Real-time monitoring of translocation of selected type-III effectors from Xanthomonas oryzae pv. oryzae into rice cells
Real-time monitoring of translocation of selected type-III effectors from Xanthomonas oryzae pv. oryzae into rice cells
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实时监测米黄单胞菌中选定的 III 型效应子的易位。
DOI：
10.1007/s12038-019-9916-0
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发表时间：
2019-08-02
影响因子：
2.9
通讯作者：
Dong, Hansong
中科院分区：
文献类型：
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作者：
Bian, Huijie;Zhang, Liyuan;Dong, Hansong
黄单胞菌黄单胞菌致病型的III型（T3）效应子PthXo 1和AvrXa 10转座到水稻细胞中，诱导感病和抗病水稻品种日本晴和IRBB 10的毒力和无毒力。这种转运需要细菌T3转运蛋白Hpa 1和水稻质膜蛋白OsPIP 1的参与;在这里，我们使用β-内酰胺酶（BlaM）报告系统来观察PthXo 1和AvrXa 10易位。建立了通过定量BlaM水解产物[P]和底物的荧光共振能量转移（FRET）来监测动物病原菌效应子的系统。通过三个标准评估了BlaM报告基因在水稻原生质体中的可行性。第一个标准表示日本晴和IRBB 10的野生型和OsPIP 1;3过表达和OsPIP 1;3沉默株系之间的[P]和FRET水平之间的差异。第二个标准表明在存在和不存在Hpa 1的情况下[P]和FRET水平之间的差异。最后一个标准阐明了PthXo 1易位与日本晴原生质体中PthXo 1靶基因的诱导表达的一致性以及AvrXa 10易位与IRBB 10原生质体中AvrXa 10靶基因的诱导表达的一致性。这些结果提供了一个实验途径，实时监测细菌T3效应易位到植物细胞的病理后果。
Type-III (T3) effectors PthXo1 and AvrXa10 of Xanthomonas oryzae pv. oryzae are translocated into rice cells to induce virulence and avirulence on susceptible- and resistant-rice varieties Nipponbare and IRBB10, respectively. The translocation needs the bacterial T3 translocator Hpa1 and rice Oryza sativa plasma membrane protein OsPIP1;3. Here, we employed the β-lactamase (BlaM) reporter system to observe PthXo1 and AvrXa10 translocation. The system was established to monitor effectors of animal-pathogenic bacteria by quantifying the BlaM hydrolysis product [P] and fluorescence resonance energy transfer (FRET) of the substrate. The feasibility of the BlaM reporter in rice protoplasts was evaluated by three criteria. The first criterion indicated differences between both [P] and FRET levels among wild types and OsPIP1;3-overexpressing and OsPIP1;3-silenced lines of both Nipponbare and IRBB10. The second criterion indicated differences between [P] and FRET levels in the presence and absence of Hpa1. The last criterion elucidated the coincidence of PthXo1 translocation with induced expression of the PthXo1 target gene in protoplasts of Nipponbare and the coincidence of AvrXa10 translocation with induced expression of the AvrXa10 target gene in protoplasts of IRBB10. These results provide an experimental avenue for real-time monitoring of bacterial T3 effector translocation into plant cells with a pathological consequence.