白桦C2H2型锌指转录因子的耐盐调控机理研究

The plant C2H2 zinc finger proteins represent a large of transcription factor family and numerous members of this family have been shown to play diverse roles in many biological processes, including plant growth, development and stress responses. In the previous study, five C2H2-type zinc finger transcription factor genes, named BpZFP, were identified that their expressions are induced by high-salt stress and four of them were confirmed that their involvement in salt tolerance in Birch. On these bases, we will further investigate two genes in two aspects: （1）To clone their promoters and define the cis-elements involved in the expression of BpZFP gene under high-salt stress and involved in signal transduction of stresses and analyze their upstream regulators and signal transduction pathway; （2）To identify the downstream target genes and the metabolic pathways regulated by these genes encoding products. This proposed project may contribute to reveal the molecular mechanism of the C2H2-type zinc finger transcription factor gene in regulation of salt stress tolerance in Birch and provide the theoretical and practical bases in genetic engineering to improve the stress tolerance in trees.

植物C2H2型锌指蛋白是成员众多的转录因子家族，在植物生长发育和非生物胁迫反应等过程中具有重要作用。我们在前期研究中鉴定了五个能被盐胁迫诱导表达的白桦C2H2型锌指蛋白基因（命名为BpZFP）, 研究证实其中的四个BpZFP基因可能参与白桦的耐盐应答。本项目在前期工作的基础上，拟对其中两个BpZFP 基因进一步开展以下的研究：（1）克隆BpZFP启动子；鉴定启动子中包含的重要耐盐相关顺式作用元件；研究其上游的调控因子和信号通路。（2）研究BpZFP所调控的下游靶基因；分析这些基因的产物所参与或调控的代谢途径。本项目的目的是研究BpZFP转录因子基因调控白桦耐盐的分子机理，为锌指蛋白基因在林木抗盐基因工程上的应用提供理论依据和优良基因。

植物锌指蛋白在植物生长、发育和非生物胁迫响应等过程中具有重要作用。本研究中我们克隆鉴定了一个C2H2型锌指蛋白基因BpZFP，其转录能被盐、干旱和ABA胁迫诱导。分子生物学分析证明白桦BpZFP蛋白是一个定位于细胞核的转录活化型转录因子。我们克隆出BpZFP基因启动子，鉴定了其中受盐、旱等胁迫诱导的重要顺式调控元件，并筛选出通过与BpZFP基因启动子上的相应顺式作用元件结合来调控BpZFP基因转录的上游调控因子。创制了过表达和抑制表达BpZFP转基因白桦，明确了BpZFP过表达增强了转基因白桦的株高、根长、鲜重、叶绿素含量、脯氨酸含量以及SOD和POD酶活性，降低了转基因白桦中MDA和H2O2含量。从全基因组水平研究了BpZFP调控的白桦基因表达谱，发现BpZFP显著上调一些逆境响应和防卫基因的表达。以上结果表明，BpZFP基因过表达显著提高了转基因白桦的生物量、增强了活性氧清除能力、提高了抗氧化酶活性以及调控了一些抗逆相关基因的表达，从而正向调控白桦对高盐、干旱等逆境的耐受性。因此，本研究为揭示白桦抗逆分子机制奠定了基础，具较重要的理论与实践意义。