Department of Genome Biology, Institute of Molecular Biology and Biotechnology, Faculty of Biology, Adam Mickiewicz University, Umultowska 89, 61-614 Poznań, Poland.
Molecular Biology Techniques Laboratory, Faculty of Biology, Adam Mickiewicz University, Umultowska 89, 61-614 Poznań, Poland.
J Plant Physiol. 2018 Apr;223:84-95. doi: 10.1016/j.jplph.2018.01.011. Epub 2018 Mar 7.
In this report, the members of PP2C, SnRK2a and Rboh oxidase families from Arabidopsis and poplar were studied in silico, and the expression profiles of the some of them were specified in Populus tremula buds and adult leaves. In poplars, the counterparts of ABI1- and ABI2-like protein phosphatases are lacking, but poplar genomes encode three HAB-like proteins denoted in this work as HAB1, HAB3a and HAB3b, and the counterparts of the two latter ones are absent in Arabidopsis. Nonetheless, they may be present in other species. In poplars, SnRK2 subclass III includes two SnRK2.6-like protein kinases denoted by us as SnRK2.6a and SnRK2.6b, and only one SnRK2.2 corresponding to SnRK2.2 and SnRK2.3 ones from Arabidopsis. In contrast to Arabidopsis, the poplar Rboh family involves two RbohD- and RbohF-like proteins denoted here as RbohD1 and RbohD2, and RbohF1 and RbohF2, respectively. The expressions of genes encoding the above components of the ABA route were studied in Populus tremula dehydrated buds and adult leaves not subjected to stress but exposed to natural daylight or to darkness, and to inhibition of ethylene biosynthesis or signaling route by cobalt or silver ions, respectively. In leaves, the light conditions seemed to be the most pronounced factor, from among the studied stimuli, controlling the expression Ptre-HAB3a, Ptre-HAB1, Ptre-SnRK2.6a and Ptre-RbohF2 genes, their expression was upregulated in darkness. This observation implies that these genes may be important for dark-induced stomatal closure regulation. Ethylene negatively affected the expression of three studied Rboh genes and Ptre-HAB1one but only at daylight, whereas its positive effect on the of Ptre-HAB3a was shown in the dark exposed leaves. In buds, three studied Rboh genes took part in the early response to dehydration, however their participation involved the visibly highest level of the Ptre-RbohD1 transcripts, followed by Ptre-RbohF2 and the lowest one of Ptre-RbohF1. Nonetheless, the further stress-induced superoxide anion generation seemed to depend on the enhanced expression of the Ptre-RbohD1 and Ptre-RbohF2 genes only, still with a significantly higher level of the Ptre-RbohD1 one. Ptre-RbohD2 transcripts were found neither in leaves nor in buds. The expression of the other genes discussed in the present work was either slightly upregulated at moderate stress or did not significantly change in response to dehydration. The protein kinase activity of overexpressed Ptre-SnRK2.6a and Ptre-SnRK2.6b was confirmed in in vitro protein kinase assay and compared to that of SnRK2.6/OST1 one from Arabidopsis.
在本报告中，我们对拟南芥和杨树中的 PP2C、SnRK2a 和 Rboh 氧化酶家族成员进行了计算机分析，并在欧洲山杨芽和成年叶片中确定了其中一些成员的表达谱。在杨树中，ABI1-和 ABI2 样蛋白磷酸酶的对应物缺失，但杨树基因组编码了三个 HAB 样蛋白，我们在本工作中分别表示为 HAB1、HAB3a 和 HAB3b，而后两者的对应物在拟南芥中不存在。然而，它们可能存在于其他物种中。在杨树中，SnRK2 亚类 III 包括两个 SnRK2.6 样蛋白激酶，我们分别表示为 SnRK2.6a 和 SnRK2.6b，并且只有一个 SnRK2.2 对应于拟南芥中的 SnRK2.2 和 SnRK2.3。与拟南芥不同的是，杨树的 Rboh 家族涉及两个 RbohD-和 RbohF 样蛋白，我们分别表示为 RbohD1 和 RbohD2，以及 RbohF1 和 RbohF2。研究了编码 ABA 途径上述成分的基因在欧洲山杨脱水芽和未受胁迫但暴露于自然光或黑暗以及分别用钴或银离子抑制乙烯生物合成或信号转导途径的成年叶片中的表达。在叶片中，光照条件似乎是最显著的因素，在研究的刺激因素中，控制 Ptre-HAB3a、Ptre-HAB1、Ptre-SnRK2.6a 和 Ptre-RbohF2 基因的表达，它们的表达在黑暗中被上调。这一观察结果表明，这些基因可能对黑暗诱导的气孔关闭调节很重要。乙烯负调控三个研究的 Rboh 基因和 Ptre-HAB1 的表达，但仅在光照下，而其对 Ptre-HAB3a 的正影响在黑暗暴露的叶片中表现出来。在芽中，三个研究的 Rboh 基因参与了对脱水的早期反应，但它们的参与涉及 Ptre-RbohD1 转录本的最高水平，其次是 Ptre-RbohF2，最低水平是 Ptre-RbohF1。然而，进一步的应激诱导的超氧阴离子生成似乎仅依赖于 Ptre-RbohD1 和 Ptre-RbohF2 基因的增强表达，仍然具有明显更高水平的 Ptre-RbohD1。在叶片和芽中均未发现 Ptre-RbohD2 转录本。本研究中讨论的其他基因的表达要么在中度胁迫下略有上调，要么在响应脱水时没有明显变化。在体外蛋白激酶测定中证实了过表达的 Ptre-SnRK2.6a 和 Ptre-SnRK2.6b 的蛋白激酶活性，并与拟南芥中的 SnRK2.6/OST1 进行了比较。
