From an EST fragment of the rainbow trout that was predicted to contain leucine-rich repeats (LRR), we cloned the whole cDNA and identified a soluble form of TLR5 ortholog (rtTLR5S), which does not exist in the mouse and human. rtTLR5S was about 38% homologous to the extracellular domains of human (hu) and mouse (mo)TLR5, while rtTLR5S showed <25% homologous to those of other human or mouse TLRs. A chimera constructed of rtTLR5S and the intra-cellular TIR of huTLR5 expressed on HeLa cells signaled the presence of flagellin A and C from V. anguillarum, resulting in NF-kappaB activation. The mRNA of rtTLR5S was predominantly detected in the liver. The hepatoma cell line of the rainbow trout RTH149 that responded to flagellin, allowed to up-regulate rtTLR5S in response to V. anguillarum or its purified flagellin within 8 h. rtTLR5S, when co-expressed with membrane huTLR5 in HeLa cells, augmented huTLR5-mediated NF-kappaB activation in response to flagellin. These results, together with the genome information of the pufferfish Fugu (Fugu rubripes), suggest that in fish the soluble TLR5 is an acute-phase protein sensing bacterial infection via recognition of a variety of bacterial flagellins to augment NF-kappaB activation, and may be important for fish to survive from bacterial infection in the water.
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