拟南芥MKK7-MPK6级联信号调控植株株型的分子机理研究

Mitogen-activated protein kinase (MAPK) cascades play an important role in transducing environmental and developmental signals into adaptive and programmed responses. In higher plants, MAPK cascades are involved in a broad spectrum of signals, which include biotic and abiotic stresses, and hormone-mediated development. In the MAPK signaling module, MKKs are of particular importance because they serve as the convergence and divergence points in the MAPK signal transduction. The complexity of MAPK cascades raise challenges not only to identify the MAPK module in planta but also to define the specific role of an individual module. Our previous study demonstrated that MPK6 and MPK3 are two major downstream targets of MKK7, and mostly strikingly, polar auxin transport and defense response are transmitted by MKK7-MPK6 and MKK7-MPK3 signalling pathways independently. It is indicated that MKK7-activated cascades involve in many development and hormone signaling pathways through phosphorylating different substrates. To identify substrates of MPK6, the yeast two-hybrid library screening was recently carried out to identify MPK6 interacting proteins. In this proposal, we will study the function of MKK7-MPK6 downstream specific substrate and explain how MKK7-MPK6 signaling pathway regulates polar auxin transport through the specific substrate to determine plant architecture in Arabidopsis.

促分裂原活化蛋白激酶级联信号（MAPK）途径是真核生物中普遍存在的非常保守和重要的信号转导通路，它通过复杂的信号转导过程将细胞的外源刺激准确地转变为细胞内的生理反应，从而调控生物体的生长、发育、代谢、凋亡及对外界生物和非生物胁迫应答等多方面的生理过程。在MAPK信号途径中，MKKs是信号转导的枢纽，起到了信号整合和发散的作用。由于MAPK级联信号的复杂性，关于它的特异性调控成为本领域具有挑战性的科学问题。我们之前的研究发现，拟南芥MKK7通过磷酸化MPK6负调控生长素极性运输从而影响植物的株型，另一方面通过激活MPK3正调控植物的基础及系统获得性抗性，暗示MKK7级联信号可能通过磷酸化不同的下游底物调控特异的生物学途径。本研究拟通过酵母双杂交筛库的方法，获得MPK6的底物蛋白，解析底物蛋白的磷酸化位点及作用机制，深入阐明MKK7-MPK6级联信号通过磷酸化特异性底物调控植物株型的分子机理。

MAPK级联信号途径的特异性调控是当前MAPK级联信号研究最具有挑战性的问题之一。在本项目中，我们以酵母双杂交筛库的结果为研究基础，鉴定了MKK7-MPK6级联信号调控植物株型的特异性底物，初步阐明了MKK7-MPK6级联信号通过磷酸化特异性底物的特定位点调控植物分枝的分子机理，为进一步解析MAPK级联信号的特异性调控机制提供了信息。具体研究进展如下：1）利用双分子荧光互补的实验(bimolecular fluorescence complementation, BiFC)对酵母双杂交筛库结果中MPK6候选互作蛋白（MPK6 interacting protein，MIPs）与MPK6的互作进行了验证。2）通过突变体表型的分析，发现MIP4蛋白的突变可以完全恢复bud1多分枝的表型。（3）鉴定了MIP4上被MKK7-MPK6级联信号特异性磷酸化的位点，并深入解析了磷酸化位点的调控机制。（4）通过生长素极性运输的测定和对PIN1蛋白极性的观察，发现MIP4蛋白的突变可以基本恢复bud1花序轴中生长素极性运输的缺陷及bud1茎基部异常的PIN1定位。本项目的成果揭示了MKK7-MPK6级联信号通过磷酸化MIP4调控生长素极性运输介导的植物分枝的分子机理，是MAPK特异性调控领域创新性的发现。