BRD7交互作用蛋白基因BRD2、BRD3在鼻咽癌组织中的表达

Expression of BRD7-interacting Proteins,BRD2 and BRD3,in Nasopharyngeal Carcinoma Tissues

背景与目的:BRD7基因是通过cDNA代表性差异分析获得的一个鼻咽癌(nasopharyngealcarcinoma,NPC)密切相关基因。采用酵母双杂交技术,已从人胎脑的cDNA文库中筛选到了两个与BRD7蛋白存在交互作用且包含溴区结构域(bromodomain)的蛋白BRD2、BRD3。本研究的目的在于进一步证实BRD7蛋白与BRD2、BRD3蛋白之间的交互作用,探讨BRD2、BRD3基因在鼻咽癌中的表达和作用模式。方法:将BRD2、BRD3基因分别与BRD7基因共转化酵母Y187后将菌落影印到尼龙膜,X-Gal检测酵母中报告基因LacZ的表达,推断BRD2、BRD3与BRD7蛋白之间的交互作用。RT-PCR方法检测BRD2、BRD3基因在正常鼻咽上皮组织和鼻咽癌活检组织中的差异表达以及在BRD7基因稳定转染的鼻咽癌细胞株(HNE1)中,BRD7基因的重表达对BRD2、BRD3基因表达水平的影响。结果:通过特异性酵母双杂交技术,发现酵母转化子的颜色呈蓝色,进一步证实BRD2、BRD3蛋白能分别与BRD7蛋白发生交互作用。RT-PCR结果显示,在鼻咽癌组织中,BRD2和BRD3基因表达下调或缺失;在BRD7基因稳定转染的HNE1细胞株中,随着BRD7基因表达的增强,BRD2、BRD3基因的表达也存在明显的上调。结论:BRD7蛋白能分别与BRD2、BRD3蛋白发生交互作用,且在mRNA水平存在一定程度的协同表达作用。

BACKGROUND & OBJECTIVE:BRD7 is a novel gene tightly associated with nasopharyngeal carcinoma(NPC) cloned by cDNA representational difference analysis (cDNA RDA). Two proteins,BRD2 and BRD3, including bromodomain and interacting with BRD7 protein had been screened from human fetal brain cDNA library by yeast two hybrid system. This study was designed to further identify the interactions of BRD2 and BRD3 with BRD7 respectively and to investigate the expression and action pattern of BRD2 and BRD3 in NPC. METHODS:BRD2 and BRD3 genes were respectively co transformed to yeast Y187 with BRD7 gene, then the yeast cotransformers were blotted to nylon membrane. And then the expression of report gene Lac Z by β Gal was determined and the interactions of BRD2 and BRD3 proteins with BRD7 protein were identified. Besides,reverse transcription polymerase chain reaction (RT PCR) was used to examine the expression of BRD2 and BRD3 genes in normal nasopharyngeal epithelium and NPC biopsies, and to detect the effect of re expression of BRD7 gene on the expression of BRD2 and BRD3 genes in HNE1 stably transfected BRD7 gene. RESULTS:The yeast transformers showed all blue by yeast two hybrid system, which further identified that BRD2 and BRD3 proteins could respectively interact with BRD7 protein. Down expression or loss of BRD2 and BRD3 genes were detectable in NPC biopsies. The expression levels of BRD2 and BRD3 were increasing with the re expression of BRD7 gene in HNE1 stably transfected with BRD7. CONCLUSION:BRD7 protein could respectively interact with proteins, BRD2 and BRD3, and BRD7 could up regulate the expression levels of BRD2 and BRD3 genes in mRNA level to some extent. Each of these three homolog proteins might be capable of forming heteromers with the others, which play important roles in the suppression of growth of NPC cells.