Alpha-kinase anchoring protein alphaKAP interacts with SERCA2A to spatially position Ca2+/calmodulin-dependent protein kinase II and modulate phospholamban phosphorylation.
The sarco-endoplasmic reticulum calcium ATPase 2a (SERCA2a) is critical for sequestering cytosolic calcium into the sarco-endoplasmic reticulum (SR) and regulating cardiac muscle relaxation. Protein-protein interactions indicated that it exists in complex with Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) and its anchoring protein alphaKAP. Confocal imaging of isolated cardiomyocytes revealed the colocalization of CAMKII and alphaKAP with SERCA2a at the SR. Deletion analysis indicated that SERCA2a and CaMKII bind to different regions in the association domain of alphaKAP but not with each other. Although deletion of the putative N-terminal hydrophobic amino acid stretch in alphaKAP prevented its membrane targeting, it did not influence binding to SERCA2a or CaMKII. Both CaMKIIdelta(C) and the novel CaMKIIbeta(4) isoforms were found to exist in complex with alphaKAP and SERCA2a at the SR and were able to phosphorylate Thr-17 on phospholamban (PLN), an accessory subunit and known regulator of SERCA2a activity. Interestingly, the presence of alphaKAP was also found to significantly modulate the Ca(2+)/calmodulin-dependent phosphorylation of Thr-17 on PLN. These data demonstrate that alphaKAP exhibits a novel interaction with SERCA2a and may serve to spatially position CaMKII isoforms at the SR and to uniquely modulate the phosphorylation of PLN.
Singh
Puneet
P
Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, Ontario K1H 8M5, Canada.
Salih
Maysoon
M
Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, Ontario K1H 8M5, Canada.
Tuana
Balwant S
BS
Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, Ontario K1H 8M5, Canada. Electronic address: btuana@uottawa.ca.
D054758:A Kinase Anchor Proteins / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000235:genetics / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008861:Microsomes / Q000201:enzymology; D009206:Myocardium / Q000166:cytology / Q000201:enzymology; D010766:Phosphorylation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D000099211:Phospholamban
D054758:A Kinase Anchor Proteins; D002135:Calcium-Binding Proteins; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D002147:Calmodulin; D007527:Isoenzymes; D011993:Recombinant Fusion Proteins; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000099211:Phospholamban