Molecular cloning of a novel gene located on chromosome 3p25.3 and an analysis of its expression in nasopharyngeal carcinoma

Objective: To obtain the novel genes associated with human nasopharyngeal carcinoma (NPC) on chromosome 3p24-26. Methods: Twenty epithelial-derived expressed sequence tags (EST) were selected from chromosome 3p24-26 where loss of heterozygosity(LOH) frequently occurs in NPC tissues. Primers were designed based on the sequences of these ESTs. RT-PCR was used to amply their corresponding cDNA fragments from NPC cell line HNE1 and primary cultures of normal nasopharyngeal epithelial cells. The differential expression of two ESTs, T93093 and R41598, was confirmed by Northern blot. Then, expression of EST T93093 was further detected in 7 normal nasopharyngeal and 19 NPC biopsies. cDNA library screening was used to get its full cDNA sequence and the sequence of this novel gene was analyzed by bioinformatics. Results: Thirteen ESTs (T62511, N39155, N68660, R61275, T95314, R06143, H52697, H66521, AA128685, AA284537, N52379, AA054180, and H98090)showed the similar expression level and 5 ESTs (R00732, R07573, R98052, H91759, H17566) showed no expression in both types of cells. EST T93093 was down-expressed, whereas EST R41598 up-expressed in NPC HNE1 cells. The EST T93093 was also found to be down-expressed in 26. 3%(5/19) of NPC biopsies. The full length cDNA of this gene was obtained and named NAG-7, which is located at chromosome 3p25.3. Its 1677 bp full length cDNA has a potential open reading frame(ORF) predicting a 94 amino acid protein with a molecular weight of 11023.87 Dalton. Bioinformatics analysis of the NAG-7 gene shows that it is a transmembrane protein containing a protein kinase C(PKC) phosphorylation site and a myristyl site. It has no significant homology to any reported genes in database of GenBank (AF086709). Conclusion: NAG-7 is a novel gene down-expressed in NPC, which may be involved in the development of NPC.