Abstract
Ataxia-telangiectasia (A-T) is a recessive genetic disease that progressively and degenerately affects a startling variety of body systems. Several epidemiological studies have found that patients with A-T heterozygotes tend to have higher risk to develop epithelial carcinoma, especially breast carcinomas. Although tumorigenic mechanisms to explainthe occurance of tissue specificity is not clear, the gene responsible for A-T (ataxia-telangiectasia mutated;ATM) has been identified recently, and it is believed that ATM plays a key role In tumor suppression this disease.In this study, we aimed at investigating how ATM down-stream ptoteins react againt ioizingradition when the ATM protein were down-regulated,and we focus on the differences among cells of tissue origins.The Tet-on gene expression system, using Doxycycline as a cofactor to control the gene expression, was used to generate an ATM antisense mRNA to suppress the expression of ATM protein. Immunoblotting was used to monitor possible functional alteration of ATM down-stream genes (p53, chk2, p95/NBS1).Our data shows that the ATM protein was successfully down regulated by adding increasing concentration of Doxycyclin in Tet-on gene expression system. however, no significantly functional differences of ATM down-stream genes was found in MCF-7 (Breast Carcinoma), HepG2 (Human Hepatocellular carcinoma), 293T (Human embryonic kidney cell) and Jurkat (human acute T-cell Leukemia). Only when the ATM protein was totally absent, the function of ATM down-stream protein was abolished.In conclusion, the outcome of using Tet-on system expressing antisense mRNA to inhibit the expression of ATM within the cells is good. When the ATM expression was down regulated, the expression of the downstream regulated proteins did not decrease significantly. This phenomenon could be explained as even low amount of ATM was expressed, but all function normally, therefore, the effect of down regulated ATM downstream proteins is slightly weaker than those not expressing antisense mRNA