Understanding the full spectrum of epigenetic vulnerability in cancer through the delineation of DNA methylation function in gene 3' end

通过描绘基因 3 端 DNA 甲基化功能，全面了解癌症的表观遗传脆弱性

• 批准号: 10765365
• 负责人: Angela H Ting
• 金额: $ 39.24万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-19 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10765365
• 关键词: 3' Untranslated Regions; Aberrant DNA Methylation; Acceleration; Affect; Binding; Binding Sites; Biochemical; Bioinformatics; Biological; Biological Assay; CCCTC-binding factor; Cancer Biology; Cancer Patient; Cancer cell line; Candidate Disease Gene; Cell Line; Cells; Chemicals; Code; Collection; Colon; Coupled; DNA; DNA Methylation; DNA Modification Process; DNA Sequence; DNA mapping; DNA-Binding Proteins; DU145; Data; Defect; Disease; Distal; Dynein ATPase; Epigenetic Process; Event; Exhibits; Gene Expression; Gene Silencing; Genes; Genetic Transcription; Genomic approach; Genomics; Goals; HCT116 Cells; Human; Hypermethylation; In Vitro; International; Knowledge; LNCaP; Light; Malignant Neoplasms; Maps; Mediating; Messenger RNA; Methylation; MicroRNAs; Modeling; Modification; Molecular; Molecular Biology; Mutate; Nuclear; Oncogenes; Pathologic; Pattern; Physiological; Poly A; Polyadenylation; Polyadenylation Pathway; Prevalence; Prevention strategy; Process; Prostate; Protein Isoforms; Proteins; Proteome; RNA Polymerase II; RNA Processing; Regulation; Reporter; Reporting; SW480; Sampling; Seminal; Shapes; Site; Testing; The Cancer Genome Atlas; Therapeutic; Tissues; Transcript; Tumor Suppressor Genes; cancer cell; cancer genome; cancer initiation; cancer therapy; cancer type; chromatin immunoprecipitation; epigenetic regulation; genome-wide; genomic locus; improved; in vivo; mRNA Precursor; methylation pattern; novel; predictive modeling; prevent; promoter; side effect; stable cell line; therapy design; transcription factor; transcription termination; transcriptome; transcriptome sequencing; treatment strategy; tumor progression; tumorigenesis

Abstract DNA methylation abnormalities occur in all genomic contexts throughout the cancer genome, and studies of such aberrant epigenetic patterns have led to seminal discoveries regarding tumor suppressor gene silencing by promoter hypermethylation. While promoter hypermethylation causes transcriptional silencing, the functions of non-promoter DNA methylation are poorly defined. Such lack of knowledge severely limits our ability to contextualize the effects of abnormal DNA methylation on cancer biology and to realize the full potential of epigenetic-based cancer therapy. In this proposal, we propose to investigate the impact of non-promoter DNA methylation on the transcriptome and will focus on studying the functions of DNA methylation near gene 3' ends. Using a pair of isogenic cancer cells (HCT116 and DKO cells) that differ specifically in their ability to maintain DNA methylation, we discovered a robust association between gene 3' end differential DNA methylation and alternative cleavage and polyadenylation (APA) events. Briefly, pre-mRNAs undergo cleavage and polyadenylation as part of normal mRNA 3' end formation, and alternative sites of cleavage and polyadenylation can be utilized to produce transcripts with varying regulatory sequences in the 3' untranslated regions (3' UTRs) or protein isoforms via APA within coding sequences. Previous studies have demonstrated that cancer cells can hijack the APA pathway to skew expression of short 3' UTRs in oncogenes to evade negative regulation, highlighting APA as an important process involved in cancer initiation and progression. By leveraging the Cancer Genome Atlas (TCGA) data, we could also observe the correlation between gene 3' DNA methylation and APA at select loci in cancer patient samples. We hypothesize that differential DNA methylation in gene 3' ends can result in cancer-promoting expression patterns through regulation of APA. In Aim 1, we will define the mechanism of DNA methylation-regulated APA using a combination of computational, biochemical, molecular biology, and genomic approaches. In Aim 2, we will validate our model and in vitro data by mapping polyadenylation site usage in additional cancer cell lines and testing for disease-relevant APA events across different cancer types using publically available RNA-seq and DNA methylation data from the International Cancer Genome Consortium (ICGC). The results of our study will improve overall functional understanding of non-promoter DNA methylation, provide a novel mechanism for APA regulation, and ultimately accelerate discovery of novel targets for cancer management. We also envision that our findings here can have broad impact on our knowledge of how epigenetic regulation shapes the transcriptome in cancer as well as in other human healthy and pathological conditions.

抽象的 DNA甲基化异常在整个癌症基因组的所有基因组中都发生，并研究 这种异常表观遗传模式导致有关肿瘤抑制基因沉默的开创性发现 通过启动子高甲基化。而启动子高甲基化会引起转录沉默，而功能 非促启发子DNA甲基化的定义很差。这种缺乏知识严重限制了我们的能力 情境化异常DNA甲基化对癌症生物学的影响，并实现全部潜力 基于表观遗传的癌症治疗。在此提案中，我们建议研究非启动者DNA的影响 转录组上的甲基化，并将重点研究基因3'基因附近的DNA甲基化的功能 结束。使用一对同源性癌细胞（HCT116和DKO细胞），这些细胞在其能力方面有着不同的差异 维持DNA甲基化，我们发现基因3'末端差分DNA之间存在稳健的关联 甲基化和替代裂解和聚腺苷酸化（APA）事件。简而言之，前MRNA进行乳沟 作为正常mRNA 3'末端形成的一部分，裂解和裂解和 聚腺苷酸化可用于在3'未翻译中产生具有不同调节序列的转录本 在编码序列中通过APA区域（3'UTRS）或蛋白质同工型。先前的研究表明 癌细胞可以劫持APA途径，以偏向于肿瘤基因中短3'UTR的表达 负调节，强调APA是癌症开始和进展涉及的重要过程。经过 利用癌症基因组图集（TCGA）数据，我们还可以观察到基因3'之间的相关性 在癌症患者样品中，DNA甲基化和APA在精选基因座处。我们假设差异DNA 基因3'末端中的甲基化可以通过调节APA导致促进癌症的表达模式。在 AIM 1，我们将使用计算的组合来定义DNA甲基化调节APA的机制 生化，分子生物学和基因组方法。在AIM 2中，我们将验证我们的模型和体外数据 通过在其他癌细胞系中绘制聚腺苷酸化位点的使用并测试与疾病相关的APA 使用公开可用的RNA-seq和DNA甲基化数据跨不同癌症类型的事件 国际癌症基因组联盟（ICGC）。我们研究的结果将改善整体功能 了解非启动者DNA甲基化，为APA调节提供了新的机制，并 最终加速了发现新颖的癌症管理目标。我们还设想我们的发现 这里可能对我们对表观遗传调节如何塑造转录组的了解有广泛的影响 癌症以及其他人类健康和病理状况。