College of Life Science/Tianjin Key Laboratory of Cyto-genetical and Molecular Regulation, Tianjin Normal University, Tianjin, 300387, China.
Cell Stress Chaperones. 2012 Jul;17(4):445-55. doi: 10.1007/s12192-012-0322-x. Epub 2012 Jan 18.
Thioredoxin (Trx) is a small redox protein existing ubiquitously in all living organisms and plays an important role in multiple cellular processes, including transcriptional regulation and immune response. To date very few studies have been carried out to examine the function of piscine Trx. In this study, we identified and analyzed the function of a Trx homologue, CsTrx1, from half-smooth tongue sole (Cynoglossus semilaevis). The deduced amino acid sequence of CsTrx1 is composed of 107 residues and shares 54.1-60.8% overall identities with the Trx of other teleosts. CsTrx1 contains the highly conserved CXXC motif, which in mammals is known to be the active site, in the form of CQPC. Expression of CsTrx1 as determined by quantitative real-time reverse transcriptase PCR was highest in liver and upregulated in time-dependent manners by bacterial infection and by exposure to iron, copper, and hydrogen peroxide. Purified recombinant CsTrx1 (rCsTrx1) exhibited insulin disulfide reductase activity and antioxidant activity, both which, however, were lost when the two cysteine residues in the CQPC motif were mutated to serine. Further analysis showed that rCsTrx1 was able to stimulate the proliferation of head kidney leukocytes, upregulate the expression of immune relevant genes, and enhance the resistance of leukocytes against bacterial infection. Taken together, these results indicate that CsTrx1 is a biologically active reductase and an antioxidant that requires the CXXC motif for activity and that CsTrx1 possesses cytokine-like immunoregulatory property. These results suggest a role for CsTrx1 in protecting cells against oxidative stress caused by oxidant exposure and pathogen infection.
硫氧还蛋白（Trx）是一种广泛存在于所有生物中的小的氧化还原蛋白，在多种细胞过程中发挥重要作用，包括转录调控和免疫反应。迄今为止，很少有研究检查鱼类 Trx 的功能。在这项研究中，我们从半滑舌鳎（Cynoglossus semilaevis）中鉴定并分析了 Trx 同源物 CsTrx1 的功能。CsTrx1 的推导氨基酸序列由 107 个残基组成，与其他硬骨鱼的 Trx 整体具有 54.1-60.8%的同一性。CsTrx1 包含高度保守的 CXXC 基序，在哺乳动物中已知该基序是活性位点，形式为 CQPC。通过定量实时逆转录 PCR 确定的 CsTrx1 表达在肝脏中最高，并以时间依赖性方式被细菌感染和暴露于铁、铜和过氧化氢上调。纯化的重组 CsTrx1（rCsTrx1）表现出胰岛素二硫键还原酶活性和抗氧化活性，但当 CQPC 基序中的两个半胱氨酸残基突变为丝氨酸时，这两种活性都丧失。进一步分析表明，rCsTrx1 能够刺激头肾白细胞的增殖，上调免疫相关基因的表达，并增强白细胞对细菌感染的抵抗力。总之，这些结果表明 CsTrx1 是一种具有生物活性的还原酶和抗氧化剂，需要 CXXC 基序才能发挥活性，并且 CsTrx1 具有细胞因子样免疫调节特性。这些结果表明 CsTrx1 在保护细胞免受氧化剂暴露和病原体感染引起的氧化应激方面发挥作用。
