微管骨架和PP1/PP2A蛋白磷酸酶在ALA-ABA调控苹果叶片气孔运动中的作用  

The Role of Microtubule Skeleton and PP1/PP2A Protein Phosphatase in ALA-ABA Regulating Stomatal Movement in Apple Leaves

作　　者：熊丽君[1] 安玉艳[1] 汪良驹[1] XIONG Lijun;AN Yuyan;and WANG Liangju(College of Horticulture,Nanjing Agricultural University,Nanjing 210095,China)

机构地区：[1]南京农业大学园艺学院,南京210095

出　　处：《园艺学报》2018年第11期2073-2088,共16页Acta Horticulturae Sinica

基　　金：国家自然科学基金项目(31772253)

摘　　要：以苹果离体叶片下表皮为材料,利用不同药剂,结合激光扫描共聚焦显微镜观察和qRT-PCR等技术,从气孔开度、保卫细胞H_2O_2水平和基因表达等方面,研究了微管骨架和PP1/PP2A蛋白磷酸酶在5–氨基乙酰丙酸(ALA)抑制脱落酸(ABA)诱导的气孔关闭过程中的作用及可能的调控机制。结果表明,微管解聚剂长春碱(Vinblastine,VBT)可以消弱ALA对苹果叶片气孔开放的促进效应,而微管稳定剂紫杉醇(Taxol)可以削弱ABA诱导的气孔关闭效应,说明ALA抑制ABA诱导的苹果叶片气孔关闭过程依赖于保卫细胞的微管聚合。与此同时,外源ALA可以逆转ABA对微管蛋白编码基因TUB1、MAP65-1和MAP65-3表达的抑制效应,说明ALA通过促进保卫细胞微管骨架聚合来促进气孔开放。另一方面,PP1/PP2A蛋白磷酸酶抑制剂冈田酸(OA)既能引起微管解聚,也能引起气孔关闭,说明保卫细胞的微管聚合和气孔开放受到PP1/PP2A的正调控;同时,OA促进保卫细胞H_2O_2积累,导致微管解聚和气孔关闭。以上结果表明,ALA可能通过促进苹果叶片保卫细胞PP1/PP2A蛋白磷酸酶活性,抑制ABA诱导的H_2O_2积累,促进微管蛋白基因表达和微管聚合,抑制ABA诱导的气孔关闭,从而为苹果叶片光合气体交换打开气孔通道。这为ALA在农业生产上的应用提供理论依据。The stomatal aperture,guard cell H 2 O 2 levels and gene expressions were studied pharmacologically in the abaxial epidermis of detached apple(Malus×domenstica Borkh.'Fuji')leaves with laser scanning confocal microscopy and qRT-PCR techniques to investigate the role and possible regulatory mechanisms of microtubule skeleton and PP1/PP2A protein phosphatase in the inhibitory effect of 5-aminolevulinic acid(ALA)on abscisic acid(ABA)-induced stomatal closure. It was observed that vinblastine(VBT),a microtubule disassemble agent,depressed stomatal opening induced by ALA,while taxol,a microtubule stabilizer,undermined stomatal closure induced by ABA. This suggests that ALA inhibits ABA-induced stomatal closure depending on the microtubule polymerization of the guard cells. Exogenous ALA reversed the down-regulation of the expressions of tubulin coding genes including TUB1, MAP65-1 and MAP65-3 induced by ABA,suggesting that ALA promoted stomatal opening by promoting microtubule polymerization in guard cells. On the other hand,okadaic acid(OA),an inhibitor of PP1/PP2A protein phosphatase caused both microtubule depolymerization and stomatal closure,indicating that microtubule polymerization and stomatal opening of the guard cells were positively regulated by PP1/PP2A. Furthermore,OA promoted H 2 O 2 accumulation in guard cells,which caused microtubule depolymerization and stomatal closure. This suggests that PP1/PP2A acts at the upstream of H 2 O 2 generation. The above results indicate that ALA may promote PP1/PP2A protein phosphatase activity in apple leaf guard cells,and then inhibit ABA-induced H 2 O 2 production,which up-regulates the expression of tubulin genes and promotes microtubule polymerization of stomatal guard cells,suppressing ABA-induced the stomatal closure. Taken together,we revealed a possible mechanism for ALA-improved photosynthesis in plants,providing a theoretical basis for the application of ALA in agricultural production.

关 键 词：苹果 ABA ALA H2O2 微管 PP1/PP2A蛋白磷酸酶 气孔 

分 类 号：S661.1[农业科学—果树学]