Polarity control and cross-talk between MAPK signalling, cascades, vesicular trafficking and the actin cytoskeleton in plant root hairs

植物根毛中 MAPK 信号、级联、囊泡运输和肌动蛋白细胞骨架之间的极性控制和串扰

• 批准号: 27050727
• 负责人: Professor Dr. Jozef Samaj
• 金额: --
• 依托单位: Biotechnological and Agricultural Research
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2006
• 资助国家: 德国
• 起止时间: 2005-12-31 至 2010-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/27050727?language=en
• 关键词: Polarity; control; cross; talk; between

Mitogen-activated protein kinases (MAPKs) are evolutionary conserved signal transducers involved in the regulation of plant development. In root hairs, crosstalks between SIMK (stress-induced MAPK), the actin cytoskeleton, and vesicular trafficking are essential for tipgrowth (Samaj et al. 2002, 2004a,b). Tip-growth is known to be controlled also by other signalling molecules such as Rop GTPases (Rop2/4), reactive oxygen species (ROS) and phospholipase D (PLD)/phosphatidic acid (PA) (Jones et al. 2002, Foreman et al. 2003, Ohashi et al. 2003) as well as by actin and actin-binding protein profilin (Baluska et al. 2000, Ramachandran et al. 2000, Ringli et al. 2002). In the context of this proposal it is important, that both ROS and PA activate MAPKs in plants and cause reorganization of actin cytoskeleton and vesicular trafficking on the cellular level. Moreover, PA and ROS can be integrated into MARK cascade involving MPK3 and MPK6 via OXI1/AGC2-1 kinase, which is a novel protein kinase that has recently been shown to be involved in Arabidopsis root hair tip growth (Anthony et al. 2004, Rentel et al. 2004). We have found that small amounts of MPK4 and MPK6, but not MPK3, are constitutivelly associated with endosomes. However, MPK3 is activated by ROS, salt and heat, and relocated to an endosomal compartment in its activated state. Upon stress activation, MPK6 relocates to broader spectrum of endomembranes including endoplasmic reticulum (ER) and endosomes. Additionally, SIMK binds to actin and MPK6 associates with actin filaments. This could be related to the actin role in the motility and other functions of ER and endosomes, representing two distinct endomembrane compartments. On the other hand, MPK4 is activated by heat, but not by ROS, and this active form is also relocated to endosomes. Here we aim to study cross-talks among ROS-, PA- and OXIl-mediated regulation of MAPKs, vesicular trafficking and the cytoskeleton in root hairs of Arabidopsis. We will isolate new Ioss-of-function (knockout) mutants for two MAPKs (MPK3 and MPK6) and characterise (using genetic, cell biological, molecular biological and biochemical approaches) those of them which are showing root hair phenotypes. We will also further characterise known mpk4 mutant showing ectopic and branched root hairs as well as other mutants defective in potential upstream signalling components such asyoda, apn2xapn3, oxil and rhd2 as well as rop2 constitutive-active and dominat-negative functional mutants, which are all showing root hair phenotypes (longer/shorter or branched root hairs). Finally, we will study two profilins PRN1 and PRN2 as potential downstream targets of MPK3/4/6 as well as MPK6 interaction with actin and MPK3/4 interactions with endosomal Rab GTPases. We expect that the data gathered in the course of this project will have a strong impact on our understanding of polarity control in plant and other eukaryotic cells.

有丝分裂原活化蛋白激酶（MAPK）是一种进化上保守的信号转导蛋白，参与植物发育的调控。在根毛中，SIMK（胁迫诱导的MAPK）、肌动蛋白细胞骨架和囊泡运输之间的串扰对于根尖生长至关重要（Samaj et al. 2002，2004 a，B）。已知尖端生长还受其他信号分子如Rop GTP酶（Rop 2/4）、活性氧（ROS）和磷脂酶D（PLD）/磷脂酸（PA）控制。（Jones et al.2002，Foreman et al.2003，Ohashi et al.2003）以及肌动蛋白和肌动蛋白结合蛋白profilin（Baluska等人，2000年; Ramachandran等人，2000年; Ringli等人，2002年）。在这一建议的背景下，重要的是，ROS和PA都激活植物中的MAPK，并导致肌动蛋白细胞骨架的重组和细胞水平上的囊泡运输。此外，PA和ROS可以通过OXI 1/AGC 2 -1激酶整合到涉及MPK 3和MPK 6的MARK级联中，OXI 1/AGC 2 -1激酶是一种新的蛋白激酶，其最近被证明参与拟南芥根毛尖生长（Anthony et al.2004，Rentel et al.2004）。我们已经发现，少量的MPK 4和MPK 6，而不是MPK 3，与内体组成性相关。然而，MPK 3被ROS、盐和热激活，并在其激活状态下重新定位到内体区室。在应激激活后，MPK 6重新定位至更广谱的内膜，包括内质网（ER）和内体。此外，SIMK与肌动蛋白结合，MPK 6与肌动蛋白丝结合。这可能与肌动蛋白在ER和内体的运动和其他功能中的作用有关，代表两个不同的内膜隔室。另一方面，MPK 4被热激活，但不被ROS激活，并且这种活性形式也被重新定位到内体。在这里，我们的目标是研究交叉谈判的ROS-，PA-和OXII介导的调节MAPKs，囊泡运输和细胞骨架在根毛的拟南芥。我们将分离出两种MAPK（MPK 3和MPK 6）的新的功能丧失（敲除）突变体，并（使用遗传学、细胞生物学、分子生物学和生物化学方法）筛选出其中显示根毛表型的突变体。我们还将进一步研究已知的显示异位和分支根毛的mpk 4突变体以及在潜在上游信号传导组分中有缺陷的其他突变体，如asyoda、apn 2xapn 3、oxil和rhd 2以及rop 2组成型活性和显性阴性功能突变体，它们都显示根毛表型（较长/较短或分支根毛）。最后，我们将研究两个profilins PRN 1和PRN 2作为MPK 3/4/6的潜在下游靶点，以及MPK 6与肌动蛋白的相互作用和MPK 3/4与内体Rab GTP酶的相互作用。我们希望在这个项目的过程中收集的数据将对我们理解植物和其他真核细胞的极性控制产生强烈的影响。