Promoter hypermethylation-induced transcriptional down-regulation of the gene MYCT1 in laryngeal squamous cell carcinoma.
BMC Cancer
; 12: 219, 2012 Jun 06.
Article
em En
| MEDLINE
| ID: mdl-22672838
ABSTRACT
BACKGROUND:
MYCT1, previously named MTLC, is a novel candidate tumor suppressor gene. MYCT1 was cloned from laryngeal squamous cell cancer (LSCC) and has been found to be down-regulated in LSCC; however, the regulatory details have not been fully elucidated.METHODS:
Here, we sought to investigate the methylation status of the CpG islands of MYCT1 and mRNA levels by bisulfite-specific PCR (BSP) based on sequencing restriction enzyme digestion, reverse transcription and real-time quantitative polymerase chain reaction (RQ-PCR). The function of specific sites in the proximal promoter of MYCT1 in LSCC was measured by transient transfection, luciferase assays, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation assay (ChIP).RESULTS:
The results suggested hypermethylation of 12 CpG sites of the promoter in both laryngeal cancer tissues and the laryngeal cancer line Hep-2 cell. The hypermethylation of the site CGCG (-695 to -692), which has been identified as the c-Myc binding site, was identified in laryngeal cancer tissues (59/73) compared to paired mucosa (13/73); in addition, statistical analysis revealed that the methylation status of this site significantly correlated with cancer cell differentiation(p < 0.01). The mRNA level of MYCT1 increased in Hep-2 cells treated with 5-aza-C (p < 0.01). The luciferase activity from mutant transfectants pGL3-MYCT1m (-852/+12, mut-695-C > A, mut-693-C > G) was significantly reduced compared with the wild type pGL3-MYCT1 (-852/+12), while the luciferase activity from wild transfectants pGL3-MYCT1 (-852/+12) rose after 5-aza treatment in Hep-2 cells. Finally, EMSA and ChIP confirmed that the methylation of the CGCG (-695 to -692) site prevented c-Myc from binding of the site and demethylation treatment of the 5' flanking region of MYCT1 by 5-aza induced the increased occupation of the core promoter by c-Myc (p < 0.01).CONCLUSION:
In summary, this study concluded that hypermethylation contributed to the transcriptional down-regulation of MYCT1 and could inhibit cancer cell differentiation in LSCC. DNA methylation of the CGCG site (-695 to -692) of MYCT1 altered the promoter activity by interfering with its binding to c-Myc in LSCC. Epigenetic therapy of reactivating MYCT1 by 5-aza should be further evaluated in clinical trails of LSCC.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Proteínas Nucleares
/
Carcinoma de Células Escamosas
/
Regulação Neoplásica da Expressão Gênica
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Neoplasias Laríngeas
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Regiões Promotoras Genéticas
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Metilação de DNA
Limite:
Humans
Idioma:
En
Revista:
BMC Cancer
Assunto da revista:
NEOPLASIAS
Ano de publicação:
2012
Tipo de documento:
Article