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Activation of RASSF2 by p300 Induces S/G2 Cell Cycle Arrest in Gastric Cancer Cell HGC-27

Author: PanYunHui
Tutor: LiXiaoXue
School: Northeast Normal University
Course: Genetics
Keywords: RASSF2 p300 S/G2 cell cycle arrest DNA demethylation histone acetylation
CLC: R735.2
Type: Master's thesis
Year: 2011
Downloads: 35
Quote: 0
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Abstract


Tumor suppressor gene RASSF2 is usually silenced by DNA hypermethylation in gastric cancer cells. The evidences have showed that histone acetyltransferase p300 was able to induce late apoptosis by activation of RASSF2 in the gastric cancer cells SGC-7901. It is known that RASSF2 was able to induce apoptosis and cell cycle. However, it remains unclear if activation of RASSF2 by p300 can induce cell cycle arrest in gastric cancer cells. In this study, the results of colony formation showed that p300 was able to inhibit cell proliferation in gastric cancer cells HGC-27, and this inhibition can be partially reversed by RASSF2 siRNA. Western Blot and RT-PCR results confirmed that p300 can upregulate RASSF2 gene expression in the HGC-27 cells. So in HGC-27 cells, p300 inhibits cell proliferation by activating RASSF2 gene expression.The flow cytometric analysis (FACS) revealed that overexpression of p300 or inhibition of RASSF2 expression can increase the number of cells in S phase, while decrease the number of cells in G2/M. When co-trasfected with p300 overexpression plasmid and RASSF2 siRNA,the cell numbers in S and G2/M phase were similar to the control. The results of Western Blot showed that the activation RASSF2 by p300 resulted in upregulation of p21, and the ratio of phosphorylation pRb to no-phosphorylation pRb in HGC-27 cells. And the expression level of phosphorylated p38 was decreased when the expression of RASSF2 was promoted by p300. These results suggested that the activation of RASSF2 by p300 induced cell cycle arrest in S/G2 through changing the protein levels of some important cell cycle regulators.In addition, chromatin immunoprecipitation (ChIP) assays showed that p300 increased the acetylation levels of histone H3 and H4 at Sp1 binding sites and the CpG islands of RASSF2 promoter. MSP and BSP results showed the activation of RASSF2 by p300 is related to CpG island demethylation. These data suggested that histone acetylation and DNA demethylation are important in upregualtion of RASSF2 transcription by p300.

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CLC: > Medicine, health > Oncology > Gastrointestinal Cancer > Gastric neoplasms
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