Mechanism of Auxin Action

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DESCRIPTION (provided by applicant): The plant hormone auxin is involved in many, perhaps all, aspects of plant growth and development. Auxin acts by stimulating the degradation of a family of transcriptional repressors called the Aux/IAA proteins, a process that requires the ubiquitin protein ligase (E3) SCFTIR1. During the last grant period we showed that auxin binds directly to the F-box protein TIR1, the substrate binding subunit in an E3 called SCFTIR1. Auxin binding stabilizes the interaction between SCFTIR1 and the Aux/IAA proteins. This is a novel mechanism of both hormone perception and E3 regulation. In addition we showed that TIR1 binds an InsP6 molecule and suggest that InsP6 may be required for TIR1 structure and/or function. Further we demonstrated that TIR1 is one of 6 related auxin sensor F-box proteins, collectively called the AFBs. The current proposal has five specific aims. The first is to investigate the molecular mechanism of auxin perception by TIR1. Based on our previous work, we propose that auxin acts like molecular glue to stabilize the TIR1- Aux/IAA complex. Biochemical studies will be performed to test various aspects of this model. In addition, we will determine the importance of InsP6 for TIR1 function. The second Aim is to characterize the biochemical diversity of the AFB proteins. A series of in vitro studies will be performed to characterize auxin and Aux/IAA binding to each member of the family. The significance of any specific interactions will be determined in vivo. The third Aim is to identify and characterize protein complexes containing TIR1 and/or the Aux/IAA proteins IAA28 with the goal of obtaining further information on the regulation of TIR1. In addition, this aim will provide additional information on the specificity of TIR1 and Aux/IAA interactions and the cellular location of TIR1-Aux/IAA complex formation. Aim four is to determine the function of cyclophilin molecules in auxin action. The fifth Aim is to characterize the auxin transcriptional network with a focus on two transcription factors called ARF5 and ARF7. These studies address a number of key issues in cellular regulation and will have important implications for human health. The ubiquitin pathway and SCF E3s in particular, are involved in diverse disease processes including numerous cancers. Because SCFTIR1 is one of the best-characterized E3 complexes in any species, this work provides a unique opportunity to advance our understanding of this critical aspect of human disease PUBLIC HEALTH RELEVANCE: Protein degradation by the ubiquitin/proteasome pathway is a central aspect of cellular regulation. Defects in the pathway contribute to many disease processes including cancers. This study will advance our understanding of the ubiquitin/proteasome pathway in cell function.