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: 10381280
• 负责人: Chengfeng Yang
• 金额: $ 41.58万
• 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 2026-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10381280
• 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; 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; base; 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-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.

项目摘要/摘要 六价铬[CR（VI）]是最常见的环境致癌物之一，导致肺癌， 但是，CR（VI）癌变的机制仍然难以捉摸。 N6-甲基丹代氨酸（M6A）的修饰为 真核信使RNA（mRNA）中最普遍的内部修饰，该修饰是动态调节的 由三组被称为作家，橡皮擦和读者的蛋白质。展示至关重要的最新进展 RNA M6A修饰在调节基因表达和多种生物过程中的作用代表 理解RNA生物学和功能的突破。此外，积累的证据表明， M6A修饰的调节在癌症进展和癌症耐药性中起关键作用。然而， M6A修饰失调是否在致癌过程中起作用，在很大程度上没有探索 特别是在环境癌变中。这项研究的目的是研究CR（VI）的机制 通过研究RNA M6A修饰失调，重点关注慢性的作用和机制，致癌 CR（VI）暴露引起的M6A作者甲基转移酶，例如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，有时会导致CR（VI）诱导的细胞 转化，CSC样性质和肿瘤发生。 （ii）在慢性中也类似地检测到Mettl3上调 CR（VI）暴露引起的人和小鼠肺肿瘤组织。 （iii）激活JAK2-STAT3途径。 （iv）JAK2 SOCS3 mRNA水平分别增加并降低CR（VI）转化的细胞。稳定敲门 下降METTL3显着降低了JAK2，但SOCS3 mRNA水平增加。 （v）抑制JAK2-Stat3 CR（VI）转化细胞中的致癌途径显着降低了其CSC样性质。 （vi）反射性 Hisstone 2A（H2A）赖氨酸119（K119）单次素激（H2AK119UB1）水平和H2AK119UB1富集 在CR（VI）转化的细胞中，METTL3启动子区域大大降低。 （vii）H2a的表达水平 去泛素酶USP28在CR（VI）转化的细胞中被上调。 USP28的敲低增加了H2AK119UB1 水平但降低了Mettl3蛋白水平。根据文献综述和我们的新型初步数据，我们的中央 假设是：“ METTL3上调增加JAK2和SOCS3 mRNA M6A修饰以上调节JAK2 但是下调SOCS3表达式，它激活了促进CR（VI）的致癌JAK2-STAT3途径 提出了三个目标：AIM 1将确定慢性CR（VI）暴露的机制 调节METTL3表达，重点是USP28介导的Hisstone 2a下调的作用 单泛素化。 AIM 2将证明METTL3上调增加JAK2和SCOS3 mRNA M6A 修改以激活JAK2-STAT3途径促进CR（VI）曝光引起的CSC样属性和 肿瘤发生。 AIM 3将证明使用Mettl3有条件敲除的Mettl3肺特异性缺失 小鼠模型会损害小鼠的CR（VI）肺肿瘤发生。