Auxin Response Factors as a model of transcriptional control

Salk Institute For Biological Studies, La Jolla CA

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Abstract

Project Summary Hormone-mediated modulation of gene activation or repression through transcription factors is central to all organisms. AUXIN RESPONSE FACTOR (ARF) transcription factors are critical modulators of plant growth and provide an ideal model for exploring hormone control of gene activation and repression. Of the 23-member ARF family, five are considered transcriptional activators and 18 are considered transcriptional repressors, allowing for study of both activator and repressor activities in a single family. Under low auxin concentrations, Aux/IAA proteins repress ARF transcription factors via direct interaction and recruitment of chromatin remodeling factors. When auxin concentrations are high, a co-receptor complex, comprised of an F-box protein from the TRANSPORT INHIBITOR REPONSE1 (TIR1) family and an Aux/IAA repressor protein, directly binds auxin. The F-box protein participates in a Skp1-Cullin-F-box (SCF) E3 ubiquitin ligase, which targets the Aux/IAA protein for degradation. This degradation event relieves ARF transcription factor repression, allowing auxin-regulated transcription. This receptor-ligand interaction allows a very short signal transduction chain to facilitate rapid transcriptional responses to auxin. To understand the molecular underpinnings of ARF-ARF and ARF-Aux/IAA interactions, our lab first solved the structure of the interaction domain, finding that it folds into a Type I/II PB1 domain. Within this domain, positively charged and negatively charged electrostatic faces occupy opposing sides, creating a miniature bar magnet that allows for front-to-back oligomerization in the packed crystal, in solution, and in the plant. Our lab further discovered that activity of a subset of ARFs is regulated by protein condensation driven by the combination of PB1 oligomerization and an intrinsically disordered region. Biomolecular condensation of these ARFs modulates auxin responsiveness in a developmentally-relevant context. We further found that many ARFs are regulated by proteasomal degradation, have identified an E3 ubiquitin ligase involved in this process, and determined that ARF stability is regulated by environmental conditions. Finally, ARF interactions are easily manipulated using PB1 domain point mutations, allowing us to direct ARF interactions for study. Using ARFs as a model will allow us to interrogate transcription factor function in an easily manipulated system to yield broad insight into many transcription factors. We are aided in our efforts by our multidisciplinary approach, extensive auxin-related molecular toolkit, and unique reagents generated by our lab. Our lab’s expertise in genetics and biochemistry/biophysics, combined with our discoveries of ARF condensation, nucleo-cytoplasmic partitioning, and proteasomal degradation, makes us well positioned to drive forward our understanding of biomolecular condensation and other mechanisms in regulation of transcription factor activity.