Cloning and mitochondrial localization of full-length D-AKAP2, a protein kinase A anchoring protein

Differential compartmentalization of signaling molecules in cells and tissu es is being recognized as an important mechanism for regulating the specifi city of signal transduction pathways. A kinase anchoring proteins (AKAPs) d irect the subcellular localization of protein kinase A (PKA) by binding to its regulatory (R) subunits. Dual specific AKAPs (D-AKAPs) interact with bo th RI and RII. A 372-residue fragment of mouse D-AKAP2 with a 40-residue C- terminal PKA binding region and a putative regulator of G protein signaling (RCS) domain was previously identified by means of a yeast two-hybrid scre en. Here, we report: the cloning of full-length human D-AKAP2 (662 residues ) with an additional putative RCS domain, and the corresponding mouse prote in less the first two exons (617 residues). Expression of D-AKAP2 was chara cterized by using mouse tissue extracts. Full-length D-AKAP2 from various t issues shows different molecular weights, possibly because of alternative s plicing or posttranslational modifications. The cloned human gene product h as a molecular weight similar to one of the prominent mouse proteins. In vi vo association of D-AKAP2 with PKA in mouse brain was demonstrated by using cAMP agarose pull-down assay. Subcellular localization for endogenous mous e, rat, and human D-AKAP2 was determined by immunocytochemistry, immunohist ochemistry, and tissue fractionation. D-AKAP2 from all three species is hig hly enriched in mitochondria. The mitochondrial localization and the presen ce of RGS domains in D-AKAP2 may have important implications for its functi on in PKA and G protein signal transduction.