Role and Mechanism of Suppressed Apoplastic Serine Hydrolase Activities during Infection with Pseudomonas syringae

植物细胞周围的细胞外空间（质外体）可以说是地球上最大的生物界面。这个表面是潮湿的，营养丰富，因此许多微生物，主要是真菌和病原菌，已经适应了在这种环境中茁壮成长。质外体也充满了水解植物酶，甚至在微生物感染引发的免疫反应期间分泌更多。这些质外体水解酶被认为降解微生物组分并释放引发免疫的激发子。新出现的情况是，微生物能够操纵质外体来抑制这些宿主酶的活性。然而，这种情况在多大程度上发生，如何以及为什么发生，仍然知之甚少。我们的初步数据表明，分泌型植物丝氨酸水解酶的活性受到抑制，在烟草感染假单胞菌注射。有趣的是，质外体蛋白质组的调查显示，植物编码的Kunitz抑制剂在感染植物的质外体中的积累，导致有趣的假设，假单胞菌有目的地诱导宿主的质外体Kunitz水平，以抑制质外体中的水解酶活性。本研究旨在证实这一假说，揭示水解酶活性受抑制的生物学相关性，并阐明更深层次的分子机制。我们将使用综合方法，包括靶向基因沉默，合成（突变）基因互补，活性分析和病原体检测。

The extracellular space surrounding plant cells (the apoplast) is arguably the largest biotic interface on earth. This surface is humid and rich in nutrients and consequently many microbes, mostly commensal and pathogenic bacteria, have adapted to thrive in this environment. The apoplast is also littered with hydrolytic plant enzymes and even more are secreted during immune responses triggered upon microbial infection. These apoplastic hydrolases are thought to degrade microbial components and release elicitors that trigger immunity. The emerging picture is that microbes are able to manipulate the apoplast to suppress these host enzyme activities. However, to what extend this occurs and how and why is still poorly understood. Our preliminary data demonstrates that the activities of secreted plant serine hydrolases are suppressed during infection of Nicotiana benthamiana with Pseudomonas syringe by secreted inhibitors. Interestingly, surveys of apoplastic proteomes revealed the accumulation of a plant-encoded Kunitz inhibitor in the apoplast of infected plants, leading to the intriguing hypothesis that Pseudomonas purposefully induces apoplastic Kunitz levels in the host to suppress hydrolase activities in the apoplast. This proposal is aimed at confirming this hypothesis and revealing the biological relevance of the suppressed hydrolase activities and elucidating the deeper underlying molecular mechanisms. We will use integrated approaches involving targeted gene silencing, complementation with synthetic (mutant) genes, activity profiling, and pathogen assays.