Disease Resistant Crops Research Unit, National Institute of Agrobiological Sciences, Ibaraki 305-8602, Japan.
BMC Plant Biol. 2013 Oct 4;13:150. doi: 10.1186/1471-2229-13-150.
The rice transcription factor WRKY45 plays a crucial role in salicylic acid (SA)/benzothiadiazole (BTH)-induced disease resistance. Its knockdown severely reduces BTH-induced resistance to the fungal pathogen Magnaporthe oryzae and the bacterial pathogen Xanthomonas oryzae pv. oryzae (Xoo). Conversely, overexpression of WRKY45 induces extremely strong resistance to both of these pathogens. To elucidate the molecular basis of WRKY45-dependent disease resistance, we analyzed WRKY45-regulated gene expression using rice transformants and a transient gene expression system.
We conducted a microarray analysis using WRKY45-knockdown (WRKY45-kd) rice plants, and identified WRKY45-dependent genes among the BTH-responsive genes. The BTH-responsiveness of 260 genes was dependent on WRKY45. Among these, 220 genes (85%), many of which encoded PR proteins and proteins associated with secondary metabolism, were upregulated by BTH. Only a small portion of these genes overlapped with those regulated by OsNPR1/NH1, supporting the idea that the rice SA pathway branches into WRKY45- regulated and OsNPR1/NH1-regulated subpathways. Dexamethazone-induced expression of myc-tagged WRKY45 in rice immediately upregulated transcription of endogenous WRKY45 and genes encoding the transcription factors WRKY62, OsNAC4, and HSF1, all of which have been reported to have defense-related functions. This was followed by upregulation of defense genes encoding PR proteins and secondary metabolic enzymes. Many of these genes were also induced after M. oryzae infection. Their temporal transcription patterns were consistent with those after dexamethazone-induced WRKY45 expression. In a transient expression system consisting of particle bombardment of rice coleoptiles, WRKY45 acted as an effector to trans-activate reporter genes in which the luciferase coding sequence was fused to upstream and intragenic sequences of WRKY62 and OsNAC4. Trans-activation of transcription occurred through a W-box-containing sequence upstream of OsNAC4 and mutations in the W-boxes abolished the trans-activation.
These data suggest a role of WRKY45 in BTH-induced disease resistance as a master regulator of the transcriptional cascade regulating defense responses in one of two branches in the rice SA pathway.
水稻转录因子 WRKY45 在水杨酸（SA）/苯并噻二唑（BTH）诱导的抗病性中发挥关键作用。其敲低严重降低了 BTH 诱导对真菌病原体稻瘟病菌和细菌病原体稻黄单胞菌 pv. （Xoo）的抗性。相反，WRKY45 的过表达诱导对这两种病原体的极强抗性。为了阐明 WRKY45 依赖性抗病性的分子基础，我们使用水稻转化体和瞬时基因表达系统分析了 WRKY45 调节的基因表达。
我们使用 WRKY45 敲低（WRKY45-kd）水稻植物进行了微阵列分析，并在 BTH 应答基因中鉴定了 WRKY45 依赖性基因。260 个基因的 BTH 反应性依赖于 WRKY45。其中，220 个基因（85%）被 BTH 上调，其中许多基因编码 PR 蛋白和与次生代谢相关的蛋白。这些基因中只有一小部分与 OsNPR1/NH1 调节的基因重叠，这支持了水稻 SA 途径分为 WRKY45 调节和 OsNPR1/NH1 调节亚途径的观点。地塞米松诱导的水稻中 myc 标记的 WRKY45 的表达立即上调了内源性 WRKY45 和编码转录因子 WRKY62、OsNAC4 和 HSF1 的基因的转录，所有这些基因都被报道具有防御相关功能。随后，编码 PR 蛋白和次生代谢酶的防御基因被上调。许多这些基因在稻瘟病菌感染后也被诱导。它们的时间转录模式与地塞米松诱导 WRKY45 表达后的转录模式一致。在由水稻 coleoptiles 的粒子轰击组成的瞬时表达系统中，WRKY45 作为效应物，转激活 WRKY62 和 OsNAC4 的上游和内含子序列与荧光素酶编码序列融合的报告基因。转录激活通过 OsNAC4 上游包含 W 框的序列发生，并且 W 框中的突变消除了转录激活。
这些数据表明 WRKY45 在 BTH 诱导的抗病性中作为水稻 SA 途径中两个分支之一的防御反应转录级联的主调节因子发挥作用。
