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Toll-like receptors (TLRs) play a critical role in the innate immune response of fish. In this study, we isolated the cDNA sequence of Nile tilapia TLR1 (OnTLR1). The deduced OnTLR1 protein contains a signal peptide, 7 leucine-rich repeats (LRRs), a C-terminal LRR (LRR-CT), a transmembrane region and a highly conserved TIR domain. In healthy Nile tilapia, the OnTLR1 transcript was broadly expressed in all examined tissues, with the highest expression levels in the spleen. After infection with Streptococcus agalactiae, the OnTLR1 transcripts were upregulated in the gill and kidney. After stimulation with polyinosinic-polycytidylic acid (poly(I:C)), the expression levels of OnTLR1 were significantly downregulated in the intestine, whereas OnTLR1 transcripts were significantly upregulated in the kidney. After challenge with lipopolysaccharide (LPS), the expression levels of OnTLR1 were significantly upregulated in the spleen and kidney. The subcellular localization showed that OnTLR1 was expressed in the cytoplasm. TLR1 significantly increased MyD88-dependent NF-κB activity. However, the results of a pull-down assay showed that OnTLR1 did not interact with MyD88 or TIRAP. Binding assays revealed the specificity of OnTLR1 for pathogen-associated molecular patterns (PAMPs) and bacteria that included S. agalactiae, Aeromonas hydrophila and poly(I:C) and LPS. Taken together, these findings suggest that OnTLR1, as a pattern recognition receptor (PRR), might play an important role in the immune response to pathogen invasion., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Toll-like receptors (TLRs) are important pattern recognition receptors (PRRs) of teleost innate immune system. However, information about TLRs is absent in Dabry's sturgeon (Acipenser dabryanus), one of the most primitive actinopterygii species. In the present study, the full lengths of adaTLR1 and adaTLR4 were cloned from Dabry's sturgeon using RT-PCR and RACE-PCR. The obtained adaTLR1 was 2957 bp in length, encoding a putative protein of 767 amino acids and adaTLR4 cDNA was 2902 bp in length, encoding a putative protein of 830 aa. Both adaTLR1 and adaTLR4 possessed several typical TLRs motifs, including signal peptides, leucine-rich repeat (LRR) motifs, a transmembrane domain and a TIR motifs. In addition, adaTLR4 contained three conserved boxes in its TIR motif, involving in TLRs signal transduction. A proline, important for LPS recognition of mammalian TLR4, was also found in adaTLR4. Physicochemical features of adaTLR1 and adaTRL4 were also analyzed. Quantitative realtime PCR showed that both transcripts were ubiquitously expressed in all 11 normal tissues selected, but they exhibited different expression patterns, with adaTLR1 highly expression in heart and adaTLR4 highly in skin. Further, adaTLR1 and adaTLR4 were up-regulated in the primary head-kidney leucocytes following LPS and polyI:C stimulation, indicating that both genes involved in the sturgeon immune response to LPS and polyI:C. To our best knowledge, this was the first report of these genes in sturgeon and these results provided the basis for further elucidating the ligand specificity and signaling pathway of fish TLRs., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Toll-like receptors (TLRs) are a group of pattern-recognition receptors which play vital roles in ligand recognition and activation of the innate immune response. As an important member of TLRs family, TLR1 is mainly responsible for PAMPs from bacteria and play a pivotal role in sensing microbial products. Recent studies revealed that TLR1 could perceive LPS stimulation and transfer signals to activate the NF-κB pathway, whereas ligands and signaling pathway of TLR1 are still unclear in fish. Growing evidence has shown that miRNAs (microRNAs) play as negative regulators in controlling the diverse of biophysical and biochemical processes at the post-transcriptional level. In this study, we used a combination of bioinformatics and experimental techniques to exhibit that both miR-8159-5p and miR-217-5p were the direct negative regulators of TLR1 in miiuy croaker. Furthermore, dual-luciferase reporter assays showed that combining miR-8159-5p and miR-217-5p exhibited a greater negative regulatory effect on TLR1 than only miR-8159-5p or miR-217-5p. Additionally, we also demonstrated that the expression of both the two miRNAs could be up-regulated by LPS stimulation in either LPS-stimulation spleen tissue or LPS-treated cultured macrophage, which indicating that miR-8159-5p and miR-217-5p could be induced by LPS and may be as the negative regulators of TLR1 involved in the immune response to LPS stimulation. These results would enhance our understanding of the miRNA regulation in fish TLR signaling pathways., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

MicroRNAs (miRNAs) are highly conserved, small non-coding RNAs which post-transcriptionally regulate various biological processes by repressing mRNA translation or degradating mRNA. It has been demonstrated that miRNAs play crucial roles in regulating the immune system. In this study, we explored the potential roles of miR-200a-3p in regulating TLR signaling pathway in miiuy croaker. Bioinformatics analysis showed that miiuy croaker TLR1 (mmiTLR1) was a putative target of miR-200a-3p. Negative expression profiles in spleen of Vibrio anguillarum challenged miiuy croaker and in lipopolysaccharide (LPS) stimulated miiuy croaker leukocytes further validated the prediction. Luciferase reporter assays showed that the dual-luciferase reporter fused to the 3'UTR of wild type mmiTLR1 cotransfected with miR-200a-3p mimics exhibited a reduction in luciferase activity compared with the controls. All of the present data provide direct evidence that miR-200a-3p is involved in TLR1 expression modulation in miiuy croaker, which will offer a basis for better understanding of miRNA regulation in fish TLR signaling pathways., (Copyright © 2016 Elsevier Ltd. All rights reserved.)