Zhou Ming, Peng Cong, Nie Xin-Min, Zhang Bi-Cheng, Zhu Shi-Guo, Yu Ying, Li Xiao-Ling, Li Gui-Yuan
Cancer Research Institute, Xiangya School of Medicine, Central South University, Changsha, Hunan, 410078, PR China.
Ai Zheng. 2003 Feb;22(2):123-7.
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.
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 beta-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.
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.
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.
BRD7是通过cDNA代表性差异分析（cDNA RDA）克隆出的与鼻咽癌（NPC）密切相关的新基因。利用酵母双杂交系统从人胎脑cDNA文库中筛选出两种含溴结构域且与BRD7蛋白相互作用的蛋白，即BRD2和BRD3。本研究旨在进一步分别鉴定BRD2和BRD3与BRD7的相互作用，并探讨BRD2和BRD3在鼻咽癌中的表达及作用模式。
将BRD2和BRD3基因分别与BRD7基因共转化至酵母Y187，然后将酵母共转化体印迹至尼龙膜上。接着通过β -半乳糖苷酶检测报告基因Lac Z的表达，鉴定BRD2和BRD3蛋白与BRD7蛋白的相互作用。此外，采用逆转录 - 聚合酶链反应（RT - PCR）检测BRD2和BRD3基因在正常鼻咽上皮及鼻咽癌活检组织中的表达，并检测稳定转染BRD7基因的HNE1细胞中BRD7基因的重新表达对BRD2和BRD3基因表达的影响。
酵母双杂交系统显示酵母转化体均呈蓝色，进一步证实BRD2和BRD3蛋白均可分别与BRD7蛋白相互作用。在鼻咽癌活检组织中可检测到BRD2和BRD3基因表达下调或缺失。在稳定转染BRD7的HNE1细胞中，随着BRD7基因的重新表达，BRD2和BRD3的表达水平升高。
BRD7蛋白可分别与BRD2和BRD3蛋白相互作用，且BRD7在mRNA水平上可在一定程度上调BRD2和BRD3基因的表达水平。这三种同源蛋白可能均能彼此形成异源二聚体，在抑制鼻咽癌细胞生长中发挥重要作用。
