质膜CYBDOM蛋白促进活性氧产生、侧根发育和抗旱性的机理

CYBDOM is a multigenic protein family ubiquitously existed in eukaryotes, whereas the physiological functions of this type of proteins have not been clearly characterized. It is composed of a C terminal CYB561 and an N terminal DOMON domain. Previously, we have identified a dehydration-induced gene from the resurrection plant Boea hygrometrica, which encoded a plasma membrane-localized CYBDOM protein and was designated BhCYBDOM1. Constitutively expression of this gene in Arabidopsis caused significant ROS accumulation and showed a series of ROS-related phenotypes, including growth retardation, late-bolting, more lateral roots and drought tolerance. Preliminary results indicated that BhCYBDOM1 expressed in the lateral root primordium (LRP), which was in agreement with the observation that ROS accumulated in LRP in the plants overexpressing BhCYBDOM1.BhCYBDOM1 recycled between plasma membrane and endocytotic vesicles in the same manner as PIN1, the major auxin efflux carrier.On the other hand,PIN1 cycling was blocked by BhCYBDOM1.Based on these data, the proposed study aims to further elucidate the roles of CYBDOM proteins in regulating ROS production, LRP formation, and drought tolerance by combinations of molecular, biochemical, cellular and genetic approaches, and by comparative analysis of BhCYBDOM1 with its homologues from Arabidopsis. This project will provide the first experimental evidence for the function of CYBDOM proteins and a new viewpoint for the integrative regulation mechanisms of dehydration tolerance in B. hygrometrica, and eventually be benefit to deepen our understanding of the interaction of ROS, auxin in drought-stressed plants.

CYBDOM蛋白是真核生物中一类含细胞色素b561结构域（CYB561）和多巴胺β-羟化酶结构域（DOMON）的功能未知的膜蛋白。我们从耐脱水的复苏植物旋蒴苣苔中克隆到一个CYBDOM基因，该基因受干旱和活性氧诱导表达，在侧根形成部位大量表达；过表达使转基因植物生长受抑，但ROS含量和侧根数目增加，抗旱性提高。ROS积累在转基因植物的侧根原基部位尤为显著。预试验结果表明该蛋白以类似PIN1内吞的方式进行胞内循环，同时抑制PIN1蛋白的胞内循环。本项目拟在此基础上，利用分子、生化、细胞和遗传学手段对CYBDOM蛋白介导ROS产生、侧根增加、抗旱性提高的功能和作用机理进行深入研究，揭示其调控逆境诱导形态反应的分子基础；并通过与模式植物同源基因的比较研究，阐明CYBDOM蛋白所介导的ROS和生长素的交互调节在植物耐旱性中的作用，为复苏植物耐旱适应的整合生物学机制提供一个新视角。

CYBDOM 蛋白是真核生物中一类含细胞色素 b561 结构域（CYB561）和多巴胺β-羟化酶结构域（DOMON）的功能未知的膜蛋白。我们从耐脱水的复苏植物旋蒴苣苔中克隆到一个 CYBDOM 基因，该基因受干旱和活性氧诱导表达，在侧根形成部位大量表达；过表达使转基因植物生长受抑，但 ROS 含量和侧根数目增加，抗旱性提高。ROS 积累在转基因植物的侧根原基部位尤为显著。预试验结果表明该蛋白以类似 PIN1 内吞的方式进行胞内循环，过表达抑制 PIN1 蛋白的胞内循环。本项目在此基础上，利用分子、生化、细胞和遗传学手段对 CYBDOM 蛋白介导 ROS 产生、侧根增加、抗旱性提高的功能和作用机理进行深入研究，揭示其调控逆境诱导形态反应的分子基础，发现其不依赖于已知的NADPH氧化酶活性，是一个新的质膜ROS信号产生因子，可引起细胞内氧化还原状态的改变和再平衡，其促进侧根增加和生长、促进抗旱性增强均依赖于其产生ROS的活性及ROS与抗旱信号途径和生长素信号途径的交互作用；该类蛋白可以以网格蛋白途径和非网格蛋白依赖的途径进行胞内循环，其胞内循环受生长素调控；其产生活性氧的活性与其与抗坏血酸/脱氢抗坏血酸的结合相关，而不受NADPH氧化酶抑制剂DPI及NADPH氧化酶编码基因突变的影响。通过与模式植物中的同源基因的比较研究，阐明了CYBDOM 蛋白所介导的 ROS 通过生长素的交互调节实现了对生长的抑制，从而为复苏植物耐旱适应提供了节能方案，这为特殊生境植物适应极端干旱逆境的整合生物学机制提供一个新视角；而在生命周期较短的拟南芥中CYBDOM蛋白基本不受干旱诱导，其功能主要在侧根发生与伸长，这可能与其生命周期短，干旱胁迫下通过气孔关闭而维持细胞内水分含量的抗旱策略有关。