| Popis: |
Serine-threonine Protein phosphatase 2 A (PP2A) is a major member of the PPP family of phosphatases. Increased PP2A activity/signaling regulates cardiac remodeling, contractility, and arrythmogenicity. The core PP2A complex is a hetero-trimeric holoenzyme consisting of a 36 kDa catalytic subunit (PP2Ac), a65 kDa scaffold subunit (PR65A or PP2Aa), and one of at least 18 variable regulatory proteins (B subunits) classified into 4 families. We have recently reported that cardiac PR65A is dephosphorylated in heart failure (Journal of Proteomics 77: 1-13, 2012). In the present study, three in vivo phosphorylations of cardiac PR65A were identified (S303, T268, S314) by MALDI-MS. In structural modeling, these are located at the concave surface and the bottom ridge of the A subunit and within HEAT repeats 7 and 8, suggesting that these residues do not directly contact PP2Ac or the regulatory subunits. Based on the structure of PP2A holoenzymes (PDB codes: 2NPP, 4I5L, 4I5N), Ser303 is buried between HEAT repeats 8 and 9 in all active PP2A holoenzymes, suggesting that significant structural shifts in HEAT repeat 7-9 are required to expose the phosphorylated Ser 303 (P-S303) and to accommodate the phosphate group, thereby alleviating the repulsive interactions between the phosphate group and internal hydrophobic structures and, as a result, stabilize the A subunit in an open conformation that hinders formation of the compact A subunit required for the holoenzymes. These phosphorylations indirectly reduce the interaction of PR65A with PP2Ac and PP2A activity was empirically demonstrated in HEK cells transfected with recombinant forms of PR65A with site-directed mutagenesis of these sites with phosphomimetic and non-phosphorylatable amino acids. Thus, phosphorylation of PR65A regulates PP2A signaling and PR65A dephosporylation may underlie increased PP2A activity in heart failure. |
