E2F and Sp1/Sp3 Synergize but are not sufficient to activate the MYCN gene in neuroblastomas.
Amplification of the MYCN gene, resulting in overexpression of MYCN, distinguishes a subset of neuroblastomas with poor prognosis. We recently identified MYCN as a target gene of the E2F transcription factors. Here we show that Sp1 and Sp3 cooperate with E2F-1 to activate the MYCN promoter. However, in a neuroblastoma cell line that does not express MYCN, overexpression of E2F-1 was not sufficient to activate the MYCN promoter even in the presence of trichostatin A and 5-aza-cytidine. This was because of a failure of E2F-1 to bind to the MYCN promoter in these cells, although access of E2F-1 to the inactive MYCN promoter was not blocked by a nucleosome. Differences in nucleosomal organization of the MYCN promoter in different cell lines did not correlate with gene activation per se but with the switch from basal to activated transcription. Binding of E2F and Sp1/Sp3 to the MYCN promoter in vivo correlated with acetylation of histones H3 and H4 and recruitment of RNA polymerase II and the protein acetyltransferase Tip60 but not with nucleosome remodeling. Our results define distinct chromatin states of the MYCN promoter, indicate that factors in addition to E2F and Sp1/Sp3 are required to activate MYCN in neuroblastomas, and provide evidence for a novel mechanism of controlling access of E2F to selected target genes.
Kramps
Christoph
C
Institute of Molecular Biology and Tumor Research, 35033 Marburg, Germany.
Strieder
Verena
V
Sapetschnig
Alexandra
A
Suske
Guntram
G
Lutz
Werner
W
D000107:Acetylation; D000123:Acetyltransferases / Q000378:metabolism; D017209:Apoptosis; D018797:Cell Cycle Proteins; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D002469:Cell Separation; D002843:Chromatin / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D005434:Flow Cytometry; D017930:Genes, Reporter; D051548:Histone Acetyltransferases; D006657:Histones / Q000378:metabolism; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D008156:Luciferases / Q000378:metabolism; D000074443:Lysine Acetyltransferase 5; D008957:Models, Genetic; D000071447:N-Myc Proto-Oncogene Protein; D009447:Neuroblastoma / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism / Q000502:physiology*; D009707:Nucleosomes / Q000378:metabolism; D015513:Oncogene Proteins / Q000378:metabolism / Q000502:physiology*; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012319:RNA Polymerase II / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012190:Retroviridae / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016329:Sp1 Transcription Factor / Q000737:chemistry / Q000502:physiology*; D051705:Sp3 Transcription Factor; D014157:Transcription Factors / Q000737:chemistry / Q000502:physiology*; D014158:Transcription, Genetic
D018797:Cell Cycle Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D006657:Histones; D006877:Hydroxamic Acids; C459801:MYCN protein, human; D000071447:N-Myc Proto-Oncogene Protein; D009687:Nuclear Proteins; D009707:Nucleosomes; D015513:Oncogene Proteins; D011993:Recombinant Fusion Proteins; C497121:SP3 protein, human; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D051705:Sp3 Transcription Factor; C012589:trichostatin A; D008156:Luciferases; D000123:Acetyltransferases; D051548:Histone Acetyltransferases; C098489:KAT5 protein, human; D000074443:Lysine Acetyltransferase 5; D012319:RNA Polymerase II