| Budget Amount *help |
¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 1996: ¥400,000 (Direct Cost: ¥400,000)
Fiscal Year 1995: ¥2,000,000 (Direct Cost: ¥2,000,000)
|
|---|
| Research Abstract |
We first studies the expression of p21 and p16 in HeLa cell, a epidermoid carcinoma cells of the cervix, Ishikawa cell, a endometrial carcinoma cells, and 2780 cell, an ovarian epithelial carcinoma cell. All three cell lines steadily express mRNA and protein of p21 and p16. The expression levels of these cell lines, however, were relatively weak. Sequence analysis of p21 and p16 of these cell lines revealed that p16 and p21 of all cell line were wild types, though codon 31 of p21 mRNA of Ishikawa cell was arginine polymorphism. Next, p53 gene of 2780 was studied by using sequence analysis, showing deletion in its codon 178. The p53 of HeLA cells is inactivated by E6 protein of papilloma virus type 18. Therefore we supposed that p21 of these cell lines was independent of p53 and gene transfer of p21 into these two cell line may make a sense. We subcloned p21 gene into pMAMneo, a eucaryotic expression vector and pXT1 a retroviral vector and introduced into these cell lines. After G418 selection, we obtained sublines of HeLa and 2780 cells. These sublines showed apparent retardation of cell growth and attenuated telomerase activity. Cell cycle analysis using flowcytometry showed that the blockage from G0G1 to S is major cause of cell growth retardation. These results suggest that introduction of p21 gene into carcinoma cells of which p53 is inactivated by viral oncoproteins will suppress cell growth more efficiently than p53 itself.
|
