Engineering plants to secrete affinity-tagged pathogen elicitors for deciphering immune receptor complex or inducing enhanced immunity
Engineering plants to secrete affinity-tagged pathogen elicitors for deciphering immune receptor complex or inducing enhanced immunity
复制标题
工程植物分泌亲和标记的病原体诱导子，用于破译免疫受体复合物或诱导增强的免疫力
DOI：
10.1111/jipb.12859
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发表时间：
2020-06-01
影响因子：
11.4
通讯作者：
Li, Jian-Feng
中科院分区：
文献类型：
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作者：
Miao, Shuang;Liu, Jiuer;Li, Jian-Feng
植物细胞内存在大量的模式识别受体（PRRs），用于检测潜在微生物病原体的微生物相关分子模式（MAMPs）。MAMPs由蛋白质模式过度代表，例如来自细菌鞭毛蛋白的flg 22肽。通过正向或反向遗传学鉴定PRR受体复合物组分可能是耗时/费力的，并且被功能冗余混淆。在这里，我们提出了一个策略，用于识别PRR复杂的组件工程植物诱导分泌亲和标记的蛋白质MAMPs的质外体。通过亲和纯化富集与自身分泌的MAMP结合的PRR蛋白复合物，并通过质谱分析进行分离。作为原理的证明，我们可以使用在雌二醇诱导下分泌FLAG标记的flg 22的拟南芥植物捕获flg 22受体FLS 2和共受体BAK 1。此外，我们确定了受体样激酶LIK 1和PEPR 1/PEPR 2作为FLS 2受体复合物中的潜在组分，并通过蛋白质-蛋白质相互作用测定和反向遗传学方法进一步验证。我们的研究展示了一种简单的方法来生化识别内源性PRR复合物组分，而不过度表达PRR或使用化学交联剂，并表明植物中不同免疫受体之间可能存在串扰。适度剂量的雌二醇也可用于诱导工程植物对细菌和真菌病原体的增强的免疫力。
Plant cells mount plenty of pattern-recognition receptors (PRRs) to detect the microbe-associated molecular patterns (MAMPs) from potential microbial pathogens. MAMPs are overrepresented by proteinaneous patterns, such as the flg22 peptide from bacterial flagellin. Identification of PRR receptor complex components by forward or reverse genetics can be time/labor-consuming, and be confounded by functional redundancies. Here, we present a strategy for identifying PRR complex components by engineering plants to inducibly secrete affinity-tagged proteinaneous MAMPs to the apoplast. The PRR protein complexes bound to self-secreted MAMPs are enriched through affinity purification and dissected by mass spectrometry. As a proof of principle, we could capture the flg22 receptor FLS2 and co-receptor BAK1 using Arabidopsis plants secreting FLAG-tagged flg22 under estradiol induction. Moreover, we identified receptor-like kinases LIK1 and PEPR1/PEPR2 as potential components in the FLS2 receptor complex, which were further validated by protein-protein interaction assays and the reverse genetics approach. Our study showcases a simple way to biochemically identify endogenous PRR complex components without overexpressing the PRR or using chemical cross-linkers, and suggests a possible crosstalk between different immune receptors in plants. A modest dose of estradiol can also be applied to inducing enhanced immunity in engineered plants to both bacterial and fungal pathogens.