The Phosphatidylinositol 3-Kinase/Akt Cassette Regulates Purine Nucleotide Synthesis

Wei Wang,Alla Fridman,William Blackledge,Stephen Connelly,Ian A Wilson,Renate B Pilz,Gerry R Boss

doi:10.1074/jbc.m806707200

Abstract

The phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway is highly conserved throughout evolution and regulates cell size and survival and cell cycle progression. It regulates the latter by stimulating procession through G(1) and the G(1)/S phase transition. Entry into S phase requires an abundant supply of purine nucleotides, but the effect of the PI3K/Akt pathway on purine synthesis has not been studied. We now show that the PI3K/Akt cassette regulates both de novo and salvage purine nucleotide synthesis in insulin-responsive mouse mesenchymal cells. We found that serum and insulin stimulated de novo purine synthesis in serum-starved cells largely through PI3K/Akt signaling, and pharmacologic and genetic inhibition of PI3K/Akt reduced de novo synthesis by 75% in logarithmically growing cells. PI3K/Akt regulated early steps of de novo synthesis by modulating phosphoribosylpyrophosphate production by the non-oxidative pentose phosphate pathway and late steps by modulating activity of the bifunctional enzyme aminoimidazole-carboxamide ribonucleotide transformylase IMP cyclohydrolase, an enzyme not previously known to be regulated. The effects of PI3K/Akt on purine nucleotide salvage were likely through regulating phosphoribosylpyrophosphate availability. These studies define a new mechanism whereby the PI3K/Akt cassette functions as a master regulator of cellular metabolism and a key player in oncogenesis.

Highlights

Several intracellular processes including protein synthesis, glucose metabolism, and cell cycle progression [1, 2]
Effect of Akt Inhibition on de Novo Purine Synthesis—To determine whether the phosphatidylinositol 3-kinase (PI3K)/Akt cassette regulates de novo purine synthesis, we treated C2C12 cells with LY294002 for 2 h and found that the drug sponding areas on the thin layer chromatography (TLC) plates were cut out, and radioactiv- reduced [14C]formate incorporation into purines by Ͼ75% over ity in formylglycinamide ribonucleotide (FGAR) was measured by liquid scintillation counting
The marked reduction of purine synthesis in serum-starved cells in the presence of some residual Akt activity suggests that serum regulates purine synthesis in part through non-PI3K/Akt mechanisms; further reduction in purine synthesis by LY294002 in serum-starved cells is consistent with this hypothesis

Summary

Introduction

Several intracellular processes including protein synthesis, glucose metabolism, and cell cycle progression [1, 2]. PI3K/Akt Regulate Purine Nucleotide Synthesis dependent hematopoietic cell line FL5.12, activity of the oxidative pentose phosphate pathway is higher in cells expressing myristoylated Akt than in control cells and does not decrease on removing IL-3 as occurs in control cells [4].

Results

Conclusion