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.

项目总结/摘要 六价铬[Cr（VI）]是导致肺癌的最常见的环境致癌物之一， 然而，Cr（VI）致癌的机制仍然不清楚。N6-甲基腺苷（m6 A）修饰是 真核生物信使RNA（mRNA）中最普遍的内部修饰， 由三组蛋白质组成，分别是写入器、擦除器和读取器。最近的进展表明， RNA m6 A修饰在调节基因表达和各种生物学过程中的作用代表了一种新的生物学机制。 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， m6 A修饰的调节在癌症进展和癌症治疗抗性中起关键作用。然而，在这方面， m6 A修饰失调是否在致癌过程中起作用， 特别是在环境致癌作用中。本研究的目的是探讨Cr（VI）的作用机理 通过研究RNA m6 A修饰失调，重点研究慢性肿瘤的作用和机制， Cr（VI）离子注入引起m6 A writer甲基转移酶样3（m6 A writer methyltransferase like 3，mL 3）上调。我们的初步研究发现： (i)慢性Cr（VI）暴露上调胃L3表达，其导致Cr（VI）诱导的细胞凋亡。 转化、CSC样性质和肿瘤发生。(ii)在慢性胃炎中也同样检测到L3上调， Cr（VI）离子注入引起的人和小鼠肺肿瘤组织。(iii)Jak 2-Stat 3通路被激活。(iv)JAK2 在Cr（VI）转化的细胞中，SOCS 3和SOCS 3 mRNA水平分别升高和降低。平稳敲击 降低胃L3显著降低Jak 2但增加SOCS 3 mRNA水平。(v)抑制Jak 2-Stat 3 Cr（VI）转化细胞中的致癌途径显著降低了它们的CSC样性质。(vi)的镇压 组蛋白2A（H2 A）赖氨酸119（K119）单泛素化（H2 AK 119 ub 1）水平和H2 AK 119 ub 1富集 在Cr（VI）转化的细胞中，胃L3启动子区域大大减少。(vii)H2 A的表达水平 去泛素化酶USP 28在Cr（VI）转化的细胞中上调。USP 28的敲除增加了H2 AK 119 ub 1 水平，但降低胃L3蛋白水平。基于文献回顾和我们的新的初步数据，我们的中心 假设是：“胃L3上调增加了Jak 2和SOCS 3 mRNA m6 A修饰，从而上调了Jak 2 但下调SOCS 3的表达，从而激活致癌的Jak 2-Stat 3通路，促进Cr（VI） 致癌作用”。提出了三个目标：目标1将确定慢性Cr（VI）暴露的机制， 调节L3表达，重点关注USP 28介导的组蛋白2A下调的作用 单倍喹他嗪目的2将证明胃L3上调增加Jak 2和SCOS 3 mRNA m6 A 修饰以激活Jak 2-Stat 3途径，促进Cr（VI）-过氧化氢诱导的CSC样性质， 肿瘤发生目的3将证明使用胃L3条件性敲除的胃L3肺特异性缺失 小鼠模型损害小鼠中Cr（VI）肺肿瘤发生。