RING类E3连接酶TEAR1调控拟南芥叶片发育

Leaves are the major organs for photosynthesis. The leaf size and morphology affect crop production directly. It is proposed that the activities of plant specific TCP transcription factors control leaf development. We have recently identified a transcriptional repressor which suppresses TCP activities at the protein level. However, the derepression of TCP activities by dissociation of TIE1 from TCP is still unknown. In this project, we find a protein named as TEAR1 which interacts with TIE1 in vitro and in vivo. TEAR1 has E3 ligase activity. Overexpression of TEAR1 causes down-curled leaves similar those observed in TIE disruption mutant, indicating that TEAR1 may play important roles in plant leaf development. We will elucidate the function and the molecular mechanism of TEAR1 in leaf development. We will examine the phenotypes of TEAR1 overexpression lines and TEAR1 knock-out mutants. We will analyze the physical and genetic interactions between TEAR1 and TIE1. The expression pattern of TEAR1 gene and the subcellular localization of TEAR1 protein will also be investigated. The downstream genes and the molecular mechanism by which TEAR1 regulates leaf development will be finally demonstrated.

叶片是植物主要的光合作用器官，其形状和大小直接影响农作物的产量。已有研究表明植物特有的转录因子TCP的活性对调控植物叶片的形状和大小起到关键作用。申请人近期发现了转录抑制因子TIE可在蛋白水平上抑制TCP的活性，进而调控叶片的发育，但TIE如何从TCP上解离的机制还不清楚。本项目在前期研究的基础上，通过酵母双杂交筛选获得了一个具有RING锌指结构域的蛋白TEAR1可和TIE相互作用。初步结果表明TEAR1可和TIE1在体内和体外直接相互作用；体外检测确定TEAR1具有E3泛素连接酶的活性；过量表达TEAR1使叶片向下卷曲，与TIE功能缺乏的表型相似。本项目拟深入研究TEAR1调控叶片发育的功能和分子机制，研究TEAR1突变体和过量表达的表型，分析TEAR1和TIE1的直接相互作用以及遗传互作，研究TEAR1的亚细胞定位和表达模式，最终找出TEAR1下游基因以及调控叶片发育的分子机制。

植物叶片形态和大小的可塑性发育对于叶片功能及植物适应多变的生存环境非常重要，但植物如何根据环境条件形成合适的叶片形态和大小的分子机制还不是很清楚。本课题组前期工作鉴定到一类以TIE1为代表的转录抑制因子，这类转录抑制因子通过招募共抑制因子TPL/TPRs在蛋白水平上抑制TCP转录因子的活性，从而调控叶片发育，但是TIE1 本身是如何调控的还未知。本研究以TIE转录抑制因子蛋白为诱饵通过酵母双杂交方法筛选到与TIE1相互作用的含有RING结构域的E3泛素连接酶TEAR1（TIE1- associated RING-type E3 ligase 1）。实验证明TEAR1具有泛素连接酶的活性，而TIE1确实在植物体内能被泛素化并通过降解调控。降低TEAR1及其同源基因的功能可导致叶片边缘过度生长和叶锯齿增多等表型，这与tcp功能缺失突变体的表型相一致。遗传分析表明过量表达TEAR1可拯救TIE1过量表达导致的叶片发育缺陷的表型，而TEAR1缺失可增强因TCP转录因子活性降低引起的叶片发育缺陷的表型。本研究不仅首次发现了一类RING类泛素连接酶在调控叶片发育过程中的重要功能，还揭示了一种新的精确调控TCP活性和叶片发育的分子机制。TEAR1根据内部条件和外部环境决定TIE1的降解来解除TIE1对TCP活性的抑制，从而正调控TCP的活性。通过这种微调机制，植物能灵活地根据内外条件的变化精确调控叶片的形态和大小。