拟南芥磷脂酶D调控微管形态和应答高温胁迫的分子机理

The growth and development of plants are exposed to abiotic stresses such as high temperature. The sructural bilayers in plant plasma membrane (PM) sense stresses and transduce downstream signals, and the components, structure and metabolism of phospholipids affect the stress resistance of plants.The PM-localized PLDδ hydrolyzes phospholipids to generate generate signal PA.Here we will study the molecular mechanism of PLDδ in regulating microtubules organization, calcium and heat shock protein 70 (HSP70) in response to high temperature stress. We will determine the specific interacting sites in PLDδ bound to microtubules and effect of PLD on microtubule dynamics, and we will explore how PLDδ regulate microtubule dynamics by polymerization or depolymerization, which would affect cellular and plant thermotolerance. We will further study the signal position between PLDδ protein and calcium signaling, and the response pattern of their interaction under high temperature stress. We will determine the effect of interaction of PLDδ and microtubules on the HSP70. The study above will reveal a lipid-, calcium- and cytoskeleton-involved signal pathway in response to high temperature stress, which would provide scientific basis and genetic resources for improving crop thermotolerance.

植物的生长发育受到高温等各种非生物胁迫。植物质膜的结构支架-磷脂双分子层参与感受胁迫和传递信号，其磷脂的组分、结构和代谢都关系到植物的抗胁迫能力。质膜蛋白磷脂酶Dδ（PLDδ）水解甘油磷脂产生信号分子磷脂酸。本项目研究高温胁迫响应途径中，拟南芥PLDδ对微管骨架的调控作用及其对钙信号、植物热休克蛋白70（HSP70）的分子调控机理。体内外探寻PLDδ蛋白与微管的结合位点及其动力学特征；结合生物化学、细胞生物学和遗传学方法，明确PLDδ蛋白如何调节微管动态，进而调控微管的解聚或聚合，并进一步影响细胞和植株的耐热性。利用钙指示探针YC3.6，研究PLDδ蛋白与钙信号的位置关系，及其二者对微管形态的调控作用和高温胁迫的应答模式；探寻PLDδ与微管的互作对HSP70的调控作用。目的是揭示由磷脂、钙信号和细胞骨架共同介导的植物抗热信号通路，为提高农作物抗高温的分子改良提供科学依据和基因资源。

细胞骨架和膜系统的互作参与调控植物发育和逆境应答等重要过程。生物学家一直致力于寻找、鉴定连接膜和细胞骨架的桥梁蛋白。我们研究表明，拟南芥PLDδ定位在质膜，并与微管共定位。PLDδ体内外结合微管，并抑制微管聚合。点突变结果表明降低PLD活性显著抑制其去稳定微管活性。热处理后诱导微管从质膜解离和解聚及其幼苗死亡，但是该效应在pldδ突变中得到缓解。总之，PLD负调控微管稳定和植物耐热性。保卫细胞微管是决定气孔应答黑暗和ABA激素的重要因子。气孔开放时，保卫细胞微管呈辐射状分布；在黑暗和ABA处理后，微管解聚。磷脂酸PA结合蛋白磷酸酶PP2CA，二者互作参与调控保卫细胞微管对ABA信号途径的调控。另外，我们利用药物学和遗传学等证据表明，PLD参与调控微管药物处理后的微管形态和细胞发育。微管稳定试剂paclitaxel处理pldδ后，与野生型相比，其根长、下胚轴和微管分布存在显著不同；外源PA能够恢复pldδ表型。总之，拟南芥PLD/PA通过调控微管分布调节细胞发育，而且是依赖于其活性的。

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