Spatiotemporal regulation of PKC via interactions with AKAP7 isoforms.
The regulation of kinases by scaffolding proteins greatly contributes to the fidelity of signal transduction. In the present study, we explored an interaction between the ubiquitous enzyme PKC (protein kinase C) and the scaffolding protein AKAP7 (A-kinase-anchoring protein 7). Using protein biochemistry and surface plasmon resonance approaches, we demonstrate that both AKAP7γ and AKAP7α are capable of high-affinity interactions with multiple isoenzymes of PKC. Furthermore, this interaction is achieved via multi-site binding on both proteins. FRET (fluorescence resonance energy transfer) analysis using a PKC activity reporter suggests that anchoring of the kinase within AKAP7 complexes enhances the phosphorylation of substrate proteins. Finally, we determined using FRAP (fluorescence recovery after photobleaching) and virtual modelling that AKAP7 restricts the mobility of PKC within cells by tethering it to subcellular compartments. Collectively, the results of the present study suggests that AKAP7 could play an integral role in dictating PKC localization and function in tissues where the two proteins are co-expressed.
Pat and Jim Calhoun Center for Cardiovascular Research, University of Connecticut Health Center, 263 Farmington Ave, Farmington, CT 06030, USA.
John M
JM
Redden
Andrew V
AV
Le
Arpita
A
Singh
Kyle
K
Federkiewicz
Samantha
S
Smith
Kimberly L
KL
Dodge-Kafka
D054758:A Kinase Anchor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D020134:Catalytic Domain; D002522:Chlorocebus aethiops; D004058:Diffusion; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011493:Protein Kinase C / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014709:Vero Cells
D054758:A Kinase Anchor Proteins; C112596:AKAP7 protein, human; D004800:Enzymes, Immobilized; D007527:Isoenzymes; D008565:Membrane Proteins; D010446:Peptide Fragments; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C496628:PRKCA protein, human; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha