人鼻咽上皮细胞CYP2E1 cDNA克隆及序列分析

目的 :比较正常人胚鼻咽上皮 (HENE)细胞、二亚硝基哌嗪转化的人胚鼻咽上皮 (HENE)所建的细胞系( 742 9)、鼻咽癌细胞系 (HNE1)细胞色素P45 0 2E1(CYP2E1)mRNA表达序列 ,探讨CYP2E1在鼻咽癌变过程中的作用特点。方法 :采用RT PCR方法和DNA重组技术从HENE ,742 9,HNE13种细胞系中克隆的CYP2E1cDNA片段 ,测序并分析其结构改变特征。结果 :①与HENE 2E1比较 ,742 9 2E1存在 846位 (A→T)、90 1位 (A→G)两个点突变 ;HNE1 2E1存在 90 1位 (A→G)一个点突变。②与成人肝来源的CYP2E1CDs序列 (GenBank号为J0 2 843)比较 ,发现HNE1 2E1序列与其完全匹配 ;HENE 2E1序列存在 90 1位 (G→A)点突变。但上述突变对应部位氨基酸序列并无改变。结论 :鼻咽癌发生过程中CYP2E1基因cDNA序列仅存在少数无义突变 ,表现出其高度保守性。

Cloning and sequence analysis of human embryonic nasopharyngeal epithelial CYP2E1 cDNA

Objectives To investigate the role of CYP2E1 gene in chemical carcinogen-induced nasopharyngeal carcinogenesis and to provide new evidence about etiology and pathogenesis of nasopharyngeal carcinoma. Methods RT-PCR was used to clone the CYP2E1 gene in human embryonic nasopharyngeal epithelial (HENE) cell, the transformed nasopharyngeal epithelial cell line (7 429),and the nasopharyngeal carcinoma cell line (HNE1). The cloned segments were inserted into pGEM-T Easy vector to sequence by DNA recombination technique. Results In comparison with HENE-2E1 cDNA, there were two point mutations at positions 846(A to T) and 901(A to G) in 7 429-2E1 cDNA as well as only one point mutation at position 901(A to G) in HNE1-2E1 cDNA. In comparison with human (adult, ethanol-inducible) liver CYP2E1 gene (GenBank NO.J02843), HENE-2E1 cDNA had one point mutation at position 901(G to A). All these point mutations didn't affect the amino acid sequence. But no base change was found in HNE1-2E1 cDNA. Conclusion There are a few of base substitutions among HENE-2E1 cDNA, 7 429-2E1 cDNA,and HNE1 cDNA sequences.All these point mutations are synonymous mutation. The study reconfirms that the human CYP2E1 gene is relatively well conserved.