Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Bunkyo-ku, Japan.
Biochem Biophys Res Commun. 2011 Aug 12;411(4):708-13. doi: 10.1016/j.bbrc.2011.07.008. Epub 2011 Jul 12.
Nitrate is a major nitrogen source for land plants but also acts as a signaling molecule that regulates gene expression, metabolism and plant growth. The genes for nitrate reductase (NR) and nitrite reductase (NIR), which are enzymes in the nitrate assimilation pathway, are typical nitrate-inducible genes. We previously identified the first authentic nitrate-responsive cis-element (NRE) for nitrate-inducible transcription by the analysis of the NIR gene promoter from Arabidopsis. Here we further characterize NRE-mediated regulation using transgenic Arabidopsis plants. First, NRE-mediated regulation is shown to be a primary response to nitrate that is caused by pre-existing components, because the NRE-mediated nitrate-inducible expression of the GUS reporter gene is unaffected by treatment with the protein synthesis inhibitor cycloheximide. Second, we show that NRE-like sequences are present in various dicotyledonous and monocotyledonous NIR gene promoters at similar positions and that they also drive nitrate-inducible expression in Arabidopsis. This suggests that NRE-mediated regulation might be conserved in higher plants. Finally, the NRE-mediated expression of the GUS reporter gene is shown to produce diurnal expression in transgenic Arabidopsis plants. This expression peaked at the beginning of the day and decreased during the day which is very similar to the reported diurnal pattern of the nitrate content, suggesting a role of NRE-mediated regulation in controlling diurnal expression in response to oscillation of the nitrate content over a day/night cycle. These findings further clarify the roles of the NRE-mediated regulatory system in higher plants.
硝酸盐是陆地植物的主要氮源，但它也作为一种信号分子，调节基因表达、代谢和植物生长。硝酸盐还原酶（NR）和亚硝酸盐还原酶（NIR）的基因，它们是硝酸盐同化途径中的酶，是典型的硝酸盐诱导基因。我们之前通过分析拟南芥的 NIR 基因启动子，鉴定了第一个真正的硝酸盐响应顺式作用元件（NRE），用于硝酸盐诱导的转录。在这里，我们使用转基因拟南芥植物进一步研究了 NRE 介导的调控。首先，证明 NRE 介导的调控是对硝酸盐的主要反应，是由预先存在的成分引起的，因为 NRE 介导的 GUS 报告基因的硝酸盐诱导表达不受蛋白质合成抑制剂环己酰亚胺处理的影响。其次，我们表明，NRE 样序列存在于各种双子叶植物和单子叶植物 NIR 基因启动子中相似的位置，并且它们也在拟南芥中驱动硝酸盐诱导表达。这表明 NRE 介导的调控可能在高等植物中保守。最后，证明 NRE 介导的 GUS 报告基因的表达在转基因拟南芥植物中产生昼夜表达。这种表达在白天开始时达到峰值，并在白天下降，这与硝酸盐含量的报道昼夜模式非常相似，表明 NRE 介导的调控在控制昼夜表达以响应一天/夜周期中硝酸盐含量的波动方面发挥作用。这些发现进一步阐明了 NRE 介导的调控系统在高等植物中的作用。
