A super strong engineered auxin-TIR1 pair.
A super strong engineered auxin-TIR1 pair.
复制标题
超强工程生长素-TIR1 对。
DOI：
10.1093/pcp/pcy127
复制
发表时间：
2018
期刊：
影响因子：
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通讯作者：
Torii KU and Hagihara S
中科院分区：
文献类型：
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作者：
Yamada R;Murai K;Uchida N;Takahashi K;Iwasaki R;Tada Y;Kinoshita T;Itami K;Torii KU and Hagihara S
生长素通过诱导运输抑制剂反应1/生长素信号F-BOX蛋白（TIR 1/AFB）和生长素/吲哚乙酸（Aux/IAA）共受体蛋白之间的相互作用以及随后的转录调控来调节植物生长发育的各个方面。内源生长素信号的人工调控将使生长素介导的生物学事件的精确描述以及生长素的农业应用成为可能。为此，我们先前开发了一种合成的生长素受体对，其由5-（3-甲氧基苯基）-IAA（convexIAA，cvxIAA）和工程化的TIR 1组成，所述工程化的TIR 1在生长素结合口袋中的位置79处的苯丙氨酸被取代为甘氨酸（TIR 1F 79 G）（concaveTIR 1，ccvTIR 1）。这种合成的生长素受体对正交的天然生长素信号转导的转基因植物中窝藏的工程TIR 1通过外源性应用5-（3-甲氧基苯基）-IAA，并有可能被利用作为新的农业/园艺工具。在本研究中，我们报告了一个改进的版本的合成cvxIAA-ccvTIR 1对，使合成IAA可以在较低的浓度下发挥作用。利用酵母双杂交系统，我们筛选了各种5-取代的IAA，并鉴定了5-金刚烷基-IAA，命名为皮科_cvxIAA，其在比原始版本5-（3-甲氧基苯基）-IAA低1，000倍的浓度下介导TIR 1F 79 G和IAA 3蛋白的相互作用。此外，我们发现TIR 1F 79 A在皮摩尔浓度的5-金刚烷基-IAA存在下与IAA 3蛋白相互作用，比我们的cvxIAA-ccvTIR 1对的原型版本低10，000倍。此外，下拉试验证实，5-金刚烷基-IAA介导的TIR 1F 79 A和IAA 7-DII肽在体外相互作用在较低的浓度。具有高亲和力的改进的合成IAA-TIR 1对将有益于基础科学以及在农业/园艺中的实际应用。
Auxin regulates diverse aspects of plant growth and development through induction of the interaction between TRANSPORT INHIBITOR RESPONSE 1/AUXIN SIGNALING F-BOX proteins (TIR1/AFBs) and AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) co-receptor proteins and the subsequent transcriptional regulation. The artificial control of endogenous auxin signaling should enable the precise delineation of auxin-mediated biological events as well as the agricultural application of auxin. To this end, we previously developed a synthetic auxin–receptor pair that consists of 5-(3-methoxyphenyl)-IAA (convexIAA, cvxIAA) and the engineered TIR1 whose phenylalanine at position 79 in the auxin-binding pocket is substituted to glycine (TIR1F79G) (concaveTIR1, ccvTIR1). This synthetic auxin–receptor pair works orthogonally to natural auxin signaling in transgenic plants harboring the engineered TIR1 by exogenous application of 5-(3-methoxyphenyl)-IAA, and has potential to be utilized as novel agricultural/horticultural tools. In the present study, we report an improved version of the synthetic cvxIAA–ccvTIR1 pair such that synthetic IAA can act at lower concentrations. Using a yeast two-hybrid system, we screened various 5-substituted IAAs and identified 5-adamantyl-IAA, named pico_cvxIAA, which mediates interaction of TIR1F79Gand IAA3 proteins at a 1,000-fold lower concentration than the original version, 5-(3-methoxyphenyl)-IAA. Furthermore, we found that TIR1F79Ainteracts with IAA3 protein in the presence of picomolar concentrations of 5-adamantyl-IAA, 10,000-fold lower than our prototype version of the cvxIAA–ccvTIR1 pair. In addition, pull-down assays confirmed that 5-adamantyl-IAA mediates in vitro interaction of TIR1F79Aand IAA7-DII peptides at lower concentrations. The improved synthetic IAA–TIR1 pair with high affinity would be beneficial for basic science as well as for practical use in agriculture/horticulture.