14-3-3 proteins activate a plant calcium-dependent protein kinase (CDPK).
Plants and protozoa contain a unique family of calcium-dependent protein kinases (CDPKs) which are defined by the presence of a carboxyl-terminal calmodulin-like regulatory domain. We present biochemical evidence indicating that at least one member of this kinase family can be stimulated by 14-3-3 proteins. Isoform CPK-1 from the model plant Arabidopsis thaliana was expressed as a fusion protein in E. coli and purified. The calcium-dependent activity of this recombinant CPK-1 was shown to be stimulated almost twofold by three different 14-3-3 isoforms with 50% activation around 200 nM. 14-3-3 proteins bound to the purified CPK-1, as shown by binding assays in which either the 14-3-3 or CPK-1 were immobilized on a matrix. Both the 14-3-3 binding and activation of CPK-1 were specifically disrupted by a known 14-3-3 binding peptide LSQRQRSTpSTPNVHMV (IC50 = 30 microM). These results raise the question of whether 14-3-3 can modulate the activity of CDPK signal transduction pathways in plants.
Institute of Molecular Biology, University of Copenhagen, Copenhagen K, Denmark.
L
L
Camoni
J F
JF
Harper
M G
MG
Palmgren
D048948:14-3-3 Proteins; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology*; D029681:Arabidopsis Proteins; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D004789:Enzyme Activation; D004926:Escherichia coli / Q000235:genetics; D007527:Isoenzymes / Q000378:metabolism; D008969:Molecular Sequence Data; D010455:Peptides; D010940:Plant Proteins; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism*; D011506:Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D011993:Recombinant Fusion Proteins; D014446:Tyrosine 3-Monooxygenase; D017871:Calcium-Calmodulin-Dependent Protein Kinases
D048948:14-3-3 Proteins; D029681:Arabidopsis Proteins; D002135:Calcium-Binding Proteins; D007527:Isoenzymes; D010455:Peptides; D010940:Plant Proteins; D011494:Protein Kinases; D011506:Proteins; D019908:Proto-Oncogene Proteins c-raf; D011993:Recombinant Fusion Proteins; D014446:Tyrosine 3-Monooxygenase; C490306:ATCDPK1 protein, Arabidopsis; D017871:Calcium-Calmodulin-Dependent Protein Kinases