果实摄取钙的途径及调控机理的研究

Ca is an important nutrient for fruit quality formation and maintenance. Its deficiency induces various physiological disorders. However, pathway of Ca uptake by fruit is still a matter of dispute, let alone to understand its regulatory mechanisms. Recently, we found a "bottleneck" in Ca transport to fruit in the pedicle of litchi and obtained some evidences suggesting phloem as the chief pathway of Ca transport to fruit. In this project, we are to investigate Ca transport pathways among different types of fruits by anatomical study of the pedicle structure, pathway dye tracing, Sr tracing and pedicle girdling experiments. At the same time, universality of the presence of abundant calcium oxalate in the pedicle will be investigated among different types of fruits, which will be clustered according to the pathways of Ca transport to fruit and the abundance and morphology of calcium oxalate crystals in the pedicle. Regulatory mechanisms in fruits with different Ca transport pathways will be examined by looking into the roles of fruit transpiration, plasma Ca channels and Ca-ATPase, and polar IAA export from fruit. Meanwhile, roles of seed in Ca uptake by fruit and mechanisms of the difference in capacity of fruit Ca uptake among cultivars in litchi will be clarified. In addition, dynamic changes and correlations between Ca accumulation in the fruit and calcium oxalate formation in the pedicle will analyzed in litchi so as to clarify whether formation of calcium oxalate causes the Ca transport "bottleneck". The study will enrich our understanding of Ca nutrition in fruit trees and provide important theoretical references for controlling Ca related physiological disorders.

钙对果实品质形成和维持具重要作用，其供应不足导致各种生理病害。但学界对果实摄取钙的途径尚存争议，更无从明确其调控机理。我们最近发现荔枝果柄存在向果实运输钙的"瓶颈"以及钙通过韧皮部输入果实的证据。在此基础上，本项目将结合果柄结构观察、染料示踪、Sr示踪、果柄环剥等实验进一步在不同类型的果树上展开果实摄取钙途径的探索，对果柄草酸钙存在的普遍性展开调查，并根据钙运输途径及果柄草酸钙丰度和形态对果树进行聚类。与此同时，针对不同运输途径类型的果实，从蒸腾、果柄质膜钙通道、钙泵等钙转运机制活性及果实极性输出IAA的调节作用的角度，探索果实摄取钙的调控机制，并揭示种子发育在果实摄取钙中的作用，探明品种间果实摄取钙能力差异的机理。分析果柄草酸钙形成的生理意义，揭示其与果实钙积累的动态关系，明确其是否导致了钙向果实运输的"瓶颈"。本项目研究将充实果树钙营养生理知识，为克服钙相关生理病害的发生供理论依据。

钙是果实品质形成和保持的重要元素，水果钙也是人们补充钙营养的重要来源，研究果实钙的摄取规律、途径和调控机制具有重要的意义。本项目展开了多树种果实组织钙含量、钙吸收动态、吸收速率和吸收活性，分析典型果树种类（荔枝等）的果实钙摄取途径，及调控机制。发现了果实钙营养生理中的一些普遍规律：1）果实钙摄取速率与果实生长速率正相关；2）12个果树种类中，果柄钙浓度比果实组织高3-10倍；3）电子探针原位分析发现果柄中的钙更多分布在韧皮部；4）以染料示踪发现，在随果实发育成熟过程中，果实-果柄木质部连续性丧失，功能止于果柄-果实连接处。其中苹果导管功能的丧失早于其他果实，但其钙摄取速率位于最高之列。这些结果显示，韧皮部/共质体也是向果实运输的重要通道。用环剥处理荔枝和柑桔果梗，果实生长和钙摄取受限制抑制；用钙离子的类似物锶离子从果梗引入，发现引入锶在果柄中主要分布在韧皮部。在荔枝中，用La3+处理，导致果实成熟和生长受到抑制，也显著降低了钙的摄取。以上结果为韧皮部/共质体通路参与钙向果实的运输提供进一步的证据。以荔枝和龙眼为材料分析了果柄钙的存在形式，表明15-40%的钙以草酸钙形成存在，推测草酸钙形成对平衡钙运输和钙信号具有积极的作用。果柄草酸钙含量和比例有明显的波动，暗示形成的草酸钙可再释放。首次从荔枝检测到草酸氧化酶活性，其活性与草酸钙含量负相关，显示草酸氧化酶可能参与草酸钙的释放。在石峡龙眼、‘岭丰糯’和‘糯米糍’中，‘糯米糍’后期果柄草酸钙最多，与其草酸钙氧化酶活性低有关。克隆了5个草酸氧化酶的潜在基因成员，发现仅LcGLP4表达水平与酶活性具有显著的正相关，其功能有待进一步验证。从荔枝中克隆了钙通道、液泡钙-氢转移体及液泡膜钙泵等3种钙转运体的基因，分析了它们的表达规律，利用VISG成功抑制了液泡膜钙泵基因（LcACAs）表达，导致果实钙积累有所提高，表明液泡钙泵可能是钙向果实运输的负调控者。