Characterization of the RbBP6 gene in colon cancer

2677 The novel RbBP6 gene was identified in Chinese Hamster Ovary cell lines, where knockouts of the gene showed a high resistance to staurosporine-induced apoptosis. As irregularities in the apoptotic system aid in the progression of a wide range of medical conditions, including cancer, it is very beneficial to characterise promoters and inhibitors of this system, with the potential for improving therapies. Colon cancer is ranked as the third most common cancer worldwide, and is primarily a genetic disease, the progression of which can be efficiently studied. The RbBP6 gene is transcribed into two mRNAs, the 1.1kb and 6.1kb. Alternative splicing of the 6.1kb mRNA at exon 16 produces two variable proteins of 200kD, while the 1.1kb mRNA is expressed as a 13kD protein. The 1.1kb and 6.1kb mRNA consists of the DWNN region which is highly conserved throughout the eukaryotes. In addition, the 6.1kb mRNA contains a zinc finger, RING finger, Rb-binding and p53-binding region. Using Digoxigenin-labelled RNA probes, colorimetric and fluorescent in situ hybridisation studies localised each mRNA in the adenomatous glands, adenocarcinomas and in surrounding dysplastic tissues, with a clear upregulation in the diseased state compared to normal colon tissue. Immunocytochemistry using antibodies against the RbBP6 protein confirmed the upregulation of the protein in these structures and in lymphocytes. When correlated with TUNEL results which indicated sites of apoptosis, RbBP6 upregulation was shown to be directly proportional to increased levels of apoptosis. Ki-67 proliferation assay showed that there was indirect relationship between cell proliferation and RbBP6 localisation, while Bcl-2 assay showed RbBP6 to be upregulated in areas where this apoptosis suppressor was downregulated, and vice versa. These results suggest that RbBP6 is a pro-apoptotic gene as it is implied that it acts in the same direction as apoptosis. Localisation studies for the Helicobacter pylori protein was also performed as a significant relationship between this bacteria and colon cancer incidence has previously been reported. Immunocytochemistry showed high levels of the protein in the adenocarcinomas but not in the adenomatous glands, suggesting that H. pylori aids in the progression of colon cancer from the non-invasive to the invasive state but is not involved in the initiation of the cancer. No relationship could be ascertained between H. pylori and RbBP6.