辣椒疫霉RXLR型效应子PcAvh2的序列多态性、基因转录特征及功能

Sequence polymorphism, gene transcriptional pattern and function analysis of the RXLR effector PcAvh2 from Phytophthora capsici

摘要:

[目的]分析辣椒疫霉中RXLR型效应子PcAvh2的序列多态性，研究该效应子在辣椒疫霉生长发育和侵染阶段的转录特征及其生物学功能。[方法]本研究通过高保真扩增，分析2个烟草疫霉、1个恶疫霉和31个辣椒疫霉菌株的PcAvh2序列；提取辣椒疫霉菌丝、游动孢子囊、游动孢子、萌发休止孢和7个侵染时间点（1.5、3、6、12、24、36、72 h）的本氏烟根部总RNA，利用RT-qPCR分析PcAvh2的转录表达水平；利用PVX瞬时表达系统，分析PcAvh2是否抑制6种效应子（BAX、INF1、PsojNIP、PsCRN63、PsAvh241、R3a/Avr3a）激发的植物免疫反应；利用CaCl₂-PEG介导的原生质体稳定转化技术，沉默PcAvh2基因，分析辣椒疫霉致病力的变化。[结果]PcAvh2为典型的RXLR效应子，在辣椒疫霉群体中该效应子具有10个等位基因，而且烟草疫霉和恶疫霉中也存在该效应子。该基因在辣椒疫霉的侵染阶段上调表达，它能够抑制6种效应子激发的植物免疫反应，进一步研究发现基因沉默导致辣椒疫霉的致病力显著下降。[结论]RXLR型效应子PcAvh2是辣椒疫霉中一个重要的侵染致病因子。

Abstract:

[Objective] The aim was to analyze the sequence polymorphism, gene transcriptional pattern and functions of the RXLR effector PcAvh2 from Phytophthora capsici.[Methods] We cloned PcAvh2 gene by high-fidelity PCR from 31 P. capsici isolates, 2 P. parasitica isolates and 1 P. cactorum isolate. Gene expression changes during the developmental stages (mycelia, zoosporangia, zoospores and germinated cysts) and infection period (1.5, 3, 6, 12, 24, 36, 72 h post-inoculation) of P. capsici were monitored by quantitative RT-PCR. PVX-based agroinfiltration assay was performed to examine if PcAvh2 could suppress plant immunity triggered by 6 effectors (BAX, INF1, PsojNIP, PsCRN63, PsAvh241 and R3a/Avr3a). The CaCl₂/PEG-mediated protoplast transformation of P. capsici was conducted to silence PcAvh2 and determine if its silencing affected the pathogen's virulence.[Results] PcAvh2 is a typical RXLR effector and possesses 10 alleles in the population. Furthermore, it also exists in P. parasitica and P. cactorum. The expression of PcAvh2 was up-regulated during the host infection by P. capsici. It can suppress the plant immunity induced by all 6 effectors. Intriguingly, silencing of PcAvh2 in P. capsici significantly compromised the pathogen's virulence on host plants.[Conclusion] RXLR effector PcAvh2 is one of important pathogenicity factors in P. capsici.