Open AccessPhosphoproteomic identification of targets of the Arabidopsis sucrose nonfermenting-like kinase SnRK2.8 reveals a connection to metabolic processes
Author(s) -
Ryoung Shin,
Sophie Alvarez,
Adrien Y. Burch,
Joseph M. Jez,
Daniel P. Schachtman
Publication year - 2007
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.0610208104
Subject(s) - biochemistry , protein kinase a , map2k7 , biology , cyclin dependent kinase 2 , mitogen activated protein kinase kinase , protein phosphorylation , kinase , arabidopsis , map kinase kinase kinase , cyclin dependent kinase complex , microbiology and biotechnology , gene , mutant
SnRK2.8 is a member of the sucrose nonfermenting-related kinase family that is down-regulated when plants are deprived of nutrients and growth is reduced. When this kinase is over expressed inArabidopsis , the plants grow larger. To understand how this kinase modulates growth, we identified some of the proteins that are phosphorylated by this kinase. A new phosphoproteomic method was used in which total protein from plants overexpressing the kinase was compared with total protein from plants in which the kinase was inactivated. Protein profiles were compared on two-dimensional gels following staining by a dye that recognizes phosphorylated amino acids. Candidate target proteins were confirmed within vitro phosphorylation assays, using the kinase and target proteins that were purified fromEscherichia coli . Seven target proteins were confirmed as being phosphorylated by SnRK2.8. Certain targets, such as 14-3-3 proteins, regulate as yet unidentified proteins, whereas other targets, such as glyoxalase I and ribose 5-phosphate isomerase, detoxify byproducts from glycolysis and catalyze one of the final steps in carbon fixation, respectively. Also, adenosine kinase and 60S ribosomal protein were confirmed as targets of SnRK2.8. Using mass spectrometry, we identified phosphorylated residues in the SnRK2.8, the 14-3-3κ, and the 14-3-3χ. These data show that the expression ofSnRK2.8 is correlated with plant growth, which may in part be due to the phosphorylation of enzymes involved in metabolic processes.