Cellular and viral trans-acting factors modulate N-myc2 promoter activity in woodchuck liver tumors.
Activation of the N-myc2 oncogene by integration of woodchuck hepatitis virus (WHV) DNA is a central event in woodchuck liver oncogenesis. In this study, we have evaluated the influence of several cellular and viral trans-acting factors and mediators of inflammation on N-myc2 promoter activity in hepatoma cell lines. Ets oncoproteins, including Ets1, Ets2 and PEA3 efficiently activated a chimeric N-myc2 promoter/luciferase reporter gene. By electrophoretic mobility shift assays, we show that Etsl and Ets2 proteins can efficiently bind two consensus Ets sites located within a 59 bp sequence upstream of the N-myc2 transcription start site. Site-directed mutagenesis of these Ets-binding motifs abolished transactivation of the N-myc2 promoter by Ets proteins. Addition of interleukin-6 (IL-6) induced a weak but reproducible activation of the N-myc2 promoter, while IL-1 was ineffective. We further show that the N-myc2 promoter can be transactivated by the hepadna-virus X protein, and that distal promoter sequences are required for both IL-6 and X responsiveness. Similar effects of these factors were observed in the context of the N-myc2 promoter activated by WHV cis-regulatory elements. In view of the high-level expression of the N-myc2 oncogene in most woodchuck liver tumors, the Ets oncoproteins, inflammation-associated cytokine IL-6 and the viral X transactivator might play important roles in hepadnavirus-associated tumorigenesis.
Flajolet
M
M
Unité de Recombinaison et Expression Génétique (INSERM U163), Institut Pasteur, Paris, France.
Gegonne
A
A
Ghysdael
J
J
Tiollais
P
P
Buendia
M A
MA
Fourel
G
G
D000818:Animals; D001665:Binding Sites; D006528:Carcinoma, Hepatocellular / Q000235:genetics*; D004268:DNA-Binding Proteins; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D016259:Genes, myc; D017919:Hepatitis B Virus, Woodchuck / Q000235:genetics; D015850:Interleukin-6 / Q000494:pharmacology; D008113:Liver Neoplasms / Q000235:genetics*; D008392:Marmota; D011401:Promoter Regions, Genetic / Q000187:drug effects; D050784:Proto-Oncogene Protein c-ets-1; D050787:Proto-Oncogene Protein c-ets-2; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000378:metabolism; D050783:Proto-Oncogene Proteins c-ets; D012097:Repressor Proteins; D015534:Trans-Activators / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014157:Transcription Factors / Q000096:biosynthesis / Q000378:metabolism; D014407:Tumor Cells, Cultured; D054334:Viral Regulatory and Accessory Proteins
D004268:DNA-Binding Proteins; C107465:ERF protein, human; C494041:ETS1 protein, human; D015850:Interleukin-6; D050784:Proto-Oncogene Protein c-ets-1; D050787:Proto-Oncogene Protein c-ets-2; D011518:Proto-Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D012097:Repressor Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein