Insulin and IGF-1 stimulate the β-catenin pathway through two signalling cascades involving GSK-3β inhibition and Ras activation

We examined the interplay between the insulin/LGF-1- and beta -catenin-regulated pathways, both of which are suspected to play a role in hepatocarcinogenesis. Insulin and IGF-1 stimulated the transcription of a Lef/Tcf-dependent luciferase reporter gene by 3-4-fold in HepG2 cells, This stimulation was mediated through the activation of phosphatidylinositol 3-kinase (PI 3-K)/Akt and the inhibition of glycogen synthase kinase-3 beta (GSK-3 beta) since the effects of insulin and IGF-I were inhibited by dominant-negative mutants of PI 3-K or Akt and an uninhibitable GSK-3 beta. Together with inhibiting GSK-3 beta, insulin and IGF-I increased the cytoplasmic levels of beta -catenin, The PI 3-K/Akt/GSK-3 beta pathway was not the sole to mediate insulin and IGF-1 stimulation of Lef/Tcf-dependent transcription. The Ras signalling pathway was also required as (i) the stimulatory effects of insulin and IGF-I were inhibited by dominant-negative Ras or the MEK1 inhibitor PD98059 and (ii) activated Ha-Res or constitutively active MEK synergized with catalytically inactive GSK-3 beta to stimulate Lef/Tcf-dependent transcription. This study provides the first evidence that insulin and IGF-I stimulate the beta -catenin pathway through tw a signalling cascades bifurcating downstream of PI 3-K and involving GSK-3 beta inhibition and Ras activation. These findings demonstrate for the first time the ability of insulin and IGF-I to activate the beta -catenin pathway in hepatoma cells and thereby provide new insights into the role of these factors in hepatocarcinogenesis.