Here, we describe functional characterization of Arabidopsis thaliana MYB4 transcription factor, a member of R2R3-subfamily of MYB domain protein, in the regulation of Cd-stress tolerance in Arabidopsis. Transgenic Arabidopsis plants overexpressing MYB4 showed appreciable Cd tolerance than wild-type plants, while MYB4 loss of function mutant lines (atmyb4) showed increased sensitivity to Cd-stress. MYB4 overexpression lines showed strong activation of anti-oxidant defense components and increased Cd accumulation than wild-type and atmyb4 mutant lines under Cd-stress. MYB4 overexpression resulted in the coordinated activation of the expression of phytochelatin (PC) synthesis related genes and specifically enhanced the transcript abundance of phytochelatin synthase 1 (PCS1) and metallothionein 1C (MT1C) genes under Cd-stress. In contrast, atmyb4 mutant lines showed reduced Cd accumulation and compromised expression of PC-synthesis related genes. Electrophoretic gel mobility shift assays have demonstrated specific binding activity of recombinant AtMYB4 to the putative MYB4-binding motifs ACCAACCAA and GGTAGGT identified in the promoters of PCS1 and MT1C genes, respectively. Further analyses have revealed that MYB4 binds directly to PCS1 and MT1C promoters in vivo and positively regulates their transcriptional expression, suggesting that PCS1 and MT1C are the key targets of MYB4. Overall, our results have provided evidence that MYB4 regulates Cd-tolerance via the coordinated activity of improved anti-oxidant defense system and through the enhanced expression of PCS1 and MT1C under Cd-stress in Arabidopsis.

中文翻译：

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MYB4 转录因子是 MYB 域蛋白 R2R3 亚家族的成员，通过增强对氧化损伤的保护来调节镉耐受性并增加拟南芥中 PCS1 和 MT1C 的表达

在这里，我们描述了拟南芥 MYB4 转录因子（MYB 结构域蛋白的 R2R3 亚家族成员）在调节拟南芥 Cd 胁迫耐受性中的功能表征。过表达 MYB4 的转基因拟南芥植物比野生型植物表现出明显的 Cd 耐受性，而 MYB4 功能丧失突变系 (atmyb4) 显示对 Cd 胁迫的敏感性增加。在镉胁迫下，与野生型和 atmyb4 突变系相比，MYB4 过表达系显示出抗氧化防御成分的强烈激活和增加的镉积累。MYB4 过表达导致植物螯合素 (PC) 合成相关基因表达的协同激活，并在镉胁迫下特异性增强植物螯合素合酶 1 (PCS1) 和金属硫蛋白 1C (MT1C) 基因的转录丰度。相比之下，atmyb4 突变体系显示出降低的 Cd 积累和受损的 PC 合成相关基因的表达。电泳凝胶迁移率变化分析表明，重组 AtMYB4 对分别在 PCS1 和 MT1C 基因启动子中鉴定的推定 MYB4 结合基序 ACCAACCAA 和 GGTAGGT 具有特异性结合活性。进一步的分析表明，MYB4 在体内直接与 PCS1 和 MT1C 启动子结合并正调节它们的转录表达，表明 PCS1 和 MT1C 是 MYB4 的关键靶点。总体而言，我们的研究结果提供了证据，表明 MYB4 通过改进的抗氧化防御系统的协同活动以及通过在拟南芥中 Cd 胁迫下增强 PCS1 和 MT1C 的表达来调节 Cd 耐受性。