Re-expression of methylation-induced tumor suppressor gene silencing is associated with the state of histone modification in gastric cancer cell lines.
OBJECTIVE
To identify the relationship between DNA hyper-methylation and histone modification at a hyperme-thylated, silenced tumor suppressor gene promoter in human gastric cancer cell lines and to elucidate whether alteration of DNA methylation could affect histone modification.


METHODS
We used chromatin immunoprecipitation (ChIP) assay to assess the status of histone acetylation and methylation in promoter regions of the p16 and mutL homolog 1 (MLH1) genes in 2 gastric cancer cell lines, SGC-7901 and MGC-803. We used methylation-specific PCR (MSP) to evaluate the effect of 5-Aza-2'-deoxycytidine (5-Aza-dC), trichostatin A (TSA) or their combination treatment on DNA methylation status. We used RT-PCR to determine whether alterations of histone modification status after 5-Aza-dC and TSA treatment are reflected in gene expression.


RESULTS
For the p16 and MLH1 genes in two cell lines, silenced loci associated with DNA hypermethylation were characterized by histone H3-K9 hypoacetylation and hypermethylation and histone H3-K4 hypomethylation. Treatment with TSA resulted in moderately increased histone H3-K9 acetylation at the silenced loci with no effect on histone H3-K9 methylation and minimal effects on gene expression. In contrast, treatment with 5-Aza-dC rapidly reduced histone H3-K9 methylation at the silenced loci and resulted in reactivation of the two genes. Combined treatment with 5-Aza-dC and TSA was synergistic in reactivating gene expression at the loci showing DNA hypermethylation. Similarly, histone H3-K4 methylation was not affected after TSA treatment, and increased moderately at the silenced loci after 5-Aza-dC treatment.


CONCLUSIONS
Hypermethylation of DNA in promoter CpG islands is related to transcriptional silencing of tumor suppressor genes. Histone H3-K9 methylation in different regions of the promoters studied correlates with DNA methylation status of each gene in gastric cancer cells. However, histone H3-K9 acetylation and H3-K4 methylation inversely correlate with DNA methylation status of each gene in gastric cancer cells. Alteration of DNA methylation affects histone modification.
Meng
Chun-Feng
CF
Department of Surgical Oncology, The First Affiliated Hospital, China Medical University, No.155, Nanjing North Street, Heping District, Shenyang 110001, Liaoning Province, China.
Zhu
Xin-Jiang
XJ
Peng
Guo
G
Dai
Dong-Qiu
DQ
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism; D001374:Azacitidine / Q000494:pharmacology; D045744:Cell Line, Tumor; D002843:Chromatin / Q000378:metabolism; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation / Q000187:drug effects; D004273:DNA, Neoplasm / Q000235:genetics* / Q000378:metabolism; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D020868:Gene Silencing / Q000502:physiology*; D016147:Genes, Tumor Suppressor / Q000502:physiology*; D006657:Histones / Q000378:metabolism*; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D000070957:MutL Protein Homolog 1; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D013274:Stomach Neoplasms / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D025803:Tumor Suppressor Protein p14ARF / Q000235:genetics / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D002843:Chromatin; D004273:DNA, Neoplasm; D004791:Enzyme Inhibitors; D006657:Histones; D006877:Hydroxamic Acids; C501486:MLH1 protein, human; D009687:Nuclear Proteins; D025803:Tumor Suppressor Protein p14ARF; C012589:trichostatin A; D000070957:MutL Protein Homolog 1; D001374:Azacitidine