Association of the type 1 protein phosphatase PP1 with the A-kinase anchoring protein AKAP220.
The cyclic AMP (cAMP)-dependent protein kinase (PKA) and the type 1 protein phosphatase (PP1) are broad-specificity signaling enzymes with opposing actions that catalyze changes in the phosphorylation state of cellular proteins. Subcellular targeting to the vicinity of preferred substrates is a means of restricting the specificity of each enzyme [1] [2]. Compartmentalization of the PKA holoenzyme is mediated through association of the regulatory subunits with A-kinase anchoring proteins (AKAPs), whereas a diverse family of phosphatase-targeting subunits directs the location of the PP1 catalytic subunit (PP1c) [3] [4]. Here, we demonstrate that the PKA-anchoring protein, AKAP220, binds PP1c with a dissociation constant (KD) of 12.1 +/- 4 nM in vitro. Immunoprecipitation of PP1 from cell extracts resulted in a 10.4 +/- 3.8-fold enrichment of PKA activity. AKAP220 co-purified with PP1c by affinity chromatography on microcystin sepharos Immunocytochemical analysis demonstrated that the kinase, the phosphatase and the anchoring protein had distinct but overlapping staining patterns in rat hippocampal neurons. Collectively, these results provide the first evidence that AKAP220 is a multivalent anchoring protein that maintains a signaling scaffold of PP1 and the PKA holoenzyme.
Schillace
R V
RV
Howard Hughes Medical Institute, L-474 Vollum Institute, Oregon Health Sciences University, 3181 S.W. Sam Jackson Park Road, Portland, Oregon 97201-3098, USA.
Scott
J D
JD
D054758:A Kinase Anchor Proteins; D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins; D002451:Cell Compartmentation; D002462:Cell Membrane / Q000737:chemistry* / Q000648:ultrastructure; D002846:Chromatography, Affinity; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000032:analysis*; D004789:Enzyme Activation; D006624:Hippocampus / Q000737:chemistry / Q000166:cytology*; D006801:Humans; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000032:analysis*; D052998:Microcystins; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000032:analysis*; D016501:Neurites / Q000737:chemistry / Q000648:ultrastructure; D009474:Neurons / Q000737:chemistry* / Q000648:ultrastructure; D010456:Peptides, Cyclic / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D054645:Protein Phosphatase 1; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000032:analysis*; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015290:Second Messenger Systems
D054758:A Kinase Anchor Proteins; C099270:AKAP11 protein, human; D002352:Carrier Proteins; D046911:Macromolecular Substances; D008565:Membrane Proteins; D052998:Microcystins; D009419:Nerve Tissue Proteins; D010456:Peptides, Cyclic; D011506:Proteins; D011993:Recombinant Fusion Proteins; C078588:microcystin; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1