Overexpression of DNA damage-induced 45 α gene contributes to esophageal squamous cell cancer by promoter hypomethylation.
BACKGROUND
Environmental factors-induced dysfunction of esophageal squamous epithelium, including genomic DNA impairment and apoptosis, play an important role in the pathogenesis of esophageal squamous cell cancer. DNA damage-induced 45α (GADD45α) has been found promoting DNA repair and removing methylation marker, Therefore, in this study we will investigate whether GADD45α expression is induced and its mechanism in esophageal squamous cell cancer.


METHODS
Two human esophageal squamous cell lines (ESCC), ECA109 and KYSE510 were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum (FBS). Lipofectamine 2000 was used to transfect cells. mRNA level of GADD45α was measured by reverse transcription-quantitive PCR (RT-qPCR), protein level of GADD45α was detected by western blot and Immunohistochemistry. Global DNA methylation of tissue sample was measured using the Methylamp Global DNA Methylation Quantification Ultra kit (Epigentek Group) and promoter methylation was measured by bisulfite sequencing.


RESULTS
GADD45a mRNA and protein levels were increased significantly in tumor tissue than that in adjacent normal tissue. Hypomethylation of global genomic DNA and GADD45α promoter were found in ESCC. The cell sensitivity to Cisplatin DDP was decreased significantly in Eca109 and Kyse510 cells, in which GADD45α expression was down-regulated by RNA interference (RNAi). In addition, silence of GADD45a expression in ESCC cells inhibited proliferation and promoted apoptosis.


CONCLUSIONS
Overexpression of GADD45α gene is due to DNA hypomethylation in ESCC. GADD45α may be a protective factor in DDP chemotherapy for esophageal squamous cell carcinoma.
Department of Cardiothoracic Surgery, Second Xiangya Hospital of Central South University, Changsha, Hunan, PR China. wangbaoxiang615@sohu.com
Bao xiang
Bx
Wang
Bang Liang
BL
Yin
Bin
B
He
Chen
C
Chen
Ming
M
Zhao
Wei xing
Wx
Zhang
Zhen Kun
ZK
Xia
Yi zhi
Yz
Pan
Jing qun
Jq
Tang
Xin min
Xm
Zhou
Ni
N
Yin
D000328:Adult; D000368:Aged; D017209:Apoptosis; D002294:Carcinoma, Squamous Cell / Q000235:genetics* / Q000473:pathology; D018797:Cell Cycle Proteins / Q000235:genetics*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D019175:DNA Methylation; D004938:Esophageal Neoplasms / Q000235:genetics* / Q000473:pathology; D005260:Female; D015870:Gene Expression; D020868:Gene Silencing; D006801:Humans; D008297:Male; D008875:Middle Aged; D009687:Nuclear Proteins / Q000235:genetics*; D011401:Promoter Regions, Genetic; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D000097642:GADD45 Proteins
D018797:Cell Cycle Proteins; D009687:Nuclear Proteins; D012333:RNA, Messenger; C439070:GADD45A protein, human; D000097642:GADD45 Proteins