Phosphodiesterase 3A binds to 14-3-3 proteins in response to PMA-induced phosphorylation of Ser428
- 8 November 2005
- journal article
- Published by Portland Press Ltd. in Biochemical Journal
- Vol. 392 (1), 163-172
- https://doi.org/10.1042/bj20051103
Abstract
PDE3A (phosphodiesterase 3A) was identified as a phosphoprotein that co-immunoprecipitates with endogenous 14-3-3 proteins from HeLa cell extracts, and binds directly to 14-3-3 proteins in a phosphorylation-dependent manner. Among cellular stimuli tested, PMA promoted maximal binding of PDE3A to 14-3-3 proteins. While p42/p44 MAPK (mitogen-activated protein kinase), SAPK2 (stress-activated protein kinase 2)/p38 and PKC (protein kinase C) were all activated by PMA in HeLa cells, the PMA-induced binding of PDE3A to 14-3-3 proteins was inhibited by the non-specific PKC inhibitors Ro 318220 and H-7, but not by PD 184352, which inhibits MAPK activation, nor by SB 203580 and BIRB0796, which inhibit SAPK2 activation. Binding of PDE3A to 14-3-3 proteins was also blocked by the DNA replication inhibitors aphidicolin and mimosine, but the PDE3A–14-3-3 interaction was not cell-cycle-regulated. PDE3A isolated from cells was able to bind to 14-3-3 proteins after in vitro phosphorylation with PKC isoforms. Using MS/MS of IMAC (immobilized metal ion affinity chromatography)-enriched tryptic phosphopeptides and phosphospecific antibodies, at least five sites on PDE3A were found to be phosphorylated in vivo, of which Ser428 was selectively phosphorylated in response to PMA and dephosphorylated in cells treated with aphidicolin and mimosine. Phosphorylation of Ser428 therefore correlated with 14-3-3 binding to PDE3A. Ser312 of PDE3A was phosphorylated in an H-89-sensitive response to forskolin, indicative of phosphorylation by PKA (cAMP-dependent protein kinase), but phosphorylation at this site did not stimulate 14-3-3 binding. Thus 14-3-3 proteins can discriminate between sites in a region of multisite phosphorylation on PDE3A. An additional observation was that the cytoskeletal cross-linker protein plectin-1 coimmunoprecipitated with PDE3A independently of 14-3-3 binding.Keywords
This publication has 46 references indexed in Scilit:
- Platelet Cyclic Adenosine Monophosphate Phosphodiesterases: Targets for Regulating Platelet-Related ThrombosisSeminars in Thrombosis and Hemostasis, 2004
- Crystal Structure of Human Phosphodiesterase 3B: Atomic Basis for Substrate and Inhibitor SpecificityBiochemistry, 2004
- Phosphorylation of the vasodilator-stimulated phosphoprotein (VASP) by the anti-platelet drug, cilostazol, in plateletsPlatelets, 2003
- 14-3-3s regulate fructose-2,6-bisphosphate levels by binding to PKB-phosphorylated cardiac fructose-2,6-bisphosphate kinase/phosphataseThe EMBO Journal, 2003
- PKD: a new protein kinase C–dependent pathway in plateletsBlood, 2003
- Membrane Localization of Cyclic Nucleotide Phosphodiesterase 3 (PDE3)Published by Elsevier BV ,2000
- Specificity and mechanism of action of some commonly used protein kinase inhibitorsBiochemical Journal, 2000
- Insulin-Induced Phosphorylation and Activation of Phosphodiesterase 3B in Rat Adipocytes: Possible Role for Protein Kinase B But Not Mitogen-Activated Protein Kinase or p70 S6 KinaseEndocrinology, 1998
- Identification of the Site in the cGMP-inhibited Phosphodiesterase Phosphorylated in Adipocytes in Response to Insulin and IsoproterenolPublished by Elsevier BV ,1996
- SB 203580 is a specific inhibitor of a MAP kinase homologue which is stimulated by cellular stresses and interleukin‐1FEBS Letters, 1995