Dissecting Nrf2-dependent HIF1a activation mechanism in arsenic-induced cancer stem-like cells

长期接触砷，尤指。无机三价砷(IAS)，一种人类致癌物， 由于自然发生在地壳或工作场所的工业环境中，一直与各种 癌症的类型，尤指肺癌。几十年来，人们已经知道有多种不同的机制 可能参与了IAS诱导的恶性转化。然而，国际会计准则是否以及如何诱导 非干细胞来源的肿瘤干细胞(CSCs)尚未被研究过。我们之前已经证明过 0.125~0.25ppbM(~9~18ppb)免疫荧光法连续处理人支气管上皮细胞 连续6个月诱导产生CSCs。除了茎的高表达外， 电路基因，包括Oct4、Sox2、myc、Klf4等，这些IAS诱导的CSCs也表现出独特的 以糖酵解活跃和线粒体氧化减少为特征的代谢模式 磷酸化(OXPHOS)。亲本细胞和IAS诱导的干细胞的CHIP-SEQ分析 揭示了组蛋白H3赖氨酸4三甲基化(H3K4me3)的浓集显著增加，以及 在茎和糖酵解的关键基因中，基因转录的活性表观遗传标记 CSCS的成员。为了了解IAS如何诱导这些代谢和表观遗传变化以及 随着CSCs的产生，我们最近还研究了IAS对Nrf2和Nrf2激活的能力 HIF1通过生化和芯片序列两种方法发出信号。数据表明，NRF2和 HIF1可作为代谢和表观遗传重编程的启动子，并获得 CSC功能。因此，我们假设ias激活的NRF2是hif1的上游激活因子。 与糖酵解有关的信号，以及随后的表观遗传重新编程和 CSCS。为了验证这一假设，将追求以下三个具体目标：Aim1，解剖Nrf2 激活途径，重点是IAS诱导的泛素化和Keap1的降解，以及 NRF2的内源性抑制物；目的2，研究NRF2依赖的HIF1的启动机制 IAS在细胞中的信号传递；以及目标3，了解IAS如何诱导NRF2-HIF1 信号将代谢从线粒体TCA循环转移到糖酵解，这与重新编程有关 组蛋白甲基化特征和CSCs的产生。我们预计来自 这项研究计划的完成将首次揭示NRF2和HIF1在IAS中的重要性- 诱导的CSCs，这可能被翻译成新的癌症治疗策略，通过靶向Nrf2- HIF1信令和CSCs。

Long-term exposure to arsenic, esp. the inorganic trivalent arsenic (iAs), a human carcinogen that occurs naturally in the earth's crust or work place due to industrial settings, has been linked to various types of cancers, esp. lung cancer. It has been known for decades that multiple different mechanisms might be involved in iAs-induced malignant transformation. However, whether and how iAs induces cancer stem-like cells (CSCs) from non-stem cells hadn’t been studied. We had previously shown that consecutive treatment of the human bronchial epithelial cells with 0.125 to 0.25 M (~9 to 18ppb) iAs for six months induced generation of the CSCs. In addition to the higher expression of the stemness circuit genes, including Oct4, Sox2, myc, Klf4, etc., these iAs-induced CSCs also exhibited a unique metabolic pattern featured with an active glycolysis and diminished mitochondrial oxidative phosphorylation (OXPHOS). ChIP-seq analysis of the parental cells and the iAs-induced CSCs revealed a substantial increase in the enrichment of histone H3 lysine4 trimethylation (H3K4me3), an active epigenetic marker for gene transcription, among genes critical for the stemness and glycolysis of the CSCs. To understand how iAs induces these metabolic and epigenetic changes and the generation of the CSCs, we recently also investigated the capability of iAs on the activation of Nrf2 and HIF1 signaling through both biochemical and ChIP-seq approaches. The data indicate that Nrf2 and HIF1 may serve as initiators for the metabolic and epigenetic reprograming, and the acquisition of the CSC features. Accordingly, we hypothesize that iAs-activated Nrf2 is an upstream activator for HIF1 signaling linked to glycolysis, and the subsequent epigenetic reprogramming and generation of the CSCs. To test this hypothesis, the following three specific aims will be pursued: Aim1, dissecting Nrf2 activation pathways with an emphasis on the iAs-induced ubiquitination and degradation of Keap1, the endogenous inhibitor of Nrf2; Aim 2, investigating mechanisms of Nrf2-dependent initiation of HIF1 signaling in the cells in response to iAs; and Aim 3, understanding how iAs-induced Nrf2-HIF1 signaling shifts the metabolism from mitochondrial TCA cycle to glycolysis that linked to reprogramming of the histone methylation profiles and the generation of CSCs. We expect that the data from the completion of this research plan will be the first to reveal the importance of Nrf2 and HIF1 in iAs- induced CSCs, which may be translated into new strategies of cancer therapy by targeting the Nrf2- HIF1 signaling and the CSCs.