A potential mechanism for fumonisin B 1 -mediated hepatocarcinogenesis: cyclin D1 stabilization associated with activation of Akt and inhibition of GSK-3β activity

Abstract

Fumonisin B1 (FB1) is a worldwide corn contaminant and has been epidemiologically linked to the high incidence of human esophageal cancer in South Africa and China. FB1 is hepatocarcinogenic in rats by an unknown mechanism. Inhibition of ceramide synthase and disruption of membrane phospholipids have been shown to be mechanisms of toxicity. Here we show overexpression of cyclin D1 protein in both preneoplastic and neoplastic liver specimens obtained from a long-term feeding study of FB1 in rats. In rats fed FB1 short-term, cyclin D1 protein levels in liver were increased up to five-fold in a dose-responsive manner. Northern blot analysis demonstrated no increase in mRNA levels of cyclin D1. 2D electrophoresis of cyclin D1 protein in FB1-treated samples showed a distinct pattern of migration (presence of less negatively charged form of the protein) that differed from controls. Recently, it has been shown that phosphorylation of cyclin D1 by glycogen synthase kinase 3? (GSK-3?) on a single threonine residue (Thr-286) positively regulates proteosomal degradation of cyclin D1. In FB1-treated samples we detected GSK-3? phosphorylated on serine 9; activated protein kinase B (Akt) appears to be responsible for this activity-inhibiting phosphorylation. These findings suggest that overexpression of cyclin D1 results from stabilization due to a lack of phosphorylation mediated by GSK-3?. We also observed an increase in cyclin dependent kinase 4 (Cdk4) complexes with cyclin D1 in FB1-treated samples; additionally, elevated Cdk4 activity was shown by increased phosphorylation of the retinoblastoma protein. In summary, the activation of Akt leads to increased survival, inhibition of GSK-3? activity and post-translational stabilization of cyclin D1, all events responsible for disruption of the cell cycle G1/S restriction point in hepatocytes. This is the first report suggesting the mechanism by which FB1 acts as a carcinogen.