Phosphatidylinositol 3-kinase (PI 3-kinase) is a cytoplasmic signaling molecule that is recruited to activated
growth factor receptors and has been shown to be involved in regulation of stimulated exocytosis and endocytosis. One of the downstream signaling molecules activated by
PI 3-kinase is the
protein kinase Akt. Previous studies have indicated that
PI 3-kinase is necessary for basal
Na(+)/H(+) exchanger 3 (NHE3) transport and for
fibroblast growth factor-stimulated NHE3 activity in PS120 fibroblasts. However, it is not known whether activation of
PI 3-kinase is sufficient to stimulate NHE3 activity or whether Akt is involved in this
PI 3-kinase effect. We used an adenoviral
infection system to test the possibility that activation of
PI 3-kinase or Akt alone is sufficient to stimulate NHE3 activity. This hypothesis was investigated in PS120 fibroblasts stably expressing NHE3 after somatic gene transfer using a replication-deficient recombinant adenovirus containing constitutively active catalytic subunit of
PI 3-kinase or constitutively active Akt. The adenovirus construct used was engineered with an upstream
ecdysone promoter to allow time-regulated expression. Adenoviral
infection was nearly 100% at 48 h after
infection. Forty-eight hours after
infection (24 h after activation of the
ecdysone promoter),
PI 3-kinase and Akt amount and activity were increased. Increases in both
PI 3-kinase activity and Akt activity stimulated NHE3 transport. In addition, a membrane-permeant synthetic 10-mer
peptide that binds
polyphosphoinositides and increases
PI 3-kinase activity similarly enhanced NHE3 transport activity and also increased the percentage of NHE3 on the plasma membrane. The magnitudes of stimulation of NHE3 by constitutively active
PI 3-kinase,
PI 3-kinase peptide, and constitutively active Akt were similar to each other. These results demonstrate that activation of
PI 3-kinase or Akt is sufficient to stimulate NHE3 transport activity in PS120/NHE3 cells.