摘要
目的探讨黑色素瘤细胞OCM-1A中表观遗传因素DNA甲基化对HLA-G分子表达的影响。方法利用DNA甲基化抑制剂5-aza-2′-deoxycytidine(ADC)处理HLA-G阴性黑色素瘤细胞OCM-1A,检测经不同浓度、不同时间的ADC处理后HLA-G基因及蛋白表达的变化。序列测定分析ADC处理前后HLA-G基因启动子区220bp片段的甲基化水平,RT-PCR检测HLA-GmRNA水平的变化,Westernblot检测细胞HLA-G分子合成总量以及FACS检测细胞表面HLA-G分子的表达水平。结果HLA-G阴性黑色素瘤细胞OCM-1A经ADC处理后,HLA-G基因启动子区的CpG甲基化程度明显降低,诱导HLA-G基因转录,HLA-G分子获得表达。HLA-G基因及蛋白的表达水平与处理时间相关,随着处理时间的延长,HLA-G表达量上升。结论OCM-1A细胞中,HLA-G基因的表达调控受DNA甲基化影响,ADC对HLA-G的表达调控具有时相性。
Objective To investigate the epigenefic mechanism of DNA methylation in regulation of HLA-G expression in a melanoma cell line OCM-1A. Methods The HLA-G negative elanoma cell line OCM-1A was treated with different concentrations of DNA methylation inhibitor 5-aza-2＇-deoxycytidine（ADC） at six particular time points. CpG methylation status in the promotor region of the HLA-G gene was evaluated with the sodium bisulfite sequencing. The HLA-G gene and protein expression was analyzed. RT-PCR was introduced to detect the HLA-G mRNA expression, Western blot and the flow cytometry were applied to measure HLA-G protein expression before and after the OCM-1A cells being treated with the ADC. Results After ADC treatment, the CpC, methylation status in the HLA-G promotor was dramatically decreased. HLA-G gene and protein were induced in a manner of time dependent after the ADC treatment in the OCM-1A cells. However, different concentrations of ADC reveals a similar effect in the study. Conclusions The epigenetic factor DNA methylation plays a vital role in the HLA-G expression regulation with a manner of time dependent in the melanoma ceil line OCM-IA.
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2006年第5期472-475,共4页
Chinese Journal of Microbiology and Immunology
