A fuzzy encounter complex precedes formation of the fully-engaged TIR1-Aux/IAA auxin co-receptor system
A fuzzy encounter complex precedes formation of the fully-engaged TIR1-Aux/IAA auxin co-receptor system
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DOI：
10.1101/781922
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发表时间：
2019-09
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通讯作者：
Sigurd Ramans Harborough;A. Kalverda;G. Thompson;M. Kieffer;M. Kubeš;M. Quareshy;V. Uzunova;J. Prusińska;Ken-ichiro Hayashi;R. Napier;I. Manfield;S. Kepinski
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作者：
Sigurd Ramans Harborough;A. Kalverda;G. Thompson;M. Kieffer;M. Kubeš;M. Quareshy;V. Uzunova;J. Prusińska;Ken-ichiro Hayashi;R. Napier;I. Manfield;S. Kepinski
植物激素生长素通过TIR 1/AFB-auxin-Aux/IAA auxin共受体复合物调节植物发育的几乎所有方面。在这种三元复合物中，生长素作为分子胶促进Aux/IAA转录抑制蛋白与SCFTIR 1/AFB泛素连接酶复合物的结合，从而催化它们的泛素介导的蛋白水解。该配合物的晶体结构的一个显著特征是在Aux/IAA降解决定子基序内存在罕见的顺式W-P键。为了研究受体复合物的组装，我们已经使用NMR来确定溶液结构的氨基末端的一半的Aux/IAA蛋白AXR 3/IAA 17，包括降解决定子，无论是在隔离和在复杂的TIR 1和生长素。我们发现，AXR 3的这个区域本质上是无序的，只有有限的结构元素，但临界降解决定子W-P键发生异常高（1：1）的顺反异构体的比例。我们发现，组装的辅助受体复合物涉及生长素依赖和独立的相互作用事件，其中的Aux/IAA的障碍被保留。此外，使用合成的生长素分子cvxIAA和通过分析特定的Aux/IAA构象，我们表明，生长素依赖的结合事件的一个子集发生远离典型的生长素结合口袋中的TIR 1的基础。我们的研究结果揭示了一个模糊的，拓扑不同的三元遇到复杂的存在，因此，生长素的感知并不限于顺序，独立的结合生长素，然后辅助/IAA到TIR 1。
The plant hormone auxin regulates almost every aspect of plant development via the TIR1/AFB-auxin-Aux/IAA auxin co-receptor complex. Within this ternary complex, auxin acts as a molecular glue to promote the binding of Aux/IAA transcriptional repressor proteins to SCFTIR1/AFB ubiquitin-ligase complexes, thereby catalysing their ubiquitin-mediated proteolysis. A conspicuous feature of the crystal structure of the complex is a rare cis W-P bond within the Aux/IAA degron motif. To study receptor complex assembly, we have used NMR to determine the solution structure of the amino-terminal half of the Aux/IAA protein AXR3/IAA17, including the degron, both in isolation and in complex with TIR1 and auxin. We show that this region of AXR3 is intrinsically-disordered with only limited elements of structure and yet the critical degron W-P bond occurs with an unusually high (1:1) ratio of cis to trans isomers. We show that assembly of the co-receptor complex involves both auxin-dependent and -independent interaction events in which the disorder of the Aux/IAA is retained. Further, using the synthetic auxin molecule cvxIAA and by analysing specific Aux/IAA conformers, we show that a subset of auxin-dependent binding events occur away from the base of the canonical auxin binding pocket in TIR1. Our results reveal the existence of a fuzzy, topologically-distinct ternary encounter complex and thus that auxin perception is not limited to sequential, independent binding of auxin and then Aux/IAA to TIR1.