MECHANISM OF HEXAVALENT CHROMIUM CARCINOGENESIS ROLE OF LONG NON-CODING RNA DYSREGULATION

项目摘要/摘要 六价铬[Cr(VI)]是一种常见的、公认的环境致癌物，会导致肺和其他 癌症，然而，铬(VI)致癌的机制仍然难以捉摸。先前的铬(VI)致癌作用 研究主要集中在其遗传毒性效应上。然而，研究表明，接触铬(VI)也会导致非 遗传毒性效应，如DNA甲基化和组蛋白失调所证明的表观遗传学变化 翻译后修饰。虽然这些观察为研究铬(VI)的致癌作用开辟了新的方向，但它 铬(VI)引起的表观遗传失调如何在铬(VI)致癌中起作用仍有待确定。 此外，人们对非编码RNA(NcRNAs)的作用知之甚少，尤其是长的非编码RNA (LncRNAs)，另一种调控基因表达的表观遗传机制，在铬(VI)致癌中。最新研究 表明lncRNAs是新出现的基因表达的关键调节因子，并在癌症中发挥关键作用。的目标是 本研究旨在探讨铬(VI)的致癌机制，重点探讨lncRNA异常调控的作用。 我们的初步研究发现：(1)慢性铬(VI)暴露增加了lncRNA ABHD11-AS1的表达， 在铬(VI)诱导的细胞转化、肿瘤干细胞(CSC)样特性和 肿瘤发生；(Ii)慢性铬(VI)暴露下调miR-182-5p的表达；但 ABHD11-AS1增加miR-182-5P水平。此外，在铬(VI)转化细胞中过表达miR-182-5p 显著减少它们转化的表型，表现出ABHD11-AS1基因敲除的效果；(Iii) Rho GTPase rac1和ERK1/2 MAPK在铬(VI)转化细胞中高度激活；(Iv) 在铬(VI)转化的细胞中，致癌的RAC-ECFTiam1显著上调；但过表达miR- 182-5P或敲除ABHD11-AS1可显著降低Tiam1的水平；(V)PARP-1的表达水平 DNA修复基因是基因表达的关键调控基因，在铬(VI)转化的细胞中显著上调； (Vi)在铬(VI)转化的细胞中，蛋白质PAR化水平显著增加；抑制PARP-1或 PARP-1基因的敲除显著降低了蛋白PAR化和ABHD11-AS1的水平 铬(VI)转化细胞的表型。基于文献回顾和我们的小说初步数据，我们的中心 假设是：“慢性铬(VI)暴露-上调的lncRNA ABHD11-AS1海绵miR-182-5p并导致其 下调，导致致癌的RAC-全环基金Tiam1促进铬(VI)水平增加 致癌作用“。提出了三个目标：目标1将确定慢性铬(VI)暴露的机制 上调ABHD11-AS1的表达。目标2将证明ABHD11-AS1海绵miR-182-5P导致其下降- 调节和促进铬(VI)暴露诱导的CSC样特性、细胞转化和肿瘤发生。和 目标3将证明miR-182-5p的下调会增加致癌的RAC-Global Tiam1的水平 促进铬(VI)暴露诱导的CSC样特性、细胞转化和肿瘤发生。Tiam1淘汰赛 小鼠将被用来证明Tiam1在铬(VI)诱导的肺癌发生中起重要作用。

PROJECT SUMMARY/ABSTRACT Hexavalent chromium [Cr(VI)] is a common and well-recognized environmental carcinogen causing lung and other cancers, however, the mechanism of Cr(VI) carcinogenesis remains elusive. Previous Cr(VI) carcinogenesis studies mostly focus on its genotoxic effects. However, studies showed that Cr(VI) exposure also causes non- genotoxic effects such as epigenetic changes as evidenced by dysregulations in DNA methylation and histone posttranslational modifications. While these observations open new directions for studying Cr(VI) carcinogenesis, it remains to be determined how Cr(VI)-caused epigenetic dysregulations contribute to Cr(VI) carcinogenesis. Moreover, very little is known about the role of non-coding RNAs (ncRNAs) especially the long non-coding RNAs (lncRNAs), another epigenetic mechanism regulating gene expression, in Cr(VI) carcinogenesis. Recent studies show that lncRNAs are emerging key regulators of gene expression and play critical roles in cancer. The goal of this study is to investigate the mechanism of Cr(VI) carcinogenesis focusing on the role of lncRNA dysregulation. Our preliminary studies found: (i) Chronic Cr(VI) exposure increases the expression of the lncRNA ABHD11-AS1, which contributes causally to Cr(VI)-induced cell transformation, cancer stem cell (CSC)-like property and tumorigenesis; (ii) Chronic Cr(VI) exposure down-regulates the expression of miR-182-5p; but knockdown of ABHD11-AS1 increases the level of miR-182-5p. Moreover, overexpressing miR-182-5p in Cr(VI)-transformed cells significantly reduces their transformed phenotypes, phenocopying the effect of ABHD11-AS1 knockdown; (iii) The Rho GTPase Rac1 and Erk1/2 MAPK are highly activated in Cr(VI)-transformed cells; (iv) The expression level of the oncogenic Rac-GEF Tiam1 is significantly up-regulated in Cr(VI)-transformed cells; but overexpressing miR- 182-5p or knockdown of ABHD11-AS1 greatly decrease Tiam1 level; (v) The expression level of PARP-1, a critical DNA repair gene and a key regulator of gene expression, is significantly up-regulated in Cr(VI)-transformed cells; (vi) The level of protein PARylation is drastically increased in Cr(VI)-transformed cells; inhibition of PARP-1 or knockdown of PARP-1 greatly decrease the levels of protein PARylation and ABHD11-AS1 and the transformed phenotype of Cr(VI)-transformed cells. Based on literature review and our novel preliminary data, our central hypothesis is: “Chronic Cr(VI) exposure-upregulated lncRNA ABHD11-AS1 sponges miR-182-5p and causes its down-regulation, which leads to increased level of the oncogenic Rac-GEF Tiam1 promoting Cr(VI) carcinogenesis”. Three aims are proposed: Aim 1 will determine the mechanism by which chronic Cr(VI) exposure up-regulates ABHD11-AS1. Aim 2 will demonstrate that ABHD11-AS1 sponges miR-182-5p causing its down- regulation and promoting Cr(VI)-exposure-induced CSC-like property, cell transformation and tumorigenesis. And Aim 3 will demonstrate that down-regulation of miR-182-5p increases levels of the oncogenic Rac-GEF Tiam1 promoting Cr(VI)-exposure-induced CSC-like property, cell transformation and tumorigenesis. Tiam1 knockout mice will be used to demonstrate that Tiam1 plays an important role in Cr(VI)-induced lung carcinogenesis.