Phosphorylation on the PPP2R5D B regulatory subunit modulates the biochemical properties of protein phosphatase 2A

Abstract

To characterize the biochemical properties of the PP2A regulatory B subunit, PPP2R5D, we analyzed its phosphorylation sites, stoichiometry and effect on holoenzyme activity. PPP2R5D was phosphorylated on Ser-53, Ser-68, Ser-81, and Ser-566 by protein kinase A, and mutations at all four of these sites abolished any significant phosphorylation in vitro. In HEK293 cells, however, the Ser-566 was the major phosphorylation site after PKA activation by forskolin, with marginal phosphorylation on Ser-81. Inhibitory tyrosine phosphorylation on Tyr-307 of the PP2A catalytic C subunit was decreased after forskolin treatment. Kinetic analysis showed that overall PP2A activity was increased with phosphorylation by PPP2R5D phosphorylation. The apparent Km was reduced from 11.25 μM to 1.175 μM with PPP2R5D phosphorylation, resulting in an increase in catalytic activity. These data suggest that PKAmediated activation of PP2A is enabled by PPP2R5D phosphorylation, which modulates the affinity of the PP2A holoenzyme to its physiological substrates. [BMB reports 2010; 43(4): 263-267]

MicroRNA-9 regulates steroid-resistant airway hyperresponsiveness by reducing protein phosphatase 2A activity

“…63 PPP2R5D is a critical PP2A regulatory subunit, and its expression and phosphorylation regulate the catalytic activity of PP2A. [43][44][45][46] To clarify the mechanism behind miR-9 function, we performed a dual luciferase reporter assay and demonstrated miR-9-mediated inhibition of PPP2R5D and PPP2R2A expression. Further investigations in pulmonary macrophages demonstrated that ant-9-mediated inhibition of miR-9 restored expression of PPP2R5D but not PPP2R2A.…”

Compact Modeling of Allosteric Multisite Proteins: Application to a Cell Size Checkpoint

“…In the case of the checkpoint pathway, the active proteins Pkc1 and PP2A have been found to have an approximate of 0.5 [42] and 1.2 [43], respectively, for specific targets. The overall concentrations of their substrates in the cell are much lower - however these proteins tend to localize at the bud, so that the resulting local concentrations are unknown and it is unclear whether a zero-order approach would apply.…”

B56δ-related protein phosphatase 2A dysfunction identified in patients with intellectual disability

“…PKA activates DARPP-32 directly by Thr-34 phosphorylation and indirectly by PP2A-B56δ-dependent activation through Thr-75 dephosphorylation (24). B56δ has several sites for PKA phosphorylation that activate PP2A-B56δ phosphatase activity (24,29). Moreover, PKA-activated PP2A-B56δ-dependent dephosphorylation of another DARPP-32 phosphorylation site (Ser-97) induces nuclear import -mediating dopamine-dependent epigenetic functions (25) -and lack of nuclear PP2A-B56δ targeting has been associated with juvenile myoclonic epilepsy (30).…”