Expression of ABCG2 (BCRP) is regulated by Nrf2 in cancer cells that confers side population and chemoresistance phenotype.
Expression of ABCG2 (BCRP) is regulated by Nrf2 in cancer cells that confers side population and chemoresistance phenotype.
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DOI：
10.1158/1535-7163.mct-10-0108
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发表时间：
2010-08
影响因子：
5.7
通讯作者：
Biswal S
中科院分区：
文献类型：
--
作者：
Singh A;Wu H;Zhang P;Happel C;Ma J;Biswal S
ATP结合盒，亚家族G，成员2（ABCG 2）在正常细胞和癌细胞中表达，并且在侧群（SP）形成和外源性物质和药物的流出中起关键作用。nrf 2是一种氧化还原敏感转录因子，在非小细胞肺癌细胞中组成性激活后，上调了参与氧化还原平衡、谷胱甘肽代谢和药物解毒的广泛基因，这些基因有助于化疗耐药性和致瘤性。本研究探讨了ABCG 2的Nrf 2依赖性表达机制及其在多药耐药表型中的作用。对ABCG 2的5 '启动子侧翼区的计算机分析鉴定了在-431 bp至-420 bp处的抗氧化反应元件。使用荧光素酶报告基因测定的详细启动子分析表明，在-431 bp至-420 bp处的抗氧化反应元件（ARE）对于Nrf 2介导的肺癌细胞中的表达是关键的。电泳迁移率变动分析（EMSA）和染色质免疫沉淀（ChIP）分析表明，Nrf 2与ABCG 2 ARE元件在-431 ~-420 bp处相互作用。通过短发夹RNA破坏肺癌和前列腺癌细胞中的Nrf 2表达，减弱了ABCG 2转录物和蛋白质的表达，并显著降低了Nrf 2耗尽的癌细胞中的SP分数。此外，这些Nrf 2敲低细胞中ABCG 2的耗尽水平使它们对米托蒽醌和拓扑替康敏感，这两种化疗药物主要由ABCG 2解毒。正如预期的那样，Nrf 2 cDNA在肺上皮细胞中的过表达导致ABCG 2表达增加和SP分数增加2倍。因此，Nrf 2介导的ABCG 2表达调节维持SP分数并赋予化学抗性。
ATP-binding cassette, sub-family G, member 2 (ABCG2) is expressed in both normal and cancer cells, and plays a crucial role in the side population (SP) formation and efflux of xenobiotics and drugs. Nrf2, a redox sensing transcription factor, upon constitutive activation in non-small-cell lung cancer cells up-regulates a wide spectrum of genes involved in redox balance, glutathione metabolism, and drug detoxification that contribute to chemoresistance and tumorigenecity. This study examined the mechanism underlying Nrf2-dependent expression of ABCG2 and its role in multidrug resistance phenotype. In silico analysis of the 5’-promoter flanking region of ABCG2 identified an antioxidant response element at -431 bp to -420 bp. A detailed promoter analysis using luciferase reporter assays demonstrated that antioxidant response element (ARE) at -431 bp to -420 bp is critical for the Nrf2-mediated expression in lung cancer cells. Electrophoresis mobility shift assays (EMSA) and chromatin-immunoprecipitation (ChIP) assays revealed that Nrf2 interacts with ABCG2 ARE element at -431 bp to -420 bp in vitro and in vivo. Disruption of Nrf2 expression in lung cancer and prostate cancer cells, by short hairpin RNA, attenuated the expression of ABCG2 transcript and protein and dramatically reduced the SP fraction in Nrf2-depleted cancer cells. Moreover, depleted levels of ABCG2 in these Nrf2-knockdown cells sensitized them to mitoxantrone and topotecan, two chemotherapy drugs detoxified mainly by ABCG2. As expected, overexpression of Nrf2 cDNA in lung epithelial cells led to an increase in ABCG2 expression and a 2-fold higher SP fraction. Thus, Nrf2-mediated regulation of ABCG2 expression maintains SP fraction and confers chemoresistance.