Auxin-induced, SCF(TIR1)-mediated poly-ubiquitination marks AUX/IAA proteins for degradation.
The plant hormone auxin (indole-3-acetic acid or IAA) regulates plant development by inducing rapid cellular responses and changes in gene expression. Auxin promotes the degradation of Aux/IAA transcriptional repressors, thereby allowing auxin response factors (ARFs) to activate the transcription of auxin-responsive genes. Auxin enhances the binding of Aux/IAA proteins to the receptor TIR1, which is an F-box protein that is part of the E3 ubiquitin ligase complex SCF(TIR1). Binding of Aux/IAA proteins leads to degradation via the 26S proteasome, but evidence for SCF(TIR1)-mediated poly-ubiquitination of Aux/IAA proteins is lacking. Here we used an Arabidopsis cell suspension-based protoplast system to find evidence for SCF(TIR1)-mediated ubiquitination of the Aux/IAA proteins SHY2/IAA3 and BDL/IAA12. Each of these proteins showed a distinct abundance and repressor activity when expressed in this cell system. Moreover, the amount of endogenous TIR1 protein appeared to be rate-limiting for a proper auxin response measured by the co-transfected DR5::GUS reporter construct. Co-transfection with 35S::TIR1 led to auxin-dependent degradation, and excess of 35S::TIR1 even led to degradation of Aux/IAAs in the absence of auxin treatment. Expression of the mutant tir1-1 protein or the related F-box protein COI1, which is involved in jasmonate signaling, had no effect on Aux/IAA degradation. Our results show that SHY2/IAA3 and BDL/IAA12 are poly-ubiquitinated and degraded in response to increased auxin or TIR1 levels. In conclusion, our data provide experimental support for the model that SCF(TIR1)-dependent poly-ubiquitination of Aux/IAA proteins marks these proteins for degradation by the 26S proteasome, leading to activation of auxin-responsive gene expression.
Maraschin
Felipe dos Santos
Fdos S
Institute of Biology, Leiden University, Clusius Laboratorium, Wassenaarseweg 64, 2333 AL Leiden, The Netherlands.
Memelink
Johan
J
Offringa
Remko
R
D017360:Arabidopsis / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D044783:F-Box Proteins / Q000235:genetics / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D007210:Indoleacetic Acids / Q000494:pharmacology*; D009687:Nuclear Proteins / Q000378:metabolism; D010937:Plant Growth Regulators / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D012097:Repressor Proteins / Q000378:metabolism; D054875:Ubiquitination
D029681:Arabidopsis Proteins; C112279:COI1 protein, Arabidopsis; D044783:F-Box Proteins; C527433:IAA12 protein, Arabidopsis; D007210:Indoleacetic Acids; D009687:Nuclear Proteins; D010937:Plant Growth Regulators; D011956:Receptors, Cell Surface; D012097:Repressor Proteins; C117994:SHY2 protein, Arabidopsis; C501405:TIR1 protein, Arabidopsis