G.Y., Lo, J.F., Luo, Y., Xiang, R., and Chuang, T.H. (2013) Toll-like receptor 9 and 21 have different ligand recognition profiles and cooperatively mediate activity of CpG-oligodeoxynucleotides in zebrafish. Proceedings of the National Academy of Sciences of the United States of America. 110(51):20711-20716.
CpG-oligodeoxynucleotides (CpG-ODNs) are potent immune stimuli currently under investigation as antimicrobial agents for different
species. Toll-like receptor (TLR) 9 and TLR21 are the cellular receptors of CpG-ODN in mammals and chickens, respectively.
The avian genomes lack TLR9, whereas mammalian genomes lack TLR21. Although fish contain both of these genes, the biological functions of fish TLR9 and TLR21 have not been investigated previously.
In this study, we comparatively investigated zebrafish TLR9 (zebTLR9) and TLR21 (zebTLR21). The two TLRs have similar expression
profiles in zebrafish. They are expressed during early development stages and are preferentially expressed in innate immune
function-related organs in adult fish. Results from cell-based activation assays indicate that these two zebrafish TLRs are
functional, responding to CpG-ODN but not to other TLR ligands. zebTLR9 broadly recognized CpG-ODN with different CpG motifs,
but CpG-ODN with GACGTT or AACGTT had better activity to this TLR. In contrast, zebTLR21 responded preferentially to CpG-ODN
with GTCGTT motifs. The distinctive ligand recognition profiles of these two TLRs were determined by their ectodomains. Activation
of these two TLRs by CpG-ODN occurred inside the cells and was modulated by UNC93B1. The biological functions of these two
TLRs were further investigated. The CpG-ODNs that activate both zebTLR9 and zebTLR21 were more potent than others that activate
only zebTLR9 in the activation of cytokine productions and were more bactericidal in zebrafish. These results suggest that
zebTLR9 and zebTLR21 cooperatively mediate the antimicrobial activities of CpG-ODN. Overall, this study provides a molecular
basis for the activities of CpG-ODN in fish.