拟南芥微管结合蛋白WDL3参与ABA诱导的气孔关闭过程  

The microtubule-associated protein WDL3 mediates ABA-induced stomatal closure in Arabidopsis

作　　者：席小慧 王攀 王朝阳 于荣[1] Xiaohui Xi;Pan Wang;Zhaoyang Wang;Rong Yu(College of Life Sciences,Capital Normal University,Beijing 100048,China)

机构地区：[1]首都师范大学生命科学学院,北京100048

出　　处：《科学通报》2019年第1期95-106,共12页Chinese Science Bulletin

基　　金：国家自然科学基金(31470011;31871351)资助

摘　　要：以拟南芥WDL3RNA干扰株系(WDL3RNAi)和Tubulin5A-YFP植株等为材料,从叶片的失水率、气孔开度、保卫细胞微管骨架动态排布以及Ca^(2+)流动等不同角度探究在脱落酸(abscisic acid, ABA)诱导的气孔关闭信号通路中,微管结合蛋白WDL3与微管骨架以及Ca^(2+)之间的功能关系,深入了解气孔运动机理.结果表明:(1)相同条件下,WDL3RNAi的叶片蒸腾速率明显慢于野生型.(2)气孔开度实验中,WDL3RNAi对ABA信号比野生型更敏感,气孔关闭更快;微管稳定剂紫杉醇(Paclitaxel)可部分阻碍ABA的作用,微管解聚剂黄草消(Oryzalin)则进一步促进ABA诱导的气孔关闭,但WDL3 RNAi与野生型之间仍存在显著差异;激光共聚焦扫描显微镜观察发现, ABA条件下WDL3 RNAi保卫细胞内微管解聚明显加快,微管成束程度(bundling)显著降低.(3)胞内Ca^(2+)螯合剂BAPTA与ABA共同处理,野生型和WDL3RNAi的气孔关闭均受到不同程度的抑制,关闭减缓,处理前后差异显著.亚细胞结构观察发现, BAPTA阻碍了ABA引起的保卫细胞微管解聚,但WDL3 RNAi与野生型相比,依然维持相对较高的微管解聚比例.此外,非损伤微测技术检测发现,ABA引起的保卫细胞Ca^(2+)内流在WDL3RNAi中较野生型的流速更快,流量加大,显示Ca^(2+)在该信号通路中具有重要作用.综上实验结果表明,微管结合蛋白WDL3通过与微管骨架及Ca^(2+)相互作用参与ABA诱导的气孔关闭过程.Stomatal movements control CO2 uptake for plant photosynthesis and water loss by transpiration, then determine plant productivity and water utilization efficiency. The microtubule dynamics is widely recognized to be essential for guard cell function. However, the molecular mechanisms underlying this process remain largely unknown. WDL3 belongs to the microtubule-associated protein WAVE-DAMPENED 2(WVD2)/WVD2-LIKE(WDL) family, which binds to and stabilizes microtubules against low-temperature and dilution-induced depolymerization. In the presence of WDL3, tubulins assemble into large microtubule bundles in vitro, otherwise only single filament patterns form. It has been intensively investigated that WDL3 participates in COP1(CONSTITUTIVE PHOTOMORPHOGENIC 1)-mediated hypocotyl cell elongation in darkness, but whether and how it modulates the microtubule behaviors during guard cell signaling transduction is still an open question. In this study, we dissected the interplay among WDL3, microtubule dynamics and Ca^2+ in ABA-induced stomatal closure. We found that transpirational water loss from detached leaves occurred slowly in the WDL3 RNA interference transgenic line. Stomatal bioassay revealed that guard cell sensitivity to ABA was promoted in the WDL3 RNAi seedlings, and this phenotype could be partially blocked by paclitaxel, a microtubule stabilizing agent. On the other hand, the microtubule-disrupting drug oryzalin, enhanced ABA-triggered stomatal closure even further, and the WDL3 RNAi guard cells were more sensitive to oryzalin treatment. Based on the pharmacological results above, we next tested the effect of WDL3 on cortical microtubules in guard cells directly by Confocal microscopy. The differences in the configurations of microtubule filaments between WDL3 RNAi and wild type(WT) were analyzed after ABA treatment. Microscopic images showed that ABA-induced microtubule disassembly took faster in WDL3 RNAi guard cells than in WT, and the extent of filament bundling decreased significantly in WDL3 RNAi seedli

关 键 词：WDL3 微管骨架 CA^2+ 气孔关闭 脱落酸 

分 类 号：Q945[生物学—植物学]