Regulation of the Dual Specificity Protein Phosphatase, DsPTP1, through Interactions with Calmodulin

• Published in: The Journal of biological chemistry Vol. 279; no. 2; pp. 848 - 858
• Main Authors: Yoo, Jae Hyuk, Cheong, Mi Sun, Park, Chan Young, Moon, Byeong Cheol, Kim, Min Chul, Kang, Yun Hwan, Park, Hyeong Cheol, Choi, Man Soo, Lee, Ju Huck, Jung, Won Yong, Yoon, Hae Won, Chung, Woo Sik, Lim, Chae Oh, Lee, Sang Yeol, Cho, Moo Je
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 09.01.2004; American Society for Biochemistry and Molecular Biology
• Subjects: Amino Acid Motifs; Amino Acid Sequence; Aniline Compounds - chemistry; Arabidopsis; Arabidopsis Proteins - biosynthesis; Arabidopsis Proteins - chemistry; Binding Sites; Binding, Competitive; Blotting, Western; Calcium - chemistry; Calcium - metabolism; Calmodulin - chemistry; Calmodulin - metabolism; DNA, Complementary - metabolism; Dose-Response Relationship, Drug; Dual-Specificity Phosphatases; Gene Deletion; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Plant; Gene Library; Glutathione Transferase - metabolism; Kinetics; Models, Genetic; Molecular Sequence Data; Mutagenesis, Site-Directed; Mutation; Organophosphorus Compounds - chemistry; Peptides - chemistry; Phosphoric Diester Hydrolases - metabolism; Phosphorylation; Phosphotyrosine - chemistry; Protein Binding; Protein Phosphatase 1; Protein Structure, Tertiary; Protein Tyrosine Phosphatases - biosynthesis; Protein Tyrosine Phosphatases - chemistry; Protein Tyrosine Phosphatases - genetics; Saccharomyces cerevisiae - metabolism; Signal Transduction
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M310709200

Reversible phosphorylation is a key mechanism for the control of intercellular events in eukaryotic cells. In animal cells, Ca2+/CaM-dependent protein phosphorylation and dephosphorylation are implicated in the regulation of a number of cellular processes. However, little is known on the functions of Ca2+/CaM-dependent protein kinases and phosphatases in Ca2+ signaling in plants. From an Arabidopsis expression library, we isolated cDNA encoding a dual specificity protein phosphatase 1, which is capable of hydrolyzing both phosphoserine/threonine and phosphotyrosine residues of the substrates. Using a gel overlay assay, we identified two Ca2+-dependent CaM binding domains (CaMBDI in the N terminus and CaMBDII in the C terminus). Specific binding of CaM to two CaMBD was confirmed by site-directed mutagenesis, a gel mobility shift assay, and a competition assay using a Ca2+/CaM-dependent enzyme. At increasing concentrations of CaM, the biochemical activity of dual specificity protein phosphatase 1 on the p-nitrophenyl phosphate (pNPP) substrate was increased, whereas activity on the phosphotyrosine of myelin basic protein (MBP) was inhibited. Our results collectively indicate that calmodulin differentially regulates the activity of protein phosphatase, dependent on the substrate. Based on these findings, we propose that the Ca2+ signaling pathway is mediated by CaM cross-talks with a protein phosphorylation signal pathway in plants via protein dephosphorylation.