p85 regulatory subunit of PI3K mediates cAMP–PKA and estrogens biological effects on growth and survival.

Cyclicadenosine3050monophosphate(cAMP)andprotein kinase A (PKA) cooperate with phosphatidylinositol 30 kinase (PI3K) signals in the control of growth and survival. To determine the molecular mechanism(s) involved, we identified and mutagenized a specific serine (residue 83) in p85aPI3K, which is phosphorylated in vivo and in vitro by PKA. Expression of p85aPI3K mutants (alanineorasparticsubstitutions)significantlyalteredthe biological responses of the cells to cAMP. cAMP protection from anoikis was reduced in cells expressing the alanine version p85aPI3K. These cells did not arrest in G1 in the presence of cAMP, whereas cells expressing the asparticmutantp85DaccumulatedinG1eveninthe absenceofcAMP.Sphasewasstillefficientlyinhibitedby cAMP in cells expressing both mutants. The binding of PI3K to Ras p21 was greatly reduced in cells expressing p85A in the presence or absence of cAMP. Conversely, expressionoftheasparticmutantstimulatedrobustlythe binding of PI3K to p21 Ras in the presence of cAMP. MutationintheSer83inhibitedcAMP,butnotPDGF stimulation of PI3K. Conversely, the p85D aspartic mutantamplifiedcAMPstimulationofPI3Kactivity. Phosphorylation of Ser 83 by cAMP–PKA in p85aPI3K was also necessary for estrogen signaling as expression of p85Aorp85Dmutantsinhibitedoramplified,respec- tively,thebindingofestrogenreceptortop85aandAKT phosphorylation induced by estrogens. The data presented indicate that: (1) phosphorylation of Ser 83 in p85aPI3K is critical for cAMP–PKA induced G1 arrest and survival in mouse 3T3 fibroblasts; (2) this site is necessary for amplification of estrogen signals by cAMP–PKA and relatedreceptors.Finally,thesedatasuggestageneral mechanism of PI3K regulation by cAMP, operating in various cell types and under different conditions.