Identification of three MAPKKKs forming a linear signaling pathway leading to programmed cell death in Nicotiana benthamiana.
Identification of three MAPKKKs forming a linear signaling pathway leading to programmed cell death in Nicotiana benthamiana.
复制标题
DOI：
10.1186/1471-2229-12-103
复制
发表时间：
2012-07-08
影响因子：
5.3
通讯作者：
Namba S
中科院分区：
文献类型：
--
作者：
Hashimoto M;Komatsu K;Maejima K;Okano Y;Shiraishi T;Ishikawa K;Takinami Y;Yamaji Y;Namba S
丝裂原活化蛋白激酶（MAPK）级联是真核细胞中发现的一种进化上古老的信号转导机制。在植物中，MAPK级联与对各种非生物和生物胁迫（如植物病原体）的响应相关。MAPK级联通过顺序磷酸化起作用：MAPK激酶激酶（MAPKKKs）磷酸化MAPK激酶（MAPKKs），并且磷酸化的MAPKKs磷酸化MAPK。在这三种类型的激酶中，MAPKKK在植物基因组中表现出最大的分歧。尽管MAPKK和MAPKs的数量相对有限，但它们的巨大多样性被认为允许MAPKK调节植物中许多特定的信号传导途径。尽管包括本氏烟草（Nicotiana benthamiana）的MAPKKKα（NbMAPKKKα）在内的一些植物MAPKKK在植物防御反应中起着重要作用，但MAPKKK基因之间的功能关系尚不清楚。在这里，我们进行了比较功能分析MAPKKKs调查信号通路导致的防御反应。我们从N.本塞姆氏菌：NbMAPKKKβ、NbMAPKKKγ和NbMAPKKKε2。全长NbMAPKKKβ或NbMAPKKKγ或其激酶结构域在N.本塞姆氏叶诱导过敏反应（HR）样细胞死亡与过氧化氢的产生。这种活性依赖于过表达的MAPKKK的激酶活性。此外，病毒诱导的NbMAPKKKβ或NbMAPKKKγ表达的沉默显著抑制了病毒感染诱导的程序性细胞死亡（PCD）。此外，在对NbMAPKKKβ、NbMAPKKKγ和NbMAPKKKα之间的功能关系的上位性分析中，（先前显示参与植物防御反应），NbMAPKKKα的沉默抑制由NbMAPKKKβ激酶结构域或NbMAPKKKγ的过表达诱导的细胞死亡，但NbMAPKKKβ的沉默不能抑制由NbMAPKKKα或NbMAPKKKγ过表达诱导的细胞死亡。NbMAPKKKγ的沉默抑制由NbMAPKKKβ激酶结构域诱导的细胞死亡，但不抑制由NbMAPKKKα诱导的细胞死亡。这些结果表明，除了NbMAPKKKα外，NbMAPKKKβ和NbMAPKKKγ也是PCD的正调控因子。此外，这三种MAPKKK形成了导致PCD的线性信号通路;该通路从NbMAPKKKβ到NbMAPKKKγ再到NbMAPKKKα。
The mitogen-activated protein kinase (MAPK) cascade is an evolutionarily ancient mechanism of signal transduction found in eukaryotic cells. In plants, MAPK cascades are associated with responses to various abiotic and biotic stresses such as plant pathogens. MAPK cascades function through sequential phosphorylation: MAPK kinase kinases (MAPKKKs) phosphorylate MAPK kinases (MAPKKs), and phosphorylated MAPKKs phosphorylate MAPKs. Of these three types of kinase, the MAPKKKs exhibit the most divergence in the plant genome. Their great diversity is assumed to allow MAPKKKs to regulate many specific signaling pathways in plants despite the relatively limited number of MAPKKs and MAPKs. Although some plant MAPKKKs, including the MAPKKKα of Nicotiana benthamiana (NbMAPKKKα), are known to play crucial roles in plant defense responses, the functional relationship among MAPKKK genes is poorly understood. Here, we performed a comparative functional analysis of MAPKKKs to investigate the signaling pathway leading to the defense response. We cloned three novel MAPKKK genes from N. benthamiana: NbMAPKKKβ, NbMAPKKKγ, and NbMAPKKKε2. Transient overexpression of full-length NbMAPKKKβ or NbMAPKKKγ or their kinase domains in N. benthamiana leaves induced hypersensitive response (HR)-like cell death associated with hydrogen peroxide production. This activity was dependent on the kinase activity of the overexpressed MAPKKK. In addition, virus-induced silencing of NbMAPKKKβ or NbMAPKKKγ expression significantly suppressed the induction of programmed cell death (PCD) by viral infection. Furthermore, in epistasis analysis of the functional relationships among NbMAPKKKβ, NbMAPKKKγ, and NbMAPKKKα (previously shown to be involved in plant defense responses) conducted by combining transient overexpression analysis and virus-induced gene silencing, silencing of NbMAPKKKα suppressed cell death induced by the overexpression of the NbMAPKKKβ kinase domain or of NbMAPKKKγ, but silencing of NbMAPKKKβ failed to suppress cell death induced by the overexpression of NbMAPKKKα or NbMAPKKKγ. Silencing of NbMAPKKKγ suppressed cell death induced by the NbMAPKKKβ kinase domain but not that induced by NbMAPKKKα. These results demonstrate that in addition to NbMAPKKKα, NbMAPKKKβ and NbMAPKKKγ also function as positive regulators of PCD. Furthermore, these three MAPKKKs form a linear signaling pathway leading to PCD; this pathway proceeds from NbMAPKKKβ to NbMAPKKKγ to NbMAPKKKα.