Natural variation reveals key amino acids in a downy mildew effector that alters recognition specificity by an Arabidopsis resistance gene
Natural variation reveals key amino acids in a downy mildew effector that alters recognition specificity by an Arabidopsis resistance gene
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DOI：
10.1111/j.1364-3703.2008.00481.x
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发表时间：
2008-07-01
影响因子：
4.9
通讯作者：
Beynon, Jim L.
中科院分区：
文献类型：
--
作者：
Allen, Rebecca L.;Meitz, Julia C.;Beynon, Jim L.
RPP 13是细胞质类抗病基因的成员，编码迄今为止鉴定的最可变的拟南芥蛋白之一。这种变异性在ATR 13中是匹配的，ATR 13是来自卵菌霜霉病病原体Hyaloperonospora parasitica的蛋白质，由RPP 13识别，这表明这些蛋白质参与了紧密的互惠共同进化。ATR 13具有5个结构域：N-末端信号肽、RXLR基序、七肽亮氨酸/异亮氨酸重复序列、11个氨基酸重复序列和C-末端结构域。我们发现，保守的RXLR-含有域的ATR 13介导的识别，其在运输到植物细胞质中的作用是一致的。对16株H.寄生虫揭示了整个蛋白质的高水平的氨基酸多样性。七肽亮氨酸重复序列的亮氨酸/异亮氨酸是保守的，特定亮氨酸或异亮氨酸残基的突变改变了RPP 13的识别。自然变异并没有利用这一途径来避免检测，这表明该结构域在致病性中的关键作用。11个氨基酸重复单位的广泛变异不影响RPP 13的识别。结构域交换分析表明，识别特异性在于ATR 13的C-末端结构域。变异分析与功能测定相结合，允许识别的四个氨基酸的位置，可能发挥作用的识别特异性。定点诱变证实，苏氨酸残基是绝对需要的RPP 13的识别和识别可以通过在其他网站的精氨酸或谷氨酸的存在下调制。这三个氨基酸的突变对ATR 13与RPP 13不相关的抗性基因的相互作用没有影响，这与它们在确定RPP 13-Nd识别特异性中的关键作用一致。
RPP13, a member of the cytoplasmic class of disease resistance genes, encodes one of the most variable Arabidopsis proteins so far identified. This variability is matched in ATR13, the protein from the oomycete downy mildew pathogen Hyaloperonospora parasitica recognized by RPP13, suggesting that these proteins are involved in tight reciprocal coevolution. ATR13 exhibits five domains: an N-terminal signal peptide, an RXLR motif, a heptad leucine/isoleucine repeat, an 11-amino-acid repeated sequence and a C-terminal domain. We show that the conserved RXLR-containing domain is dispensable for ATR13-mediated recognition, consistent with its role in transport into the plant cytoplasm. Sequencing ATR13 from 16 isolates of H. parasitica revealed high levels of amino acid diversity across the entire protein. The leucines/isoleucines of the heptad leucine repeat were conserved, and mutation of particular leucine or isoleucine residues altered recognition by RPP13. Natural variation has not exploited this route to detection avoidance, suggesting a key role of this domain in pathogenicity. The extensive variation in the 11-amino-acid repeat units did not affect RPP13 recognition. Domain swap analysis showed that recognition specificity lay in the C-terminal domain of ATR13. Variation analyses combined with functional assays allowed the identification of four amino acid positions that may play a role in recognition specificity. Site-directed mutagenesis confirmed that a threonine residue is absolutely required for RPP13 recognition and that recognition can be modulated by the presence of either an arginine or glutamic acid at other sites. Mutations in these three amino acids had no effect on the interaction of ATR13 with a resistance gene unlinked to RPP13, consistent with their critical role in determining RPP13-Nd recognition specificity.