A potential mechanism for fumonisin B(1)-mediated hepatocarcinogenesis: cyclin D1 stabilization associated with activation of Akt and inhibition of GSK-3beta activity.
Fumonisin B(1) (FB(1)) is a worldwide corn contaminant and has been epidemiologically linked to the high incidence of human esophageal cancer in South Africa and China. FB(1) 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 FB(1) in rats. In rats fed FB(1) 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 FB(1)-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 3beta (GSK-3beta) on a single threonine residue (Thr-286) positively regulates proteosomal degradation of cyclin D1. In FB(1)-treated samples we detected GSK-3beta 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-3beta. We also observed an increase in cyclin dependent kinase 4 (Cdk4) complexes with cyclin D1 in FB(1)-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-3beta activity and post-translational stabilization of cyclin D1, all events responsible for disruption of the cell cycle G(1)/S restriction point in hepatocytes. This is the first report suggesting the mechanism by which FB(1) acts as a carcinogen.
Laboratory of Comparative Carcinogenesis, National Cancer Institute, Frederick Cancer Research and Development Center, Building 538, Room 205E, Frederick, MD 21702, USA. ramljak@mail.ncifcrf.gov
D
D
Ramljak
R J
RJ
Calvert
P W
PW
Wiesenfeld
B A
BA
Diwan
B
B
Catipovic
W F
WF
Marasas
T C
TC
Victor
L M
LM
Anderson
W C
WC
Gelderblom
D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D002264:Carboxylic Acids / Q000633:toxicity*; D002274:Carcinogens, Environmental / Q000633:toxicity*; D019938:Cyclin D1 / Q000096:biosynthesis / Q000378:metabolism*; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D020536:Enzyme Activators / Q000633:toxicity; D037341:Fumonisins; D016193:G1 Phase / Q000187:drug effects / Q000502:physiology; D015972:Gene Expression Regulation, Neoplastic; D011905:Genes, ras / Q000235:genetics; D038362:Glycogen Synthase Kinase 3; D038341:Glycogen Synthase Kinases; D008099:Liver / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D008114:Liver Neoplasms, Experimental / Q000139:chemically induced* / Q000201:enzymology / Q000378:metabolism*; D009154:Mutation; D009183:Mycotoxins / Q000633:toxicity*; D011230:Precancerous Conditions / Q000139:chemically induced / Q000201:enzymology / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects*; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D011916:Rats, Inbred F344; D016196:S Phase / Q000187:drug effects / Q000502:physiology
D002264:Carboxylic Acids; D002274:Carcinogens, Environmental; D020536:Enzyme Activators; D037341:Fumonisins; D009183:Mycotoxins; D011518:Proto-Oncogene Proteins; D019938:Cyclin D1; C056933:fumonisin B1; D038341:Glycogen Synthase Kinases; C494920:Akt1 protein, rat; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C495903:Cdk4 protein, rat; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases; D038362:Glycogen Synthase Kinase 3