小麦Metacaspase I型基因在小麦细胞死亡及抗条锈中的作用机理

Wheat stripe rust caused by Puccinia striiformis f. sp. tritici (Pst), is an important biological disaster, which severely affects the security of wheat production in China. The rapid virulence variation of Pst results in "resistance loss" of wheat cultivars, and leads to the frequent epidemic of the disease. So it is particularly urgent to reveal the mechanism of wheat and Pst interactions for durable disease control pertinently. In previous studies, we identified two metacasepase I genes by analyzing the whole transcriptome of wheat infected by Pst, which were significantly induced during the compatible wheat-Pst interactions. Silencing of them enhanced the resistance of wheat to Pst, indicating their roles in the susceptibility of wheat to Pst. Previous reports suggested that metacaspase could regulate the hypersensitive response (HR) in plants, and then affect the plant resistance to pathogens. Thus, in this project, we plan to further study the function of metacasepase I genes in the susceptible interaction of wheat and Pst, analyse their regulation roles in cell death and caspase activity, identify their target genes by using the yeast and Brachypodium research system via transient expression, gene silencing, yeast two hybrid, RNAi technique and so on. Our results will reveal the modulation mechanism of metacaspase I cell death and wheat susceptibility to Pst, and lay the foundation for clarifying the mechanism of wheat and stripe rust interactions, which will furthermore provide theoretical basis and technical support to develop new strategies to contro disease persistently.

小麦条锈菌引起的小麦条锈病是影响我国小麦安全生产的重要生物灾害。由于条锈菌毒性变异快，小麦品种抗性不断"丧失"，导致该病害频繁爆发。因而解析小麦与条锈菌互作机理，有针对性持久控制病害显得尤为迫切。在前期小麦与条锈菌互作全转录组分析中，获得了两个metacaspase I基因，它们在小麦与条锈菌亲和互作中诱导表达，沉默后可增强小麦抗病性，推测其与小麦感病性有关。而研究表明，metacaspase通过调控HR影响植物对病原菌的抗性。因此，本项目拟通过瞬时表达、基因沉默、酵母双杂交、RNAi等技术方法，借助酵母、短柄草等研究体系，深入解析其在小麦感条锈菌中作用，明确其caspase活性及在细胞坏死中的功能，鉴定其互作靶标，以揭示metacaspase I基因在小麦细胞坏死及感病中的作用机理，为阐明小麦与条锈菌互作机理奠定基础，进而为利用感病基因制定持久防治病害新策略提供理论依据和技术支撑。

小麦条锈菌引起的小麦条锈病是影响我国小麦安全生产的重要生物灾害。由于条锈菌毒性变异快，小麦品种抗性不断“丧失”，导致该病害频繁爆发。因而解析小麦与条锈菌互作机理，有针对性持久控制病害显得尤为迫切。本项目通过瞬时表达、基因沉默、酵母双杂交、RNAi 等技术方法，借助酵母、短柄草、小麦等研究体系，开展了系列研究工作，取得了重要进展，达到了预期目标。1）鉴定获得了小麦半胱氨酸蛋白酶 I 型基因 TaMCA1 和 TaMCA3。证明TaMCA1在体外不具有动物典型半胱氨酸蛋白酶 Caspase-1的酶活性，TaMCA3 蛋白在体外不具有 Caspase-3 活性，但二者在体内均具有Caspase酶的活性；2）TaMCA1 和 TaMCA3 基因不能诱导烟草或小麦叶片产生细胞坏死，但可以抑制由细胞坏死诱导因子 Bax 诱导的细胞坏死反应；3）TaMCA1 和 TaMCA3 基因分别沉默后，小麦对条锈菌的抗性增强，表明其参与了小麦与条锈菌的亲和反应过程；4）筛选鉴定TaMCA3 的互作靶标为左旋天冬酰胺酶 TaASP ，进一步的研究表明 TaMCA3 可能通过激活天冬酰胺酶的活性，天冬酰胺水解成天冬氨酸和氨，参与植物细胞的程序性死亡过程，从而调控小麦的抗病性。通过该项目的研究，揭示了 metacaspase I 基因在小麦细胞坏死及小麦感条锈病中的作用机理，为深入阐明小麦与条锈菌的互作机理奠定了基础，进而为利用感病性基因创制新型抗病材料提供了支撑。