SKP1-SnRK protein kinase interactions mediate proteasomal binding of a plant SCF ubiquitin ligase.
Arabidopsis Snf1-related protein kinases (SnRKs) are implicated in pleiotropic regulation of metabolic, hormonal and stress responses through their interaction with the kinase inhibitor PRL1 WD-protein. Here we show that SKP1/ASK1, a conserved SCF (Skp1-cullin-F-box) ubiquitin ligase subunit, which suppresses the skp1-4 mitotic defect in yeast, interacts with the PRL1-binding C-terminal ... domains of SnRKs. The same SnRK domains recruit an SKP1/ASK1-binding proteasomal protein, alpha4/PAD1, which enhances the formation of a trimeric SnRK complex with SKP1/ASK1 in vitro. By contrast, PRL1 reduces the interaction of SKP1/ASK1 with SnRKs. SKP1/ASK1 is co-immunoprecipitated with a cullin SCF subunit (AtCUL1) and an SnRK kinase, but not with PRL1 from Arabidopsis cell extracts. SKP1/ASK1, cullin and proteasomal alpha-subunits show nuclear co-localization in differentiated Arabidopsis cells, and are observed in association with mitotic spindles and phragmoplasts during cell division. Detection of SnRK in purified 26S proteasomes and co-purification of epitope- tagged SKP1/ASK1 with SnRK, cullin and proteasomal alpha-subunits indicate that the observed protein interactions between SnRK, SKP1/ASK1 and alpha4/PAD1 are involved in proteasomal binding of an SCF ubiquitin ligase in Arabidopsis.
Mesh Terms:
Amino Acid Sequence, Arabidopsis, Arabidopsis Proteins, Binding Sites, Conserved Sequence, Models, Molecular, Molecular Sequence Data, Peptide Hydrolases, Peptide Synthases, Plant Proteins, Proteasome Endopeptidase Complex, Protein Binding, Protein Subunits, Protein-Serine-Threonine Kinases, Recombinant Proteins, Restriction Mapping, SKP Cullin F-Box Protein Ligases, Saccharomyces cerevisiae, Schizosaccharomyces pombe Proteins, Sequence Alignment, Sequence Homology, Amino Acid, Trans-Activators
Amino Acid Sequence, Arabidopsis, Arabidopsis Proteins, Binding Sites, Conserved Sequence, Models, Molecular, Molecular Sequence Data, Peptide Hydrolases, Peptide Synthases, Plant Proteins, Proteasome Endopeptidase Complex, Protein Binding, Protein Subunits, Protein-Serine-Threonine Kinases, Recombinant Proteins, Restriction Mapping, SKP Cullin F-Box Protein Ligases, Saccharomyces cerevisiae, Schizosaccharomyces pombe Proteins, Sequence Alignment, Sequence Homology, Amino Acid, Trans-Activators
EMBO J.
Date: Jun. 01, 2001
PubMed ID: 11387208
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