Acclimation to diverse environmental stresses caused by a suppression of cytosolic ascorbate peroxidase in tobacco BY-2 cells
Acclimation to diverse environmental stresses caused by a suppression of cytosolic ascorbate peroxidase in tobacco BY-2 cells
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DOI：
10.1093/pcp/pci135
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发表时间：
2005-08-01
影响因子：
4.9
通讯作者：
Shigeoka, S
中科院分区：
文献类型：
--
作者：
Ishikawa, T;Morimoto, Y;Shigeoka, S
正常代谢过程中产生的活性氧受到各种抗氧化机制的严格控制。AOS是调节包括环境应激反应在内的许多生理过程的重要信号分子。在这项工作中，我们研究了在花椰菜花叶病毒（CaMV） 35S启动子下，拟南芥APX1 cDNA的表达在两个转化的by -2细胞系cAPX-S2和cAPX-S3的共同抑制下，降低了烟草by -2细胞的胞质抗坏血酸过氧化物酶（APX）活性的影响。与未转化的细胞相比，cAPX-S2和cAPX-S3的胞质APX活性分别降低了50%和75%。化学荧光分析表明，两种apx抑制细胞系的AOS水平明显高于野生型细胞。然而，其他各种抗氧化酶的活性水平没有实质性差异。有趣的是，apx抑制的细胞对环境胁迫（如高温和盐度）表现出不同的反应和耐受性。抑制减法杂交发现，在cAPX-S3细胞中，一些热胁迫和盐胁迫诱导基因被上调。组成型诱导基因包括HSP70、dnaj样蛋白和紫色酸性磷酸酶。凝胶内激酶分析表明，在apx抑制的细胞中，约46 kDa的丝裂原活化蛋白激酶（MAP）激酶主要活跃，烟叶蛋白激酶1 （NPK1）和核苷二磷酸激酶2 （NDPK2）的转录水平均上调。这些数据表明，MAP激酶级联反应可能是由胞质内APX活性降低引起的细胞氧化还原状态稳态的微妙失衡所激活的。
The active oxygen species (AOS) that arise from normal metabolic processes are kept under tight control by various antioxidant mechanisms. AOS are important signal molecules that regulate many physiological processes, including environmental stress responses. In this work, we have investigated the effect of lowered cytosolic ascorbate peroxidase (APX) activity in transgenic tobacco BY-2 cells, using two transformed BY-2 cell lines, cAPX-S2 and cAPX-S3, resulting from co-suppression by expression of Arabidopsis APX1 cDNA under the cauliflower mosaic virus (CaMV) 35S promoter. cAPX-S2 and cAPX-S3 possessed 50 and 75% lower cytosolic APX activity, respectively, compared with that in the untransformed cells. Chemical fluorescence analysis indicated that the AOS levels were markedly higher in the two APX-suppressed cell lines than in the wild-type cells. However, there were no substantial differences in the activity levels of the various other antioxidant enzymes. Interestingly, the APX-suppressed cells showed different responses and tolerances to environmental stresses, such as heat and salinity. Suppression subtractive hybridization revealed that several heat- and salt stress-inducible genes were up-regulated in cAPX-S3 cells. HSP70, DnaJ-like protein and purple acid phosphatase were among the constitutively induced genes. An in-gel kinase assay suggested that a mitogen-activated protein (MAP) kinase of approximately 46 kDa was predominantly active in the APX-suppressed cells, and transcript levels of both nicotiana protein kinase 1 (NPK1) and nucleoside diphosphate kinase 2 (NDPK2) were up-regulated. These data suggest the possibility that MAP kinase cascades are activated by subtle imbalances in the homeostasis of the cellular redox status caused by lowered cytosolic APX activity.