Abscisic acid increases Arabidopsis ABI5 transcription factor levels by promoting KEG E3 ligase self-ubiquitination and proteasomal degradation.
The Arabidopsis thaliana RING-type E3 ligase KEEP ON GOING (KEG) is a negative regulator of abscisic acid (ABA) signaling. Seedlings homozygous for T-DNA insertions in KEG accumulate high levels of the ABA-responsive transcription factor ABSCISIC ACID-INSENSITIVE5 (ABI5). Here, we demonstrate that KEG E3 ligase activity is required for the regulation of ABI5 abundance. KEG ubiquitinates ABI5 in vitro, and a functional KEG RING domain is required to restore the levels of ABI5 in keg-1 to that of the wild type. Overexpression of KEG leads to ABA insensitivity, which correlates with KEG protein levels. In the presence of ABA, ABI5 levels increase drastically via a decrease in ubiquitin-meditated proteasomal degradation. Our results indicate that ABA promotes ABI5 accumulation by inducing the ubiquitination and proteasomal degradation of KEG. A functional RING domain is required for the ABA-induced degradation of KEG, suggesting that the loss is due to self-ubiquitination. Mutations within KEG's kinase domain or treatments with kinase inhibitors prohibit the ABA-induced ubiquitination and degradation of KEG, indicating that phosphorylation, possibly self-phosphorylation, is involved in the ABA regulation of KEG protein levels. We discuss a model for how ABA may negatively regulate KEG protein abundance, leading to accumulation of ABI5 and ABA-dependent cellular responses.
Liu
Hongxia
H
Department of Biology, Dalhousie University, Halifax, Nova Scotia, Canada.
Stone
Sophia L
SL
D000040:Abscisic Acid / Q000494:pharmacology*; D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D050976:Basic-Leucine Zipper Transcription Factors / Q000235:genetics / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D030821:Plants, Genetically Modified / Q000201:enzymology / Q000235:genetics; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
C409743:ABI5 protein, Arabidopsis; D029681:Arabidopsis Proteins; D050976:Basic-Leucine Zipper Transcription Factors; D000040:Abscisic Acid; C518495:KEG protein, Arabidopsis; D044767:Ubiquitin-Protein Ligases; D046988:Proteasome Endopeptidase Complex