Dysregulations of functional RNA modifications and hexavalent chromium lung carcinogenesis

功能性RNA修饰失调与六价铬肺癌发生

• 批准号: 10565860
• 负责人: Chengfeng Yang
• 金额: --
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-08 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10565860
• 关键词: Affect; Biological Assay; Biological Process; Biology; CISH gene; Cancer Etiology; Carcinogens; Cells; Chemicals; Chronic; Control Groups; Country; DNA; Data; Down-Regulation; Environmental Carcinogens; Environmental Health; Enzyme-Linked Immunosorbent Assay; Exposure to; Gene Expression; Goals; Histones; Human; Impairment; Incidence; Janus kinase; Knockout Mice; Lung; Lung Neoplasms; Lysine; Malignant neoplasm of lung; Mediating; Messenger RNA; Metal Carcinogenesis; Methyltransferase; Microarray Analysis; Modification; Monoubiquitination; Mus; Nucleotides; Occupational; Oncogenic; Pathway interactions; Persons; Play; Post-Translational Protein Processing; Process; Promoter Regions; Property; Proteins; RNA; RNA Polymerase II; RNA Stability; Reader; Resistance; Review Literature; Ribose; Role; STAT protein; Small Interfering RNA; Tumor Burden; Tumor Tissue; Tumorigenicity; United States; Untranslated RNA; Up-Regulation; Western Blotting; cancer stem cell; cancer therapy; carcinogenesis; carcinogenicity; cell transformation; chromium hexavalent ion; conditional knockout; environmental carcinogenesis; insight; knock-down; lung carcinogenesis; lung tumorigenesis; mouse model; novel; overexpression; pollutant; promoter; recruit; stem cell self renewal; stem-like cell; transcriptome sequencing; tumor progression; tumorigenesis; ubiquitin isopeptidase; ubiquitin-protein ligase

PROJECT SUMMARY/ABSTRACT Hexavalent chromium [Cr(VI)] is one of the most common environmental carcinogens causing lung cancer, however, the mechanism of Cr(VI) carcinogenesis remains elusive. The N6-methyladenosine (m6A) modification is the most prevalent internal modification in eukaryotic messenger RNAs (mRNAs), which is dynamically regulated by three groups of proteins known as writers, erasers and readers. Recent advances in demonstrating the vital roles of RNA m6A modifications in regulating gene expression and a variety of biological processes represents a breakthrough in understanding RNA biology and functions. Moreover, accumulating evidence has shown that up- regulation of m6A modifications plays critical roles in cancer progression and cancer therapy resistances. However, it remains largely un-explored whether the m6A modification dysregulation plays a role in the carcinogenic process especially in environmental carcinogenesis. The goal of this study is to investigate the mechanism of Cr(VI) carcinogenesis by studying RNA m6A modification dysregulations, focusing on the role and mechanism of chronic Cr(VI) exposure-caused m6A writer methyltransferase like 3 (METTL3) up-regulation. Our preliminary studies found: (i) Chronic Cr(VI) exposure up-regulates METTL3 expression, which contributes causally to Cr(VI)-induced cell transformation, CSC-like property and tumorigenesis. (ii) METTL3 up-regulation is also similarly detected in chronic Cr(VI) exposure-caused human and mouse lung tumor tissues. (iii) The Jak2-Stat3 pathway is activated. (iv) Jak2 and SOCS3 mRNA levels are increased and decreased in Cr(VI)-transformed cells, respectively. Stably knocking down METTL3 significantly decreases Jak2 but increases SOCS3 mRNA levels. (v) Inhibition of the Jak2-Stat3 oncogenic pathway in Cr(VI)-transformed cells significantly reduces their CSC-like property. (vi) The repressive histone 2A (H2A) lysine 119 (K119) monoubiquitination (H2AK119ub1) levels and H2AK119ub1 enrichment at METTL3 promoter region are greatly reduced in Cr(VI)-transformed cells. (vii) The expression level of a H2A deubiquitinase USP28 is up-regulated in Cr(VI)-transformed cells. Knockdown of USP28 increases H2AK119ub1 levels but reduces METTL3 protein levels. Based on literature review and our novel preliminary data, our central hypothesis is: “METTL3 up-regulation increases Jak2 and SOCS3 mRNA m6A modifications to up-regulate Jak2 but down-regulate SOCS3 expressions, which activates the oncogenic Jak2-Stat3 pathway promoting Cr(VI) carcinogenesis”. Three aims are proposed: Aim 1 will determine the mechanism of how chronic Cr(VI) exposure up- regulates METTL3 expression focusing on the role of USP28-mediated down-regulation of histone 2A monoubiquitination. Aim 2 will demonstrate that METTL3 up-regulation increases Jak2 and SCOS3 mRNA m6A modifications to activate the Jak2-Stat3 pathway promoting Cr(VI)-exposure-induced CSC-like property and tumorigenesis. Aim 3 will demonstrate that METTL3 lung-specific deletion using a METTL3 conditional knockout mouse model impairs Cr(VI) lung tumorigenesis in mice.

