Epigenetic regulation of the human mucin gene MUC4 in epithelial cancer cell lines involves both DNA methylation and histone modifications mediated by DNA methyltransferases and histone deacetylases.
The human gene MUC4 encodes a transmembrane mucin, ligand of ErbB2, that is associated with pancreatic tumor progression. In the normal pancreas, MUC4 is not expressed, whereas activation of its expression is observed in the early steps of pancreatic carcinogenesis. The molecular mechanisms responsible for MUC4 gene activation are however still unknown. The MUC4 5'-flanking region being GC-rich and including two CpG islands, we hypothesized that epigenetic regulation may be involved and undertook to decipher the molecular phenomenons implied. By treating cancer cell lines with 5-aza-2'-deoxycytidine (5-aza) and trichostatin A (TSA), we were able to restore MUC4 expression in a cell-specific manner. We showed by bisulfite-treated genomic DNA sequencing and chromatin immunoprecipitation that methylation of five CpG sites and establishment of a repressive histone code at the 5'-untranslated region were associated with MUC4 silencing and impaired its activation by Sp1. Direct involvement of DNMT3A, DNMT3B, HDAC1, and HDAC3 was demonstrated by RNA interference and chromatin immunoprecipitation. Moreover, inhibition of histone deacetylation by TSA was associated with strong MUC4 repression in high-expressing cells. In conclusion, this work shows for the first time the importance of epigenetics in regulating MUC4 expression and may represent a new strategy to inhibit its expression in epithelial tumors.
Vincent
Audrey
A
Inserm, U837, Pl. de Verdun, 59045 Lille cedex, France.
Ducourouble
Marie-Paule
MP
Van Seuningen
Isabelle
I
D020121:5' Untranslated Regions; D001483:Base Sequence; D045744:Cell Line, Tumor; D018899:CpG Islands; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000378:metabolism; D019175:DNA Methylation; D000090205:DNA Methyltransferase 3A; D015254:DNA Modification Methylases / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D004273:DNA, Neoplasm / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D044127:Epigenesis, Genetic; D056284:Histone Deacetylase 1; D006655:Histone Deacetylases / Q000378:metabolism*; D006657:Histones / Q000378:metabolism; D006801:Humans; D055264:Mucin-4; D009077:Mucins / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D009375:Neoplasms, Glandular and Epithelial / Q000235:genetics* / Q000378:metabolism*; D010190:Pancreatic Neoplasms / Q000235:genetics* / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D016329:Sp1 Transcription Factor / Q000378:metabolism; D013274:Stomach Neoplasms / Q000235:genetics / Q000378:metabolism; D000097584:DNA Methyltransferase 3B; D000099186:Histone Deacetylase 3
D020121:5' Untranslated Regions; D004248:DNA (Cytosine-5-)-Methyltransferases; D000090205:DNA Methyltransferase 3A; D015254:DNA Modification Methylases; D017931:DNA Primers; D004273:DNA, Neoplasm; D056284:Histone Deacetylase 1; D006655:Histone Deacetylases; D006657:Histones; D055264:Mucin-4; D009077:Mucins; D034741:RNA, Small Interfering; D016329:Sp1 Transcription Factor; D000097584:DNA Methyltransferase 3B; D000099186:Histone Deacetylase 3; C000717967:DNMT3A protein, human; C524658:MUC4 protein, human; C110550:HDAC1 protein, human