Preliminary Study On The Role Of BRD7 Gene In Nasopharyngeal Carcinoma Cells

Nasopharyngeal carcinoma (NPC) is a common cancer in South China and Southeast Asia , but is rare in other parts of the world. The tumorigenesis of NPC is characteristic of obvious family history and genomic allelic imbalance. Significant loss of heterozygosity at 3p, 9p,llq, 13q and 14q in nasopharyngeal carcinoma (NPC) had been detected by comparative genomic hybridization (CGH) and genome-wide analysis of genetic alteration with microsatellite allelotyping. A novel NPC related gene, BRD7, was isolated through cDNA representational difference analysis (RDA). GenBank accession number is AF 152604.Our previous studies showed that the BRD7 gene was down-regulated in Nasopharyngeal carcinoma (NPC) cells and NPC biopsy. Three coding-region single nucleotide polymorphisms (cSNPs) were detected in BRD7 cDNA sequence. In addition, six BRD7-interacting proteins (EIPs) were identified by yeast two-hybrid system. These proteins included Bromodomain-containing 3 protein (BRD3), Bromodomain-containing 2 protein (BRD2), IkB kinase-beta , KIAA1375 protein, interleukin 7 ( IL-7) and the delta 1 subunit of adaptor-related protein complex 3. In this study , we further investigated the function of BRD7 in pathogenesis ofNPC and its possible molecular mechanism.Five kinds of proteins homologous to BRD7 of bromodomain were selected through Homology-based sequence analyses. These proteins have the similar structure containing four a helices (Z, A, B and C), and "a hydrophobic pocket" which has the ability to recognize acetylated histone peptide. So, we predicted that the bromodomain of BRD-7 may have the similarity structure. In addition, BRD7 gene expression was detected in brain, lung, liver, muscle, kidney, prostate and in other normal tissues by virtual Northern. The virtual Northern resulted in mainly similar results with laboratory assay.To elucidate the mechanism of BRD7 effect on carcinogenesis in NPC, BRD7 gene was transfected into BRD7 down-expressed HNE1 cell by liposome and a stable cell line overexpressing BRD7 was established. RT-PCR and Northern blot were performed to detect the overexpression of BRD7 in transfected cell.To confirm the effect of BRD7 on the malignant phenotype of HNE1 cell line, three assays, including growth curves ,colony formation and tumor xenografts in nude mice were performed. The results showed that compared with HNE1 and HNE1 cells transfected with pcDNA3.1(+) empty vector, BRD7 transfected cells resulted in declined growth curve, fewer colonies formation, and development of a smaller tumor size. Pathological analysis showed that xenograft were low differentiatesquamous cell carcinoma. These results indicated that the BRD7 can play negative role in the progression of NPC and may partly reverse malignant phenotype of NPC cell line.To determine whether BRD7 gene have effect on cell cycle and apoptosis, the flow cytometry (FCM) was performed to detect the distribution of cell cycle change and expression of cyclins in BRD7 transfected HNE1 cells. The results indicated that the overexpression of BRD7 could delay the progression of GO/G1 to S phase in cell cycle, and the expression cyclinDl and B were significantly decreased, while it had no effect on the apoptosis of cells.In summary, the BRD7 gene acted as a candidate of tumor suppressor gene with NPC. The overexpression of BRD7 can partly reverse malignant phenotype of NPC cell line. The suppression effect of BRD7 on NPC tumorigenesis my be achieved by recognizing acetylated histone peptide through their motif-bromodomain, then modulating gene transcription by taking part in histone acetylating and chromosome remodeling, finally influencing signal-transduction pathways.