Studies on Induction and Functions of Thioredoxin Reductase in Stress Responses

硫氧还蛋白还原酶应激反应的诱导及功能研究

• 批准号: 13672345
• 负责人: HARA Shuntaro
• 金额: $ 2.24万
• 依托单位: Kitasato University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13672345/
• 关键词: Thiorecoxin reductase; Transcription factor; Redox; NF-κB; Stress response; Cadmium; レドックス制御; ストレス応答

Thioredoxin reductase (TrxR) is an antioxidant enzyme that has an ability to reduce thioredoxin (Trx). It is indicated that Trx plays an important role in stress responses, but the role of TrxR in the stress responses remains unclear. To date, three TrxR isozymes, TrxR1, TrxR2 and Trx3, have been identified. Among these isozymes, only TrxR1 is induced by various kinds of stresses. In this study, in order to reveal the role of TrxR in stress responses, the mechanism of induction and the functions of TrxR1 were studied as follows:(1) Regulation of stress-induced TrxR1 expression - Exposure of bovine arterial endothelial cells (BAEC) to TNFα, PMA+A23187, or Cd upregulated the expression of TrxR1 but neither TrxR2 nor TrxR3. First, role of the promoter region in the huma TrxR1 gene was evaluated by the transient transfection of luciferase reporter vectors into BAEC. Both PMA+A23187 and Cd elevated luciferase activity via the region between-646 and 46 bp. On the other hand, TNFα did not affect the luciferase activity, but the mRNA decay experiments using actinomycin D showed that TNFα stabilized TrxR1 mRNA. These results indicate that both transcriptional activation and posttranscriptional mRNA stabilization may contribute to the stress-induced expression of TrxR1, and that the contributive mechanisms may vary with kinds of stresses.(2) Role of TrxR1 in stress-responsive transcription factor-mediated gene expression - Reporter gene analysis revealed that the overexpression of TrxR1 enhanced AP-1- and NF-κB-dependent gene expression. The catalytic selenocysteine residue of TrxR1, which is essential for reducing Trx, was required for this activation, and aurothiomalate, an inhibitor of TrxR, suppressed this activation. These results suggest that TrxR1 may act as a positive regulator of stress-responsive transcription factors, AP-1 and NF-κB via its ability to reduce Trx.

硫氧还蛋白还原酶（TrxR）是一种具有还原硫氧还蛋白（Trx）能力的抗氧化酶。表明Trx在应激反应中起重要作用，但TrxR在应激反应中的作用尚不清楚。到目前为止，TrxR同工酶TrxR1、TrxR2和Trx3已经被鉴定出来。在这些同工酶中，只有TrxR1受到各种胁迫的诱导。为了揭示TrxR在应激反应中的作用，本研究对TrxR1的诱导机制和功能进行了如下研究：(1)应激诱导TrxR1的表达调控——牛动脉内皮细胞（BAEC）暴露于TNFα、PMA+A23187或Cd后，TrxR1的表达上调，但TrxR2和TrxR3均未上调。首先，通过将荧光素酶报告载体瞬时转染到BAEC中，评估了启动子区域在人类TrxR1基因中的作用。PMA+A23187和Cd均通过-646 ~ 46 bp区域提高荧光素酶活性。另一方面，TNFα不影响荧光素酶活性，但放线菌素D的mRNA衰减实验表明，TNFα稳定TrxR1 mRNA。这些结果表明，TrxR1的转录激活和转录后mRNA稳定都可能参与胁迫诱导的TrxR1的表达，其作用机制可能因胁迫的不同而不同。(2) TrxR1在应激应答转录因子介导的基因表达中的作用——报告基因分析显示，TrxR1过表达可增强AP-1和NF-κ b依赖性基因的表达。TrxR1的催化硒代半胱氨酸残基是还原Trx所必需的，而TrxR的抑制剂金硫硫酸盐抑制了这种激活。这些结果表明，TrxR1可能通过其减少Trx的能力，作为应激反应转录因子AP-1和NF-κB的正调节因子。