IDENTIFICATION OF GENES OVEREXPRESSED IN THE SQCC/Y1 HUMAN BUCCAL CARCINOMA CELL-LINE USING THE DIFFERENTIAL DISPLAY METHOD

Although several oncogenes and tumor suppressor genes have been suggested to be of relevance for the development of oral cancer, it is likely that additional genes are involved in this complex process. Therefore, in an attempt to isolate such genes, the aim of this study was to investigate changes in gene expression in human buccal carcinoma cells as compared to normal buccal epithelial cells, and identify mRNA overexpressed in the carcinoma cell line. The method of differential display of mRNA was used to isolate differentially expressed genes (Liang P et al, Science 257:967-971, 1992). A key step of this method, a polymerase chain reaction amplification, was optimized in terms of choice of thermostable DNA polymerase, annealing temperature, molar ratios and concentrations of primers. The comparative analysis of expression in tumor and normal buccal epithelial cells led to the isolation of three different mRNAs overexpressed in human oral carcinoma cells, as confirmed by Northern blot analysis. Cloning and sequence analysis revealed that these genes, which were termed OTEX as in Oral Tumor EXpressed, included a novel, previously not characterized, human gene, OTEX-1. OTEX-2 was identical to the gene coding for the L26 ribosomal protein, a protein known to be overexpressed also in other tumor cell types. OTEX-3 showed a perfect match to a sequence isolated during the human genome sequencing project, with a hitherto unknown function.