Unknown

Dataset Information

0

Multisite phosphorylation of 14-3-3 proteins by calcium-dependent protein kinases.


ABSTRACT: Plant 14-3-3 proteins are phosphorylated at multiple sites in vivo; however, the protein kinase(s) responsible are unknown. Of the 34 CPK (calcium-dependent protein kinase) paralogues in Arabidopsis thaliana, three (CPK1, CPK24 and CPK28) contain a canonical 14-3-3-binding motif. These three, in addition to CPK3, CPK6 and CPK8, were tested for activity against recombinant 14-3-3 proteins ? and ?. Using an MS-based quantitative assay we demonstrate phosphorylation of 14-3-3 ? and ? at a total of seven sites, one of which is an in vivo site discovered in Arabidopsis. CPK autophosphorylation was also comprehensively monitored by MS and revealed a total of 45 sites among the six CPKs analysed, most of which were located within the N-terminal variable and catalytic domains. Among these CPK autophosphorylation sites was Tyr463 within the calcium-binding EF-hand domain of CPK28. Of all CPKs assayed, CPK28, which contained an autophosphorylation site (Ser43) within a canonical 14-3-3-binding motif, showed the highest activity against 14-3-3 proteins. Phosphomimetic mutagenesis of Ser72 to aspartate on 14-3-3?, which is adjacent to the 14-3-3-binding cleft and conserved among all 14-3-3 isoforms, prevented 14-3-3-mediated inhibition of phosphorylated nitrate reductase.

SUBMITTER: Swatek KN 

PROVIDER: S-EPMC4127189 | biostudies-literature | 2014 Apr

REPOSITORIES: biostudies-literature

altmetric image

Publications

Multisite phosphorylation of 14-3-3 proteins by calcium-dependent protein kinases.

Swatek Kirby N KN   Wilson Rashaun S RS   Ahsan Nagib N   Tritz Rebecca L RL   Thelen Jay J JJ  

The Biochemical journal 20140401 1


Plant 14-3-3 proteins are phosphorylated at multiple sites in vivo; however, the protein kinase(s) responsible are unknown. Of the 34 CPK (calcium-dependent protein kinase) paralogues in Arabidopsis thaliana, three (CPK1, CPK24 and CPK28) contain a canonical 14-3-3-binding motif. These three, in addition to CPK3, CPK6 and CPK8, were tested for activity against recombinant 14-3-3 proteins χ and ε. Using an MS-based quantitative assay we demonstrate phosphorylation of 14-3-3 χ and ε at a total of  ...[more]

Similar Datasets

| S-EPMC4568647 | biostudies-literature
| S-EPMC3971370 | biostudies-literature
| S-EPMC3006207 | biostudies-literature
| S-EPMC3259650 | biostudies-literature
| S-EPMC3151052 | biostudies-literature
| S-EPMC8126791 | biostudies-literature
| S-EPMC7700312 | biostudies-literature
| S-EPMC3675764 | biostudies-literature
| S-EPMC7588190 | biostudies-literature
| S-EPMC4485940 | biostudies-literature