title: Sphingosine kinase interacting protein is an A-kinase anchoring protein specific for type I cAMP-dependent protein kinase.
abstract: The compartmentalization of kinases and phosphatases plays an important role in the specificity of second-messenger-mediated signaling events. Localization of the cAMP-dependent protein kinase is mediated by interaction of its regulatory subunit (PKA-R) with the versatile family of A-kinase-anchoring proteins (AKAPs). Most AKAPs bind avidly to PKA-RII, while some have dual specificity for both PKA-RI and PKA-RII; however, no mammalian PKA-RI-specific AKAPs have thus far been assigned. This has mainly been attributed to the observation that PKA-RI is more cytosolic than the more heavily compartmentalized PKA-RII. Chemical proteomics screens of the cAMP interactome in mammalian heart tissue recently identified sphingosine kinase type 1-interacting protein (SKIP, SPHKAP) as a putative novel AKAP. Biochemical characterization now shows that SPHKAP can be considered as the first mammalian AKAP that preferentially binds to PKA-RIalpha. Recombinant human SPHKAP functions as an RI-specific AKAP that utilizes the characteristic AKAP amphipathic helix for interaction. Further chemical proteomic screening utilizing differential binding characteristics of specific cAMP resins confirms SPHKAPs endogenous specificity for PKA-RI directly in mammalian heart and spleen tissue. Immunolocalization studies revealed that recombinant SPHKAP is expressed in the cytoplasm, where PKA-RIalpha also mainly resides. Alignment of SPHKAPs' amphipathic helix with peptide models of PKA-RI- or PKA-RII-specific anchoring domains shows that it has largely only PKA-RIalpha characteristics. Being the first mammalian PKA-RI-specific AKAP with cytosolic localization, SPHKAP is a very promising model for studying the function of the less explored cytosolic PKA-RI signaling nodes.
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics Group, Bijvoet Center forBiomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Padualaan 8, 3584 CH, Utrecht, The Netherlands.
authors.forename: Duangnapa
authors.initials: D
authors.lastname: Kovanich
authors.forename: Marcel A G
authors.initials: MA
authors.lastname: van der Heyden
authors.forename: Thin Thin
authors.initials: TT
authors.lastname: Aye
authors.forename: Toon A B
authors.initials: TA
authors.lastname: van Veen
authors.forename: Albert J R
authors.initials: AJ
authors.lastname: Heck
authors.forename: Arjen
authors.initials: A
authors.lastname: Scholten
mesh_terms: D054758:A Kinase Anchor Proteins / Q000235:genetics / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D002460:Cell Line; D000242:Cyclic AMP / Q000737:chemistry*; D054743:Cyclic AMP-Dependent Protein Kinase Type I / Q000032:analysis / Q000378:metabolism*; D006352:Heart Ventricles / Q000201:enzymology; D006801:Humans; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D054758:A Kinase Anchor Proteins; D048868:Adaptor Proteins, Signal Transducing; D011994:Recombinant Proteins; C466999:SPHKAP protein, human; D000242:Cyclic AMP; D054743:Cyclic AMP-Dependent Protein Kinase Type I