Auxin (Indole-3-acetic acid; IAA) can be considered one of the most important hormones in plant development as it coordinates plant development through transcriptional regulation.1 Auxin is shown to bind to an F-box protein Transport Inhibitor Response 1 (TIR1) 2 of which there are 5 other homologues; Auxin Signalling F-Box (AFB1 – 5). TIR1 and AFB5 are the most distantly related in terms of sequence homology and will be studied for this project.
At low auxin levels, Aux/IAA transcriptional repressors, together with co-repressor proteins repress genes targeted by auxin response factor (ARF) transcriptional activators. F-box proteins determine substrate specificity of Skp1/Cullin/F-box (SCF) complexes that ubiquitinate target proteins, marking them as substrates for proteasomal degradation. At higher levels of auxin; once auxin binds to TIR1 the Aux/IAA protein complexes with TIR1 via a binding domain in which auxin is the “molecular glue” between the two. Thereafter, TIR1 tethers to an SCF complex which ubiquitinates the Aux/IAA protein and subsequently the Aux/IAA is degraded by 26S proteasomes. Hence the ARF’s are no longer repressed and transcription commences. 3
As yet there is no comprehensive understanding of how and why complexes like the SCF recruit their components and what drives their combinations and partnering. 4,5 The aims of this project are to observe the binding of Auxin to the TIR1/AFB5 receptors and elucidate the events that lead up to the complex formation with the Aux/IAA proteins. Surface plasmon resonance (SPR) and Isothermal titration calorimetry (ITC) will enable me to obtain kinetic and thermodynamic data respectively. We also aim to find novel compounds that are selective towards a particular member of the TIR1/AFB family with AUX/IAA proteins. This can be achieved by using in silico docking and ligand chemometrics to gain insights into novel functional groups and compound scaffolds for the screening process.
1. Westfall, C. S., Muehler, A. M. & Jez, J. M. Enzyme Action in the Regulation of Plant Hormone Responses. J. Biol. Chem. 288, 19304–19311 (2013).
2. Kepinski, S. & Leyser, O. The Arabidopsis F-box protein TIR1 is an auxin receptor. Nature 435, 446–51 (2005).
3. Woodward, A. W. & Bartel, B. A receptor for auxin. Plant Cell 17, 2425–9 (2005).
4. Hayashi, K. The interaction and integration of auxin signaling components. Plant Cell Physiol. 53, 965–75 (2012).
5. Sauer, M., Robert, S. & Kleine-Vehn, J. Auxin: simply complicated. J. Exp. Bot. 64, 2565–77 (2013).