Protein Kinase A Opposes the Phosphorylation-dependent Recruitment of Glycogen Synthase Kinase 3β to A-kinase Anchoring Protein 220

• Published in: The Journal of biological chemistry 2015-08, Vol.290 (32), p.19445-19457
• Main Authors: Whiting, Jennifer L., Nygren, Patrick J., Tunquist, Brian J., Langeberg, Lorene K., Seternes, Ole-Morten, Scott, John D.
• Format: Article
• Language: English
• Subjects: A Kinase Anchor Proteins - genetics; A Kinase Anchor Proteins - metabolism; A-kinase anchoring protein (AKAP); Amino Acid Sequence; Animals; Binding Sites; Cell Line; cyclic AMP (cAMP); Cyclic AMP-Dependent Protein Kinases - genetics; Cyclic AMP-Dependent Protein Kinases - metabolism; Epithelial Cells - cytology; Epithelial Cells - enzymology; Gene Expression Regulation - drug effects; Genetic Engineering; glycogen synthase kinase 3 (GSK-3); Glycogen Synthase Kinase 3 - genetics; Glycogen Synthase Kinase 3 - metabolism; Glycogen Synthase Kinase 3 beta; HEK293 Cells; Holoenzymes - genetics; Holoenzymes - metabolism; Humans; Mice; Molecular Sequence Data; Phosphorylation; Protein Binding; protein complex; Protein Interaction Domains and Motifs; protein kinase; protein kinase A (PKA); protein phosphorylation; Protein Structure, Secondary; Protein Subunits - genetics; Protein Subunits - metabolism; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Sequence Alignment; Signal Transduction; β-catenin (B-catenin)
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1074/jbc.M115.654822

The proximity of an enzyme to its substrate can influence rate and magnitude of catalysis. A-kinase anchoring protein 220 (AKAP220) is a multivalent anchoring protein that can sequester a variety of signal transduction enzymes. These include protein kinase A (PKA) and glycogen synthase kinase 3β (GSK3β). Using a combination of molecular and cellular approaches we show that GSK3β phosphorylation of Thr-1132 on AKAP220 initiates recruitment of this kinase into the enzyme scaffold. We also find that AKAP220 anchors GSK3β and its substrate β-catenin in membrane ruffles. Interestingly, GSK3β can be released from the multienzyme complex in response to PKA phosphorylation on serine 9, which suppresses GSK3β activity. The signaling scaffold may enhance this regulatory mechanism, as AKAP220 has the capacity to anchor two PKA holoenzymes. Site 1 on AKAP220 (residues 610–623) preferentially interacts with RII, whereas site 2 (residues 1633–1646) exhibits a dual specificity for RI and RII. In vitro affinity measurements revealed that site 2 on AKAP220 binds RII with ∼10-fold higher affinity than site 1. Occupancy of both R subunit binding sites on AKAP220 could provide a mechanism to amplify local cAMP responses and enable cross-talk between PKA and GSK3β. Background: AKAPs integrate intracellular signals by sequestering PKA with other kinases. Results: Phosphorylation of Thr-1132 on AKAP220 initiates GSK3β recruitment, and PKA activity drives the release of GSK3β from the complex. Conclusion: Cross-talk between PKA and GSK3β is optimized in the context of AKAP220 multienzyme complexes. Significance: Signal responsive assembly of enzyme complexes may represent a general mechanism to diversify transduction through AKAPs.