Inhibition of Protein Phosphatase 2A Activity by PI3Kγ Regulates β-Adrenergic Receptor Function

Abstract

Phosphoinositide 3-kinaseγ (PI3Kγ) is activated by G-protein coupled receptors (GPCRs). We show here that PI3Kγ inhibits protein phosphatase 2A (PP2A) at the β-adrenergic receptor (βAR, a GPCR) complex altering G-protein coupling. PI3Kγ inhibition results in significant increase of βAR-associated phosphatase activity leading to receptor dephosphorylation and resensitization preserving cardiac function. Mechanistically, PI3Kγ inhibits PP2A activity at the βAR complex by phosphorylating an intracellular inhibitor of PP2A (I2PP2A) on serine residues 9 & 93 resulting in enhanced binding to PP2A. Indeed, enhanced phosphorylation of β2ARs is observed with phosphomimetic I2PP2A mutant that was completely reversed with a mutant mimicking dephosphorylated state. siRNA depletion of endogenous I2PP2A augments PP2A activity despite active PI3K resulting in β2AR dephosphorylation and sustained signaling. Our study provides the underpinnings of a PI3Kγ mediated regulation of PP2A activity that has significant consequences on receptor function with broad implications in cellular signaling.

Antagonistic activities of the immunomodulator and PP2A-activating drug FTY720 (Fingolimod, Gilenya) in Jak2-driven hematologic malignancies

“…Reportedly, SET undergoes PI-3Kg-mediated serine 9 and 24 phosphorylation by PKC that controls SET nuclear export and, therefore, its ability to act as a inhibitor of PP2A. 23,25,26 Thus, Jak2 V617F -generated signals may suppress PP2A activity through the PI-3Kg-PKC-mediated SET phosphorylation on these serine residues ( Figure 2A). In Ba/F3 cells, ectopic expression of WT and phosphomimetic mutation of SET serine 9 into glutamic acid (S9E) resulted in an ;80% inhibition of PP2A activity ( Figure 2B, bottom left).…”

The Rac1 hypervariable region in targeting and signaling

“…121 This interaction is enhanced using a SET mutant that encodes a S9E mutation, which was used to mimic phosphorylation of SET. 120,121 In addition, this mutant of SET does no longer dimerize, but still binds to PP2A suggesting that phosphorylation of SET on Ser9 promotes monomerization, binding to Rac1 and PP2A concomitant with shuttling toward the cytoplasm. Using a membranetargeted SET mutant, we could show that increased plasma membrane localization of SET cooperated with active Rac1 to stimulate cell migration 121 (Fig.…”

SET Protein Interacts with Intracellular Domains of the Gonadotropin-releasing Hormone Receptor and Differentially Regulates Receptor Signaling to cAMP and Calcium in Gonadotrope Cells

“…7). Introduction of ICL1 peptide into ␣T3-1 cells decreased by 33 Ϯ 4% cAMP production in response to GnRHa but did not have any impact on cAMP production by PACAP 38 (Fig. 7, ICL1 WT).…”

“…The 38-amino acid form of pituitary adenylyl cyclase-activating polypeptide (PACAP 38 ) was from Calbiochem (VWR International, Strasbourg, France). Iodine-125 radionuclide was purchased from PerkinElmer Life Sciences.…”

“…Cells were preincubated with a cellpermeable ICL1 peptide and challenged either with GnRHa or PACAP 38 , and intracellular cAMP production was measured in real time in ␣T3-1 living cells (Fig. 7).…”