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背景与目的:染色体3p21.3区域的杂合性丢失是鼻咽癌细胞中高发和早期的细胞遗传学变异,提示缺失区域内可能存在与鼻咽癌发生相关的抑癌基因。BLU基因定位于3p21.3,蛋白质结构中含有MYND功能域。已有研究表明,BLU基因在鼻咽癌细胞中发生高频率的启动子甲基化和表达缺失。本研究构建了野生型BLU基因和其突变体的表达载体,分析BLU基因对鼻咽癌细胞恶性增殖的可能抑制作用。方法:用RT-PCR方法钓取BLU基因全长cDNA;用定点突变技术分别构建缺失MYND结构域的突变体、携带Ser402Phe点突变及缺失第405位Cys和第406位Ser的突变体、以及携带Gly160Arg点突变的BLU基因突变体的表达载体。将野生型BLU基因和MYND缺失型突变体转染鼻咽癌细胞CNE1和CNE2,用TUNEL方法检测、观察细胞凋亡情况;通过细胞计数和克隆形成实验分析稳定转染细胞的生长特性;分析稳定转染BLU基因的载体对CNE2细胞裸鼠致瘤性的影响。结果:瞬时转染野生型或MYND缺失突变型BLU基因不能诱导CNE2细胞凋亡;外源表达BLU基因对CNE2的细胞凋亡以及对CNE1、CNE2的细胞增殖和克隆形成能力均无明显影响;BLU基因对CNE2细胞的裸鼠成瘤能力也没有明显抑制作用。结论:尽管BLU基因在鼻咽癌中发生高频率变异,但其对鼻咽癌细胞的恶性增殖并没有明显抑制作用,因此该基因
BACKGROUND AND PURPOSE: Loss of heterozygosity on chromosome 3p21.3 is a high incidence and early cytogenetic variation in nasopharyngeal carcinoma cells. It suggests that there may be tumor suppressor genes associated with the occurrence of nasopharyngeal carcinoma in the absence of this gene. The BLU gene is located at 3p21.3, and the protein structure contains the MYND domain. Studies have shown that BLU gene in nasopharyngeal carcinoma cells with high frequency of promoter methylation and lack of expression. In this study, we constructed the expression vector of wild type BLU gene and its mutants, and analyzed the possible inhibitory effect of BLU gene on the malignant proliferation of nasopharyngeal carcinoma cells. Methods: The full-length cDNA of BLU gene was obtained by RT-PCR. The mutated MYND domain, the point mutation Ser402Phe, the Cys at position 405 and the Ser at position 406 were deleted by site-directed mutagenesis. Gly160Arg Point Mutant BLU Gene Mutant Expression Vector. The wild-type BLU gene and MYND deletion mutant were transfected into nasopharyngeal carcinoma cells CNE1 and CNE2, and detected by TUNEL method to observe the cell apoptosis; cell growth and stability were analyzed by cell counting and clonogenic assay; Effect of BLU gene transfection on tumorigenicity of CNE2 cells in nude mice. Results: Transfection of wild type or MYND deletion mutant BLU gene could not induce apoptosis of CNE2 cells. Exogenous expression of BLU gene had no effect on CNE2 cell apoptosis and CNE1 and CNE2 cell proliferation and clonality. BLU Gene CNE2 cells in nude mice tumorigenic ability did not significantly inhibit the role. Conclusion: Although BLU gene has high frequency mutation in nasopharyngeal carcinoma, it has no obvious inhibitory effect on the malignant proliferation of nasopharyngeal carcinoma cells. Therefore,