| Summary: | 碩士 === 國立臺灣海洋大學 === 生物科技研究所 === 98 === Though it is known that hypoxic microenvironments in solid tumor shape tumor progression, little is understood about how hypoxia regulates cancer cell proliferation. We have identified that Cited2, a novel transcriptional coactivator, is induced by hypoxia, and regulates hypoxic cell cycle progression in lung cancer. Immunoblotting and Q-PCR analysis showed that overexpression of HIF-2α but not HIF-1α promoted Cited2 expression. Knockdown of HIF-2α but not HIF-1αattenuated hypoxia-induced Cited2 level, suggesting that hypoxia induces Cited2 expression through a HIF-2α dependent pathway. Whereas overexpression of HIF-2α promoted cell proliferation as well as anchorage–independent cell growth, knockdown of Cited2 inhibited HIF-2α-mediated oncogenesis of lung cancer, suggesting that hypoxia-induced Cited2 is important for HIF-2α-mediated oncogenic properties. Consistent with these observations, cell cycle analysis showed that hypoxia induced G0/G1 cell arrest, and knockdown of Cited2 further increased hypoxia-mediated cell cycle arrest. In addition, overexpression of Cited2 promoted cell growth under the treatment of hypoxia-mimetic agents. These data support the notion that Cited2 regulates hypoxia-mediated cell cycle progression. Microarray analysis identified that Cited2 controlled E2F3, a key factor regulating G0/G1 cell cycle transition. Immunoblotting and Q-PCR analysis demonstrated that both hypoxia and HIF-2αinduced E2F3 expression, indicating E2F3 as a hypoxia inducible genes. Cited2-silencing attenuated hypoxia-induced E2F3 expression in lung cancer cells, suggesting that Cited2 regulates hypoxia-mediated E2F3 expression. It has been shown that HIF-2αpromotes cell proliferation through enhancing c-Myc transcription. Coimmunoprecipitation indicated that Cited2 interacted with HIF-2αand c-Myc. E2F3 promoter reporter analysis further showed that Cited2 enhanced HIF-2α/c-Myc-mediated transactivation of E2F3 promoter. Thus, our study provides a novel mechanism that shows how HIF-2α induces Cited2 expression, which further promotes HIF-2α c-Myc-mediated transcription in a feed-forward manner, leading to increased cell proliferation.
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