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Now showing items 1 - 3 of 3

  • Identification and characterization of the TLR18 gene in grass carp (Ctenopharyngodon idella)

    Huang, Wen-Ji   Shen, Yubang   Xu, Xiao-Yan   Hu, Mo-Yan   Li, Jia-le  

    Toll-like receptors (TLRs) play a critical role in the innate immune system. Although TLR18 is an important member of this family of receptors in fish, the role of the tlr18 gene in responses to pathogen infection is still unclear. In this study, we identified and characterized the grass carp tlr18 gene (gctlr18) to further clarify the function of TLR18 in teleost fish. Gctlr18 spans over 3600 bp and encodes a polypeptide of 852 amino acids. Analysis of the deduced amino acid sequence showed that gctlr18 encodes structures typical of the TLR family, including a signal peptide, seven leucine-rich repeats (LRRs), a transmembrane region, and a (Toll-interleukin-1 receptor) TIR domain. Quantitative RT-PCR analysis showed that gctlr18 was constitutively expressed in all investigated tissues, with abundant expression in spleen, gill, heart, intestine, kidney and fin and low expression in skin, liver and brain. Following grass carp reovirus-challenge and Aeromonas hydrophila inoculation, gctlr18 transcripts were upregulated significantly in immune-relevant tissues. Stimulation of Ctenopharyngodon idella kidney (CIK) cells with purified flagellin from Salmo typhimurium, lipopolysaccharide and polyinosinic-polycytidylic acid stimulation in vitro resulted in significantly increased gctlr18 expression, reaching a peak followed by restoration of normal levels. Overexpression of gctlr18 reduced A. hydrophila invasion by 83.4%. In CIK cells, gctlr18 induced the expression of proinflammatory cytokines, including il-8, inf-1 and trif-alpha. Our results indicate that gctlr18 plays a key role in innate immune responses in teleost fish. (C) 2015 Elsevier Ltd. All rights reserved.
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  • Identification,characterization and immunological analysis of Ras related C3 botulinum toxin substrate 1 (Rac1) from grass carp Ctenopharyngodon idella

    Hu, Mo-Yan   Shen, Yu-Bang   Xu, Xiao-Yan   Yu, Hong-Yan   Zhang, Meng   Dang, Yun-Fei   Li, Jia-Le  

    Rac1, a Rho GTPase, serves critical immunological functions in mammals. Here, a Rac1 homolog (gcRac1) was identified in grass carp (Ctenopharyngodon idella). The full-length 2023-base pair gcRac1 cDNA contained a 579-bp open reading frame encoding a 192-residue protein, including a conserved RHO domain and nuclear localization signal. The gcRac1 protein shares high identity with other Rac1 counterparts and phylogenetically clustered with Danio redo Rac1. The gcRac1 transcript showed wide tissue distribution and was inducible by Aeromonas hydrophila in vivo and in vitro; its expression also fluctuated with LPS or flagellin stimulation in vitro. With gcRac1 over-expression, gcPAK1, gcIL1-beta, gcTNF-alpha and gcIFN were basically up-regulated by A. hydrophila and bacterial PAMPs induction, while gcRac1 knockdown decreased these transcripts after A. hydrophila challenge. Over-expression of gcRac1 reduced, while its suppression facilitated, bacterial invasion. Moreover, gcRac1 could activate NF-kappa B signaling. These findings implicate the vital role of gcRac1 in grass carp innate immunity. (C) 2015 Elsevier Ltd. All rights reserved.
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  • Identification, characterization and immunological analysis of Ras related C3 botulinum toxin substrate 1 (Rac1) from grass carp Ctenopharyngodon idella

    Hu, Mo-Yan   Shen, Yu-Bang   Xu, Xiao-Yan   Yu, Hong-Yan   Zhang, Meng   Dang, Yun-Fei   Li, Jia-Le  

    Highlights • A Rac1 gene was identified from grass carp Ctenopharyngodon idella. • Over-expression of gcRac1 induces IL1-β, TNF-α and IFN during pathogen stimulation. • Knocking down gcRac1 inhibits PAK1, IL1-β, TNF-α, IFN post Aeromonas hydrophila infection. • gcRac1 can induce NF-κB activity. • gcRac1 induces an effective immune response against A. hydrophila invasion in vitro. Abstract Rac1, a Rho GTPase, serves critical immunological functions in mammals. Here, a Rac1 homolog (gcRac1) was identified in grass carp (Ctenopharyngodon idella). The full-length 2023-base pair gcRac1 cDNA contained a 579-bp open reading frame encoding a 192-residue protein, including a conserved RHO domain and nuclear localization signal. The gcRac1 protein shares high identity with other Rac1 counterparts and phylogenetically clustered with Danio rerio Rac1. The gcRac1 transcript showed wide tissue distribution and was inducible by Aeromonas hydrophila in vivo and in vitro; its expression also fluctuated with LPS or flagellin stimulation in vitro. With gcRac1 over-expression, gcPAK1, gcIL1-β, gcTNF-α and gcIFN were basically up-regulated by A. hydrophila and bacterial PAMPs induction, while gcRac1 knockdown decreased these transcripts after A. hydrophila challenge. Over-expression of gcRac1 reduced, while its suppression facilitated, bacterial invasion. Moreover, gcRac1 could activate NF-κB signaling. These findings implicate the vital role of gcRac1 in grass carp innate immunity.
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