Farrall A L, Whitelaw M L
Discipline of Biochemistry, School of Molecular & Biomedical Science, University of Adelaide, Adelaide, South Australia, Australia.
Oncogene. 2009 Oct 15;28(41):3671-80. doi: 10.1038/onc.2009.228. Epub 2009 Aug 10.
The short isoform of single-minded 2 (SIM2s), a basic helix-loop-helix/PAS (bHLH/PAS) transcription factor, is upregulated in pancreatic and prostate tumours; however, a mechanistic role for SIM2s in these cancers is unknown. Microarray studies in prostate DU145 cells identified the pro-cell death gene, BNIP3 (Bcl-2/adenovirus E1B 19 kDa interacting protein 3), as a novel putative target of SIM2s repression. Further validation showed BNIP3 repression in several prostate and pancreatic carcinoma-derived cell lines with ectopic expression of human SIM2s. BNIP3 levels are enhanced in prostate carcinoma cells upon short interfering (si)RNA-mediated knockdown of endogenous SIM2s. Chromatin immunoprecipitation and promoter studies show that SIM2s represses BNIP3 through its activities at the proximal promoter hypoxia response element (HRE), the site through which the bHLH/PAS family member, hypoxia-inducible factor 1alpha (HIF1alpha), induces BNIP3. SIM2s attenuates BNIP3 hypoxic induction via the HRE, and increased hypoxic induction of BNIP3 occurs with siRNA knockdown of endogenous SIM2s in prostate PC3AR+ cells. BNIP3 is implicated in hypoxia-induced cell death processes. Prolonged treatment of PC3AR+ cells with hypoxia mimetics, DP and DMOG, confers hypoxia-induced autophagy, measured by enhanced LC3-II levels and SQSTM1/p62 turnover. We show that PC3AR+ cells expressing ectopic SIM2s have enhanced survival in these conditions. Induction of LC3-II and turnover of SQSTM1/p62 are attenuated in PC3AR+/SIM2s DMOG and hypoxia-treated cells, suggesting that SIM2s may attenuate autophagic cell death processes, perhaps through BNIP3 repression. These data show, for the first time, SIM2s cross-talk on an endogenous HRE. SIM2s' functional interference with HIF1alpha activities on BNIP3 may indicate a novel role for SIM2s in promoting tumourigenesis.
单 minded 2（SIM2）的短异构体是一种碱性螺旋-环-螺旋/ PAS（bHLH / PAS）转录因子，在胰腺和前列腺肿瘤中上调；然而，SIM2 在这些癌症中的作用机制尚不清楚。对前列腺 DU145 细胞的微阵列研究确定促细胞死亡基因 BNIP3（Bcl-2 /腺病毒 E1B 19 kDa 相互作用蛋白 3）是 SIM2 抑制的一个新的假定靶点。进一步验证表明，在几种前列腺和胰腺癌来源的细胞系中，随着人 SIM2 的异位表达，BNIP3 受到抑制。在内源性 SIM2 经短干扰（si）RNA 介导的敲低后，前列腺癌细胞中的 BNIP3 水平升高。染色质免疫沉淀和启动子研究表明，SIM2 通过其在近端启动子缺氧反应元件（HRE）处的活性抑制 BNIP3，bHLH / PAS 家族成员缺氧诱导因子 1α（HIF1α）通过该位点诱导 BNIP3。SIM2 通过 HRE 减弱 BNIP3 的缺氧诱导，并且在前列腺 PC3AR + 细胞中，随着内源性 SIM2 的 siRNA 敲低，BNIP3 的缺氧诱导增加。BNIP3 与缺氧诱导的细胞死亡过程有关。用缺氧模拟物 DP 和 DMOG 对 PC3AR + 细胞进行长时间处理，可导致缺氧诱导的自噬，通过增强的 LC3-II 水平和 SQSTM1 / p62 周转来衡量。我们表明，在这些条件下，表达异位 SIM2 的 PC3AR + 细胞具有更高的存活率。在 PC3AR + / SIM2s DMOG 和缺氧处理的细胞中，LC3-II 的诱导和 SQSTM1 / p62 的周转减弱，这表明 SIM2 可能通过抑制 BNIP3 来减弱自噬性细胞死亡过程。这些数据首次表明 SIM2 在内源性 HRE 上的相互作用。SIM2 对 HIF1α在 BNIP3 上的活性的功能干扰可能表明 SIM2 在促进肿瘤发生中具有新作用。
