Beyond glycolysis: GAPDHs are multi-functional enzymes involved in regulation of ROS, autophagy, and plant immune responses.
Beyond glycolysis: GAPDHs are multi-functional enzymes involved in regulation of ROS, autophagy, and plant immune responses.
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DOI：
10.1371/journal.pgen.1005199
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发表时间：
2015-04
期刊：
影响因子：
4.5
通讯作者：
Coaker G
中科院分区：
文献类型：
--
作者：
Henry E;Fung N;Liu J;Drakakaki G;Coaker G
甘油醛-3-磷酸脱氢酶（GAPDH）是脊椎动物能量代谢中的重要酶，具有多种细胞调节作用，但植物GAPDH异构体除了参与糖酵解外的重要性研究较少。虽然动物具有一种GAPDH同种型，但植物具有多种同种型。本研究采用细胞生物学和遗传学方法研究GAPDH在植物免疫应答中的作用。单个拟南芥GAPDH敲除（KO系）在接种细菌性植物病原体假单胞菌pv.番茄KO系表现出加速的程序性细胞死亡和增加的电解质渗漏响应效应触发的免疫。此外，KO系显示增加的基础ROS积累，如使用荧光探针H2 DCFDA可视化的。gapa 1 -2和gapc 1科斯在缺乏营养饥饿的情况下表现出组成性自噬表型。由于脊椎动物和植物胞质GAPDH之间的高度序列保守性，我们的实验集中在使用互补的GAPC 1-GFP系的胞质GAPC 1细胞动力学。与内吞膜标记物（FM 4 -64）和内体运输抑制剂（BFA，Wortmannin）结合的共聚焦成像证明，除了胞质溶胶和细胞核之外，胞质GAPC 1还定位于质膜和内膜系统。感知细菌鞭毛蛋白后，GAPC 1动态响应，荧光斑点大小显着增加，核积累增强。总之，这些结果表明，植物GAPDH可以影响植物免疫的多个方面，在不同的亚细胞区室。植物可以被所有病原体种类感染，严重影响作物生产和粮食安全。天然免疫反应对植物生存至关重要，但必须严格调控，以避免对生长和发育产生负面影响。在这里，我们研究的作用，甘油醛3-磷酸脱氢酶（GAPDH）蛋白在模式植物拟南芥，芥菜的亲戚。动物有一种GAPDH同种型，已被深入研究，并显示出多种兼职或非传统活动。植物具有存在于不同亚细胞区室中的多种GAPDH同种型。使用特定GAPDH敲除的遗传学研究与显微镜相结合，我们发现GAPDH调节活性氧的积累和细胞死亡，以响应细菌病原体假单胞菌的接种。GAPC 1亚型表现出不同的亚细胞定位和动态响应细菌鞭毛蛋白的感知。GAPC 1和GAPA 1同种型也负调节自噬，这是植物免疫应答的重要组成部分。综上所述，我们的结果表明，多种GAPDH异构体起负调控植物防御反应的作用。负调节因子对于精确调节免疫应答的持续时间和幅度是重要的。
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is an important enzyme in energy metabolism with diverse cellular regulatory roles in vertebrates, but few reports have investigated the importance of plant GAPDH isoforms outside of their role in glycolysis. While animals possess one GAPDH isoform, plants possess multiple isoforms. In this study, cell biological and genetic approaches were used to investigate the role of GAPDHs during plant immune responses. Individual Arabidopsis GAPDH knockouts (KO lines) exhibited enhanced disease resistance phenotypes upon inoculation with the bacterial plant pathogen Pseudomonas syringae pv. tomato. KO lines exhibited accelerated programmed cell death and increased electrolyte leakage in response to effector triggered immunity. Furthermore, KO lines displayed increased basal ROS accumulation as visualized using the fluorescent probe H2DCFDA. The gapa1-2 and gapc1 KOs exhibited constitutive autophagy phenotypes in the absence of nutrient starvation. Due to the high sequence conservation between vertebrate and plant cytosolic GAPDH, our experiments focused on cytosolic GAPC1 cellular dynamics using a complemented GAPC1-GFP line. Confocal imaging coupled with an endocytic membrane marker (FM4-64) and endosomal trafficking inhibitors (BFA, Wortmannin) demonstrated cytosolic GAPC1 is localized to the plasma membrane and the endomembrane system, in addition to the cytosol and nucleus. After perception of bacterial flagellin, GAPC1 dynamically responded with a significant increase in size of fluorescent puncta and enhanced nuclear accumulation. Taken together, these results indicate that plant GAPDHs can affect multiple aspects of plant immunity in diverse sub-cellular compartments. Plants can be infected by all pathogen classes, significantly impacting crop production and food security. Innate immune responses are critical to plant survival but must be tightly regulated in order to avoid negative impacts on growth and development. Here, we investigated the role of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) proteins in the model plant Arabidopsis thaliana, a mustard relative. Animals have one GAPDH isoform, which has been intensely investigated and shown to exhibit diverse moonlighting, or non-traditional, activities. Plants possess multiple GAPDH isoforms that reside in distinct sub-cellular compartments. Using a combination of genetic investigation of specific GAPDH knockouts coupled with microscopy, we found that GAPDHs regulate accumulation of reactive oxygen species and cell death in response to inoculation with the bacterial pathogen Pseudomonas syringae. The GAPC1 isoform exhibits diverse sub-cellular localizations and dynamically responds to perception of bacterial flagellin. The GAPC1 and GAPA1 isoforms also negatively regulate autophagy, which is an important component of plant immune responses. Taken together, our results demonstrate that multiple GAPDH isoforms act to negatively regulate plant defense responses. Negative regulators are important for precisely regulating the duration and amplitude of immune responses.
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