Stathmin, a microtubule-destabilizing phosphoprotein, is highly expressed in ovarian cancer, but the pathophysiological significance of this protein in ovarian carcinoma cells remains poorly understood. This study reports the involvement of stathmin in the mTOR/HIF-1α/VEGF pathway in ovarian clear cell adenocarcinoma (CCA) during hypoxia. HIF-1α protein and VEGF mRNA levels were markedly elevated in RMG-1 cells, a CCA cell line, cultured under hypoxic conditions. Rapamycin, an inhibitor of mTOR complex 1, reduced the level of HIF-1α and blocked phosphorylation of ribosomal protein S6 kinase 1 (S6K), a transcriptional regulator of mTOR, demonstrating that hypoxia activates mTOR/S6K/HIF-1α signaling in CCA. Furthermore, stathmin knockdown inhibited hypoxia-induced HIF-1α and VEGF expression and S6K phosphorylation. The silencing of stathmin expression also reduced Akt phosphorylation, a critical event in the mTOR/HIF-1α/VEGF signaling pathway. By contrast, stathmin overexpression upregulated hypoxia-induced HIF-1α and VEGF expression in OVCAR-3 cells, another CCA cell line. In addition, suppression of Akt activation by wortmannin, a phosphoinositide 3-kinase (PI3K) inhibitor, decreased HIF-1α and VEGF expression. These results illustrate that regulation of HIF-1α through the PI3K/Akt/mTOR pathway is controlled by stathmin in CCA. Our findings point to a new mechanism of stathmin regulation during ovarian cancer. 1. Introduction Most ovarian cancers are believed to arise from epithelial cells residing on the outer surface of the ovary. Histologically, human ovarian cancers are classified as serous cyst, clear cell (CCA), and endometrioid adenocarcinomas [1–3]. CCA accounts for 20% of ovarian cancers and 25% of all surface epithelial tumors. Because no symptoms are present during early stages of ovarian cancer, its diagnosis is usually delayed. This has contributed to an increase in the number of individuals with CCA in Japan. CCA is also resistant to chemotherapy; thus, it associates with a poor prognosis. Tumors often express vascular endothelial growth factor (VEGF) in response to local hypoxia [3, 4]. VEGF expression is an indicator of angiogenesis, and cancer cell proliferation and invasion [3–7]. Increased VEGF expression can also stimulate neovascularization and contribute to tumor growth. VEGF expression in different tissues is regulated by hypoxia inducible factor (HIF)-1α [8]. Hypoxia inhibits the hydroxylation of HIF-1α and its subsequent proteasomal degradation, resulting in the translocation of HIF-1α into the nucleus and in the transcription
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