NECAB3 Promotes Activation of Hypoxia-inducible factor-1 during Normoxia and Enhances Tumourigenicity of Cancer Cells.
NECAB3 Promotes Activation of Hypoxia-inducible factor-1 during Normoxia and Enhances Tumourigenicity of Cancer Cells.
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DOI：
10.1038/srep22784
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发表时间：
2016-03-07
影响因子：
4.6
通讯作者：
Sakamoto T
中科院分区：
文献类型：
--
作者：
Nakaoka HJ;Hara T;Yoshino S;Kanamori A;Matsui Y;Shimamura T;Sato H;Murakami Y;Seiki M;Sakamoto T
与大多数细胞不同，癌细胞激活缺氧诱导因子-1 （HIF-1）进行糖酵解，即使在正常氧水平或常氧条件下也是如此。因此，HIF-1在癌症治疗中是一个有吸引力的靶点。然而，虽然最近发现Mint3通过抑制HIF-1抑制剂、抑制HIF-1因子（FIH-1）在癌细胞和巨噬细胞中激活HIF-1，但在常氧条件下HIF-1的调节尚未得到很好的表征。在这项研究中，我们分析了Mint3结合蛋白来研究Mint3调节HIF-1的机制。以Mint3为诱饵进行酵母双杂交筛选，发现n端EF-hand钙结合蛋白3 （NECAB3）是通过Mint3调控HIF-1活性的新因子。NECAB3与min3的磷酸酪氨酸结合域结合，与min3和FIH-1形成三元配合物，并与min3在高尔基体共定位。在常氧癌细胞中，NECAB3的缺失降低了HIF-1靶基因的表达，并减少了糖酵解。结合Mint3但缺乏完整单加氧酶结构域的NECAB3突变体也抑制HIF-1的激活。通过表达shrna或产生显性阴性突变体抑制癌细胞中的NECAB3可降低致瘤性。综上所述，这些数据表明NECAB3是一个很有希望的癌症治疗新靶点。
Unlike most cells, cancer cells activate hypoxia inducible factor-1 (HIF-1) to use glycolysis even at normal oxygen levels, or normoxia. Therefore, HIF-1 is an attractive target in cancer therapy. However, the regulation of HIF-1 during normoxia is not well characterised, although Mint3 was recently found to activate HIF-1 in cancer cells and macrophages by suppressing the HIF-1 inhibitor, factor inhibiting HIF-1 (FIH-1). In this study, we analysed Mint3-binding proteins to investigate the mechanism by which Mint3 regulates HIF-1. Yeast two-hybrid screening using Mint3 as bait identified N-terminal EF-hand calcium binding protein 3 (NECAB3) as a novel factor regulating HIF-1 activity via Mint3. NECAB3 bound to the phosphotyrosine-binding domain of Mint3, formed a ternary complex with Mint3 and FIH-1, and co-localised with Mint3 at the Golgi apparatus. Depletion of NECAB3 decreased the expression of HIF-1 target genes and reduced glycolysis in normoxic cancer cells. NECAB3 mutants that binds Mint3 but lacks an intact monooxygenase domain also inhibited HIF-1 activation. Inhibition of NECAB3 in cancer cells by either expressing shRNAs or generating a dominant negative mutant reduced tumourigenicity. Taken together, the data indicate that NECAB3 is a promising new target for cancer therapy.