Phosphorylation of the Nicotiana benthamiana WRKY8 Transcription Factor by MAPK Functions in the Defense Response
Phosphorylation of the Nicotiana benthamiana WRKY8 Transcription Factor by MAPK Functions in the Defense Response
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DOI：
10.1105/tpc.110.081794
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发表时间：
2011-03-01
期刊：
影响因子：
11.6
通讯作者：
Yoshioka, Hirofumi
中科院分区：
文献类型：
--
作者：
Ishihama, Nobuaki;Yamada, Reiko;Yoshioka, Hirofumi
丝裂原活化蛋白激酶（MAPK）级联在植物先天免疫中起关键作用。然而，植物防御相关MAPKs的下游信号传导尚不清楚。本研究表明，本烟WRKY8转录因子是SIPK、NTF4和WIPK的生理底物。在体外实验中，这些MAPKs磷酸化了WRKY8 n端区域的簇状前导向丝氨酸残基（SP簇），这些簇状前导向丝氨酸残基在I组WRKY蛋白中是保守的。抗磷酸肽抗体表明WRKY8 SP簇丝氨酸残基在体内可被SIPK、NTF4和WIPK磷酸化。WRKY8与mapk的相互作用依赖于它的D结构域，这是一个mapk相互作用的基序，这种相互作用是WRKY8在植物中有效磷酸化所必需的。WRKY8的磷酸化增加了其与同源W-box序列的DNA结合活性。WRKY8的磷酸化模拟突变体表现出较高的转激活活性，其异位表达诱导了3-羟基-3-甲基戊二酰辅酶a还原酶2和nadp -苹果酸酶等防御相关基因的表达。相比之下，WRKY8的沉默降低了防御相关基因的表达，增加了对病原菌疫霉（Phytophthora infestans）和炭疽杆菌（Colletotrichum orbiculare）的易感性。因此，mapk介导的WRKY8磷酸化通过激活下游基因在防御反应中发挥重要作用。
Mitogen-activated protein kinase (MAPK) cascades have pivotal roles in plant innate immunity. However, downstream signaling of plant defense-related MAPKs is not well understood. Here, we provide evidence that the Nicotiana benthamiana WRKY8 transcription factor is a physiological substrate of SIPK, NTF4, and WIPK. Clustered Pro-directed Ser residues (SP cluster), which are conserved in group I WRKY proteins, in the N-terminal region of WRKY8 were phosphorylated by these MAPKs in vitro. Antiphosphopeptide antibodies indicated that Ser residues in the SP cluster of WRKY8 are phosphorylated by SIPK, NTF4, and WIPK in vivo. The interaction of WRKY8 with MAPKs depended on its D domain, which is a MAPK-interacting motif, and this interaction was required for effective phosphorylation of WRKY8 in plants. Phosphorylation of WRKY8 increased its DNA binding activity to the cognate W-box sequence. The phospho-mimicking mutant of WRKY8 showed higher transactivation activity, and its ectopic expression induced defense-related genes, such as 3-hydroxy-3-methylglutaryl CoA reductase 2 and NADP-malic enzyme. By contrast, silencing of WRKY8 decreased the expression of defense-related genes and increased disease susceptibility to the pathogens Phytophthora infestans and Colletotrichum orbiculare. Thus, MAPK-mediated phosphorylation of WRKY8 has an important role in the defense response through activation of downstream genes.