Protein phosphatase 2A is regulated by protein kinase Cα (PKCα)-dependent phosphorylation of its targeting subunit B56α at Ser41.
Protein phosphatase 2A (PP2A) is a family of multifunctional serine/threonine phosphatases consisting of a catalytic C, a structural A, and a regulatory B subunit. The substrate and therefore the functional specificity of PP2A are determined by the assembly of the enzyme complex with the appropriate regulatory B subunit families, namely B55, B56, PR72, or PR93/PR110. It has been suggested that additional levels of regulating PP2A function may result from the phosphorylation of B56 isoforms. In this study, we identified a novel phosphorylation site at Ser(41) of B56α. This phosphoamino acid residue was efficiently phosphorylated in vitro by PKCα. We detected a 7-fold higher phosphorylation of B56α in failing human hearts compared with nonfailing hearts. Purified PP2A dimeric holoenzyme (subunits C and A) was able to dephosphorylate PKCα-phosphorylated B56α. The potency of B56α for PP2A inhibition was markedly increased by PKCα phosphorylation. PP2A activity was also reduced in HEK293 cells transfected with a B56α mutant, where serine 41 was replaced by aspartic acid, which mimics phosphorylation. More evidence for a functional role of PKCα-dependent phosphorylation of B56α was derived from Fluo-4 fluorescence measurements in phenylephrine-stimulated Flp293 cells. The endoplasmic reticulum Ca(2+) release was increased by 23% by expression of the pseudophosphorylated form compared with wild-type B56α. Taken together, our results suggest that PKCα can modify PP2A activity by phosphorylation of B56α at Ser(41). This interplay between PKCα and PP2A represents a new mechanism to regulate important cellular functions like cellular Ca(2+) homeostasis.
From the Institut für Pharmakologie und Toxikologie, Universitätsklinikum Münster, 48149 Münster, Germany.
Uwe
U
Kirchhefer
Alexander
A
Heinick
Simone
S
König
Torsten
T
Kristensen
Frank U
FU
Müller
Matthias D
MD
Seidl
Peter
P
Boknik
D058646:Adrenergic alpha-1 Receptor Agonists / Q000494:pharmacology; D019943:Amino Acid Substitution; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006706:Homeostasis / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D020125:Mutation, Missense; D010656:Phenylephrine / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects / Q000502:physiology; D051571:Protein Kinase C-alpha / Q000235:genetics / Q000378:metabolism*; D054648:Protein Phosphatase 2 / Q000235:genetics / Q000378:metabolism*; D012694:Serine / Q000235:genetics / Q000378:metabolism; D061987:Sf9 Cells; D018411:Spodoptera
D058646:Adrenergic alpha-1 Receptor Agonists; C525690:PPP2R5A protein, human; D010656:Phenylephrine; D012694:Serine; C496628:PRKCA protein, human; D051571:Protein Kinase C-alpha; D054648:Protein Phosphatase 2; D002118:Calcium
Calcium Intracellular Release; Heart Failure; Protein Kinase C (PKC); Protein Phosphorylation; Serine/Threonine Protein Phosphatase