Distinct Amino Acids of the Phytophthora infestans Effector AVR3a Condition Activation of R3a Hypersensitivity and Suppression of Cell Death
Distinct Amino Acids of the Phytophthora infestans Effector AVR3a Condition Activation of R3a Hypersensitivity and Suppression of Cell Death
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DOI：
10.1094/mpmi-22-3-0269
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发表时间：
2009-03-01
影响因子：
3.5
通讯作者：
Kamoun, Sophien
中科院分区：
文献类型：
--
作者：
Bos, Jorunn I. B.;Chaparro-Garcia, Angela;Kamoun, Sophien
疫霉菌AVR3a蛋白是RXLR类细胞质效应蛋白的多态性成员，具有双重功能。AVR3a（KI）而不是AVR3a（EM）激活马铃薯抗性蛋白R3a触发的先天免疫，并且是由INF1剔除素诱导的细胞死亡反应的强抑制因子，INF1剔除素是一种分泌的P. infestans蛋白，具有病原体相关分子模式的特征。为了深入了解AVR3a活性的分子基础，我们对两种AVR3a形式进行了结构-功能分析。我们利用饱和高通量突变体筛选来鉴定对R3a激活重要的氨基酸。在6500个AVR3a（EM）克隆中，我们鉴定出136个AVR3a（EM）突变克隆获得了诱导R3a超敏反应的能力。这组突变克隆中有15个氨基酸位点受到影响。这些突变体中的大多数对细胞死亡的抑制程度与AVR3a（KI）相似。对4500个AVR3a（KI）克隆进行类似的功能丧失筛选，只发现了13个活性改变的突变体。这些结果表明，AVR3a通过与一种或多种宿主蛋白相互作用发挥作用，与通过酶活性识别AVR3a不一致。获得R3a激活但没有细胞死亡抑制活性的突变体的鉴定表明，不同的氨基酸决定了两种AVR3a效应物的活性。
The AVR3a protein of Phytophthora infestans is a polymorphic member of the RXLR class of cytoplasmic effectors with dual functions. AVR3a(KI) but not AVR3a(EM) activates innate immunity triggered by the potato resistance protein R3a and is a strong suppressor of the cell-death response induced by INF1 elicitin, a secreted P. infestans protein that has features of pathogen-associated molecular patterns. To gain insights into the molecular basis of AVR3a activities, we performed structure-function analyses of both AVR3a forms. We utilized saturated high-throughput mutant screens to identify amino acids important for R3a activation. Of 6,500 AVR3a(EM) clones tested, we identified 136 AVR3a(EM) mutant clones that gained the ability to induce R3a hypersensitivity. Fifteen amino-acid sites were affected in this set of mutant clones. Most of these mutants did not suppress cell death at a level similar to that of AVR3a(KI). A similar loss-of-function screen of 4,500 AVR3a(KI) clones identified only 13 mutants with altered activity. These results point to models in which AVR3a functions by interacting with one or more host proteins and are not consistent with the recognition of AVR3a through an enzymatic activity. The identification of mutants that gain R3a activation but not cell-death suppression activity suggests that distinct amino acids condition the two AVR3a effector activities.