A Super Strong Engineered Auxin–TIR1 Pair

• Published in: Plant and cell physiology Vol. 59; no. 8; pp. 1538 - 1544
• Main Authors: Yamada, Ryotaro, Murai, Keiichiro, Uchida, Naoyuki, Takahashi, Koji, Iwasaki, Rie, Tada, Yasuomi, Kinoshita, Toshinori, Itami, Kenichiro, Torii, Keiko U, Hagihara, Shinya
• Format: Journal Article
• Language: English
• Published: Japan Oxford University Press 01.08.2018
• Subjects: Arabidopsis Proteins - metabolism; F-Box Proteins - metabolism; Indoleacetic Acids - metabolism; Nuclear Proteins - metabolism; Protein Binding; Receptors, Cell Surface - metabolism; Special Focus Issue – Regular Papers; Bump-and-hole; Auxin; TIR1
• ISSN: 0032-0781; 1471-9053
• DOI: 10.1093/pcp/pcy127

Abstract 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 TIR1F79G and IAA3 proteins at a 1,000-fold lower concentration than the original version, 5-(3-methoxyphenyl)-IAA. Furthermore, we found that TIR1F79A interacts 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 TIR1F79A and 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.