项目概要/摘要 六价铬[Cr(VI)]是最常见的导致肺癌的环境致癌物之一， 然而，Cr(VI) 致癌的机制仍不清楚。 N6-甲基腺苷 (m6A) 修饰是 真核生物信使 RNA (mRNA) 中最普遍的内部修饰，受到动态调节 由三组蛋白质组成，即写入器、擦除器和读取器。展示重要的最新进展 RNA m6A 修饰在调节基因表达和多种生物过程中的作用代表了 在理解 RNA 生物学和功能方面取得了突破。此外，越来越多的证据表明， m6A 修饰的调节在癌症进展和癌症治疗耐药性中发挥着关键作用。然而， m6A 修饰失调是否在致癌过程中发挥作用尚未得到充分研究 尤其是在环境致癌方面。本研究的目的是研究 Cr(VI) 的作用机制 通过研究 RNA m6A 修饰失调，重点关注慢性癌的作用和机制 Cr(VI) 暴露导致 m6A writer 甲基转移酶样 3 (METTL3) 上调。我们的初步研究发现： (i) 慢性 Cr(VI) 暴露上调 METTL3 表达，这对 Cr(VI) 诱导的细胞有因果作用 转化、CSC 样特性和肿瘤发生。 (ii) METTL3 上调也类似地在慢性病中检测到。 Cr(VI) 暴露引起的人和小鼠肺肿瘤组织。 (iii) Jak2-Stat3 途径被激活。 (四) Jak2 Cr(VI) 转化细胞中，SOCS3 和 SOCS3 mRNA 水平分别增加和减少。稳定敲击 下调 METTL3 显着降低 Jak2，但增加 SOCS3 mRNA 水平。 (v) Jak2-Stat3 的抑制 Cr(VI) 转化细胞中的致癌途径显着降低了其 CSC 样特性。 (vi) 镇压 组蛋白 2A (H2A) 赖氨酸 119 (K119) 单泛素化 (H2AK119ub1) 水平和 H2AK119ub1 富集 Cr(VI) 转化细胞中 METTL3 启动子区域大大减少。 (vii) H2A的表达水平 去泛素酶 USP28 在 Cr(VI) 转化细胞中上调。 USP28 的敲低增加了 H2AK119ub1 水平，但降低 METTL3 蛋白水平。根据文献综述和我们新颖的初步数据，我们的中央 假设是：“METTL3 上调会增加 Jak2 和 SOCS3 mRNA m6A 修饰，从而上调 Jak2 但下调 SOCS3 表达，从而激活致癌 Jak2-Stat3 通路，促进 Cr(VI) 提出了三个目标：目标 1 将确定慢性 Cr(VI) 暴露如何升高的机制。 调节 METTL3 表达，重点关注 USP28 介导的组蛋白 2A 下调的作用 单泛素化。目标 2 将证明 METTL3 上调会增加 Jak2 和 SCOS3 mRNA m6A 激活 Jak2-Stat3 通路的修饰可促进 Cr(VI) 暴露诱导的 CSC 样特性， 肿瘤发生。目标 3 将证明使用 METTL3 条件敲除 METTL3 肺部特异性缺失 小鼠模型会损害小鼠 Cr(VI) 肺肿瘤的发生。