| Summary: | 碩士 === 國防醫學院 === 生物化學研究所 === 92 === p53 is identified as a tumor suppressor and more than 50% of human cancers p53’s function is lost via mutations in its gene. In normal or unstressed cells, the protein level of p53 remains low and inactive state. Upon exposing various stresses, p53 protein becomes stabilized and activated. The activated p53 protein can induce a variety of cellular responses such as growth arrest, apoptosis, and DNA repair. However, the exact molecular mechanism of p53 for its biological functions is still remains elusive.
We cloned and identified a p53-interacting protein, named pip by Sos recruitment system. We demonstrated that pip interacts with p53 in vitro and in vivo, however, the functional significant of this interaction is still quite limited. In order to understand the biological meaning of the interaction, we established adenovirus and retrovirus expression systems to overexpress p53 and pip protein in experimental cells. We would like to know how pip affects p53’s biological function.
From our data indicated when overexpress pip in mouse lymphoma cells EL4 by retrovirus (Retro Q) system, we observe pip inhibited cells growth. This phenomenon cause we can’t ask any of EL4 cells. On the other hand, we also established stably express pip human osteosarcoma cells by retrovirus. One is SaOS-2 cells which have no endogenous p53, another is U2OS cells which have wild-type p53. To employ the difference between SaOS-2 and U2OS, we treated various DNA damage agents alone or infected ad-p53 alone or both DNA damage agents and ad-p53 treatment for SaOS-2, U2OS cells and stably express pip SaOS-2 and U2OS cells. From the flow cytometer analysis, we hypothesize a critical molecular determinant in p53-induced cell cycle arrest and apoptosis. We find out pip’s function in osteosarcoma cells depend on p53 is activated or inactivated. When cells treated DNA damage agents, it can promote cell’s p53 activated. We observe pip can promote cell apoptosis and G2/M phase decrease. If the p53 is inactivated in cells, we observe pip reduces apoptosis and G1 phase increase. In conclusion, we provide complete experimental data to explain pip function may decide to cellular p53 is activated or inactivated, and p53 induced cells downstream biological effect is depend on pip existence.
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