In vivo transcriptional regulation of N-Myc target genes is controlled by E-box methylation.
N-Myc is a transcription factor that forms heterodimers with the protein Max and binds gene promoters by recognizing a DNA sequence, CACGTG, called E-box. The identification of N-myc target genes is an important step for understanding N-Myc biological functions in both physiological and pathological contexts. In this study, we describe the identification of N-Myc-responsive genes through chromatin immunoprecipitation and methylation-sensitive restriction analysis. Results show that N-Myc is a direct regulator of several identified genes, and that methylation of the CpG dinucleotide within the E-box prevents the access of N-Myc to gene promoters in vivo. Furthermore, methylation profile of the E-box within the promoters of EGFR and CASP8, two genes directly controlled by Myc, is cell type-specific, suggesting that differential E-box methylation may contribute to generating unique patterns of Myc-dependent transcription. This study illuminates a central role of DNA methylation in controlling N-Myc occupancy at gene promoters and modulating its transcriptional activity in cancer cells.
Perini
Giovanni
G
Department of Biology, University of Bologna, Via Selmi 3, 40126 Bologna, Italy.
Diolaiti
Daniel
D
Porro
Antonio
A
Della Valle
Giuliano
G
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000378:metabolism; D015152:Blotting, Northern; D053181:Caspase 8; D020169:Caspases / Q000235:genetics; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D019175:DNA Methylation; D017931:DNA Primers; D024721:E-Box Elements / Q000235:genetics / Q000502:physiology*; D024202:Electrophoretic Mobility Shift Assay; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D018773:Genes, erbB-1 / Q000235:genetics; D006801:Humans; D015151:Immunoblotting; D008156:Luciferases; D011401:Promoter Regions, Genetic / Q000235:genetics; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism*; D012097:Repressor Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D017931:DNA Primers; C106465:MNT protein, human; D016271:Proto-Oncogene Proteins c-myc; D012097:Repressor Proteins; D014157:Transcription Factors; D008156:Luciferases; C505530:CASP8 protein, human; D053181:Caspase 8; D020169:Caspases