Altering Specificity and Autoactivity of Plant Immune Receptors Sr33 and Sr50 Via a Rational Engineering Approach.
Altering Specificity and Autoactivity of Plant Immune Receptors Sr33 and Sr50 Via a Rational Engineering Approach.
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DOI：
10.1094/mpmi-07-22-0154-r
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发表时间：
2023-07
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作物中许多抗病基因是胞内核苷酸结合(NB)富含亮氨酸重复(LRR)受体(NLR)。合理设计NLRs特异性的能力将在应对新出现的作物疾病方面至关重要。修改NLR识别的成功尝试一直局限于非靶向的方法，或者依赖于先前可用的结构信息或对病原体-效应物靶标的了解。但是，对于大多数NLR效应器对，此信息不可用。在这里，我们展示了在两个密切相关的NLR之间精确预测和随后转移效应器识别所涉及的残基，而不需要通过实验确定它们的结构或详细了解它们的病原体效应器靶标。通过结合系统发育、等位基因多样性分析和结构建模，我们成功地预测了介导Sr50与其同源效应子AvrSr50相互作用的残基，并将Sr50的识别特异性转移到了密切相关的NLR Sr33。我们创造了含有来自Sr50的氨基酸的合成版本的Sr33，包括Sr33syn，它获得了识别具有12个氨基酸替换的AvrSr50的能力。此外，我们发现LRR结构域中需要将识别特异性转移到Sr33的位点也影响Sr50的自身活性。结构模拟表明，这些残基与NB-ARC结构域的一部分相互作用，我们称之为NB-ARC闩锁，可能维持受体的非活性状态。我们的方法展示了NLRs的合理修改，这可能有助于增强现有的优秀作物种质。
Many resistance genes deployed against pathogens in crops are intracellular nucleotide-binding (NB) leucine-rich repeat (LRR) receptors (NLRs). The ability to rationally engineer the specificity of NLRs will be crucial in the response to newly emerging crop diseases. Successful attempts to modify NLR recognition have been limited to untargeted approaches or depended on previously available structural information or knowledge of pathogen-effector targets. However, this information is not available for most NLR-effector pairs. Here, we demonstrate the precise prediction and subsequent transfer of residues involved in effector recognition between two closely related NLRs without their experimentally determined structure or detailed knowledge about their pathogen effector targets. By combining phylogenetics, allele diversity analysis, and structural modeling, we successfully predicted residues mediating interaction of Sr50 with its cognate effector AvrSr50 and transferred recognition specificity of Sr50 to the closely related NLR Sr33. We created synthetic versions of Sr33 that contain amino acids from Sr50, including Sr33syn, which gained the ability to recognize AvrSr50 with 12 amino-acid substitutions. Furthermore, we discovered that sites in the LRR domain needed to transfer recognition specificity to Sr33 also influence autoactivity in Sr50. Structural modeling suggests these residues interact with a part of the NB-ARC domain, which we named the NB-ARC latch, to possibly maintain the inactive state of the receptor. Our approach demonstrates rational modifications of NLRs, which could be useful to enhance existing elite crop germplasm.