Epigenetic mechanisms for silencing glutathione S-transferase m2 expression by hypermethylated specificity protein 1 binding in lung cancer.
BACKGROUND
Glutathione S-transferases M2 (GST-M2) is a detoxifying enzyme. Low expression levels of GST-M2 have been detected in lung cancer cells. However, little is known about the regulation of GST-M2 in lung cancer cells. In this study, the authors investigated the epigenetic regulatory mechanisms of GST-M2 in lung cancer cells.


METHODS
The authors evaluated the promoter methylation of GST-M2 in lung cancer cells after treatment with the DNA methyltransferase (DNMT) inhibitor 5'-aza-2'-deoxycytidine (5'-aza-dC). Reporter activity assays, chromatin immunoprecipitation (ChIP), electrophoretic mobility-shift assays, and small interfering RNA (siRNA) assays were used to determine whether the methylation of specificity protein 1 (Sp1) affected binding to the GST-M2 promoter or regulated GST-M2 transcription. Real-time polymerase chain reaction was used to determine GST-M2 and DNMT-3b messenger RNA levels in 73 nonsmall cell lung cancer (NSCLC) tissues.


RESULTS
GST-M2 expression was restored after treatment with 5'-aza-dC in lung cancer cells. GST-M2 exhibited high frequency of promoter hypermethylation in lung cancer cells and NSCLC tumor tissues. CpG hypermethylation abated Sp1 binding to the GST-M2 promoter in lung cancer. Knockdown of Sp1 in normal lung cells reduced GST-M2 expression, and silencing of DNMT-3b increased GST-M2 expression in lung cancer cells. In addition, DNMT-3b expression was significantly higher in lung tumors with low levels of GST-M2 expression than in lung tumors with high levels of GST-M2 expression, especially among women and among patients who had stage I disease.


CONCLUSIONS
Epigenetic silencing of GST-M2 was distinguished from Sp1-mediated GST-M2 transcriptional expression. The authors concluded that this represents a mechanism that leads to decreased expression of GST-M2 in lung cancer cells.
Institute of Medical and Molecular Toxicology, Chung Shan Medical University, Taichung, Taiwan, Republic of China.
Sheau-Chung
SC
Tang
Ming-Fang
MF
Wu
Ruey-Hong
RH
Wong
Yu-Fan
YF
Liu
Lee-Chun
LC
Tang
Chien-Hung
CH
Lai
Chung-Ping
CP
Hsu
Jiunn-Liang
JL
Ko
D000368:Aged; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics*; D045744:Cell Line, Tumor; D019175:DNA Methylation; D000077209:Decitabine; D004791:Enzyme Inhibitors / Q000494:pharmacology; D044127:Epigenesis, Genetic; D005260:Female; D005982:Glutathione Transferase / Q000037:antagonists & inhibitors / Q000235:genetics*; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics*; D008297:Male; D016329:Sp1 Transcription Factor / Q000235:genetics*
D004791:Enzyme Inhibitors; D016329:Sp1 Transcription Factor; D000077209:Decitabine; D005982:Glutathione Transferase; C412602:glutathione S-transferase Mu 2; D001374:Azacitidine