受低毒病毒调控的板栗疫病菌总蛋白质组学

Proteomic analysis of Cryphonectria parasitica infected by a virulence-attenuating hypovirus

摘要:

摘要:【目的】为解析低毒病毒调控板栗疫病菌生理性状和致病性的机制，在总蛋白质组水平上寻找受病毒侵染调控的宿主蛋白及其编码基因。【方法】使用双向电泳方法对野生型菌株EP155和受低毒病毒感染的菌株EP713进行差异蛋白质组分析，同时，对差异表达蛋白质编码基因的mRNA水平进行定量分析。【结果】共找到71 个因病毒侵染而发生差异表达的蛋白质点，分别属于58种不同蛋白质。以EP155为对照，表现为上调的19个，下调的52个，主要涉及能量代谢，蛋白质、核酸和碳水化合物代谢，信号传导以及压力应激和氧化还原反应。对10个受病毒调控的蛋白的编码基因进行了mRNA水平定量，其中7个基因受病毒感染后的mRNA水平变化与蛋白水平变化趋势一致，3个基因的mRNA水平变化与蛋白水平变化趋势不相符，表明低毒病毒对板栗疫病菌不同基因的调控可以发生在不同的调控层面上。【结论】低毒病毒的侵染弱化了宿主TCA循环的能量流动，调控了宿主体内的甲基化过程及真菌致病因子的表达。

Abstract:

Abstract:［Objective］Chestnut blight fungus Cryphonectria parasitica and hypovirus constitute a model system to study fungal pathogenesis and host-virus interaction． Proteomic analysis of chestnut blight fungus upon hypovirus infection was conducted to find the differentially expressed host proteins．［Methods］According to the characteristics of this filamentous fungus，an optimized extraction protocol for fungal total protein was developed． Two-dimensional electrophoresis (2-DE) was used for comparative proteomic analysis of wild strain EP155 and hypovirus-infected strain EP713．The quantitative RT-PCR was applied to analyze mRNA expression level of protein-coding genes.［Results］In total 71 protein spots were detected to be differentially expressed on the base of EP155 of which 19 up-regulated and 52 down-regulated. Fifty-eight unique proteins were identified by mass spectrometry． Further study on quantitative RT-PCR indicated that the regulation of related host genes by hypovirus occurred at different levels.［Conclusion］ The TCA cycle of C．parasitica was weakened after hypovirus infection and the process of methylation was regulated by hypovirus． Meanwhile，viral regulation of virulence factors also contributed to the phenomenon of hypovirulence．