MAP Kinase in Plant Disease Resistance Signaling Pathway

蛋白质磷酸化和去磷酸化在植物抗病过程中起着重要作用，从感知入侵病原体到激活植物防御反应。WIPK是一种烟草丝裂原活化蛋白激酶（MAPK），在非宿主特异性激发子或“基因对基因”相互作用引发的抗性反应中被激活。此外，WIPK mRNA和蛋白的诱导是系统性的，并与系统性获得性耐药（SAR）的建立相关。最近对酵母和动物的广泛研究表明，MAPK级联反应是将细胞外刺激（包括各种应激）转化为细胞反应的主要途径。与细胞中预先存在且只需要磷酸化激活的所有其他特征mapk相反，WIPK激活需要翻译后磷酸化和先前的基因转录以及WIPK蛋白的从头合成。非常有趣的是，WIPK基因的激活是由H2O2介导的，H2O2是植物防御反应中另一个重要的信号分子。这个项目的目标是从功能上定义WIPK在局部和系统抵抗中的作用。分子、生物化学和遗传方法的结合将被用来解决这一目标。为了阐明WIPK的功能，我们将鉴定WIPK的上游激酶，磷酸化和激活WIPK的WIPK激酶（WIPKK）。在体内，WIPK的活性将通过过表达一个组成活性的WIPKK突变体来操纵。这些转基因植物对病原体或激发子的表型变化应该提供WIPK的第一个功能定义。为了补充“功能获得”研究，将尝试使用显性负突变激酶的反义、共抑制和过表达来抑制WIPK或WIPKK表达和/或激活的“功能丧失”转基因系。此外，将鉴定拟南芥中WIPK和WIPKK的同源基因，这将有助于从T-DNA插入突变体池中分离出敲除植株。这项工作的一个实际扩展是，在植物防御信号通路中识别一个重要的调控成分可能导致作物植物的抗病性增强。制定这样一种以增强植物自身防御为基础的战略，对维持农业生产和改善我们的环境质量很重要。对WIPK级联的深入了解还可以为以下方面提供基础：(1)研究植物mapk对其他胁迫（如寒冷、盐和干旱）或植物激素（如乙烯、生长素和ABA）的调控；(2)比较植物、酵母和哺乳动物中应激激活的mapk；(3)识别植物mapk调控和功能的独特方面。同样重要的是，了解H2O2下游的调控成分WIPK的作用，将极大地扩展我们对氧化爆发在局部抗性和SAR建立中的信号作用的认识。

Protein phosphorylation and dephosphorylation play important roles in plant disease resistance at multiple steps from the perception of invading pathogens to the activation of plant defense responses. WIPK, a tobacco mitogen-activated protein kinase (MAPK) is activated during resistance responses initiated by either non-host specific elicitors or 'gene-for-gene' interactions. Furthermore, induction of WIPK mRNA and protein occur systemically and correlates with the establishment of systemic acquired resistance (SAR). Extensive recent research in yeast and animals demonstrated that MAPK cascades are major pathways that transduce extracellular stimuli, including various stresses, into cellular responses. In contrast to all other characterized MAPKs that pre-exist in cells and require only phosphorylation activation, WIPK activation requires both post-translational phosphorylation and a preceding gene transcription and de novo synthesis of WIPK protein. Very interestingly, the activation of WIPK gene is mediated by H2O2, another important signaling molecule in plant defense responses.The goal of this project is to functionally define the role(s) of WIPK in both local and systemic resistance. A combination of molecular, biochemical and genetic approaches will be utilized to address this goal. To elucidate the function of WIPK, the upstream kinase of WIPK, WIPK kinase (WIPKK), that phosphorylates and activates WIPK, will be identified. In vivo WIPK activity will then be manipulated by overexpressing a constitutively active WIPKK mutant. The phenotypic change of such transgenic plants in response to pathogens or elicitors should provide the first functional definition of WIPK. To compliment the "gain-of-function" study, "loss-of-function" transgenic lines with suppressed WIPK or WIPKK expression and/or activation will be attempted by using anti-sense, co-suppression and overexpression of dominant negative mutant kinase. In addition, orthologs of WIPK and WIPKK in Arabidopsis will be identified which will facilitate the isolation of knock-out plants from T-DNA insertional mutant pools. One practical extension of this work is that the identification of an important regulatory component in the plant defense signaling pathway may lead to the engineering of crop plants with enhanced disease resistance. The development of such a strategy, which is based on the enhancement of the plant's own defenses, is important for sustaining agricultural production and improving the quality of our environment. Insights into the WIPK cascade should also provide a basis for (1) studying the regulation of plant MAPKs involved in responses to other stresses (e.g. cold, salt, and drought) or phytohormones (e.g. ethylene, auxin and ABA), (2) comparing stress-activated MAPKs in plants, yeast and mammals, (3) identifying unique aspects in the regulation and function of plant MAPKs. Equally important, understanding the role of WIPK, a regulatory component down-stream of H2O2, will greatly extend our knowledge about the signaling role of the oxidative burst in both local resistance and establishment of SAR.