Bacterial Rogue Methyltransferases Inducing Human Epimutations

细菌流氓甲基转移酶诱导人类表观突变

基本信息

批准号：
10439471
负责人：
Emily E Fink
金额：
$ 2.14万
依托单位：
CLEVELAND CLINIC LERNER COM-CWRU
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-06-01 至 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10439471
关键词：
ATAC-seq Adenine Aerobic Affect Anaerobic Bacteria Anatomy BRAF gene Bacillus Bacteria Bacterial DNA Bacterial Genome Bioinformatics Biological Assay Cancer Etiology Candidate Disease Gene Cell Nucleus Cells ChIP-seq Chromatin Clinical Coculture Techniques Colon Colon Carcinoma Colorectal Cancer Core Facility CpG Island Methylator Phenotype Cytosine DNA DNA Binding DNA Methylation DNA Modification Methylases DNA Repair DNA Restriction Enzymes DNA Restriction-Modification Enzymes DNA biosynthesis DNA methylation profiling Data Defense Mechanisms Detection Development Enzymes Epigenetic Process Epithelial Cells Experimental Models Fusobacterium nucleatum Genes Genetic Genetic Transcription Genome Genomics HCT116 Cells Human Human Genome Hypermethylation In Vitro Infection Invaded Kilogram Kinetics Knowledge Link Location Malignant Neoplasms Methylation Methyltransferase Microsatellite Instability Modification Molecular Mutation Pathway interactions Patients Play Proteins Publishing Quantitative Reverse Transcriptase PCR RNA Splicing Research Institute Risk Management Role Side Signal Pathway Signal Transduction Source Specificity Specimen T cell response Testing The Cancer Genome Atlas Tumor Suppressor Genes Virulence Factors Virus Western Blotting XCL1 gene advanced disease anticancer research bisulfite sequencing cancer cell cancer type chromatin immunoprecipitation chromatin remodeling cohort colon cancer cell line demethylation dysbiosis epigenome experimental study fascinate gut dysbiosis gut microbiome gut microbiota histone methyltransferase host microbiome human DNA in vitro activity in vivo methyl group microbial microbial genome microbial host microbiome microorganism next generation sequencing novel novel therapeutics opportunistic pathogen promoter transcriptome sequencing treatment risk tumor viral DNA

项目摘要

PROJECT SUMMARY: DNA methylation is dysregulated in every form of cancer. While there is substantial knowledge regarding the location and function of abnormal DNA methylation across tumor types, we are severely lacking in understanding how such abnormal DNA methylation is established during cancer formation. Colorectal cancer (CRC), the third most common cancer type worldwide, has been heavily associated with alterations in DNA methylation, however, mutations in DNA methylation/demethylation pathway genes are rare and cannot account for the exaggerated DNA hypermethylation. Roughly 1.5 kg of bacteria reside in our gut and have been shown to play a substantial role in the development of CRC. Recent evidence suggests that these bacteria can alter the colon cell epigenome by inducing aberrant DNA hypermethylation, although the mechanism promoting this is still not completely understood. One such microorganism, Fusobacterium nucleatum, is able to invade into colon epithelial cells and has been heavily associated with CRC development, as well as hypermethylation of tumor suppressor genes. Roughly 90% of bacteria, including Fusobacterium nucleatum, contain DNA methyltransferase enzymes as a component of their restriction-modification system; a defense mechanism aimed at protecting their genome against invading viruses by methylating specific motifs within their DNA as to distinguish between their own and invading viral DNA. To date, >3500 different R-M motifs have been identified, thus our gut microbiome is filled with microbial genome-modifying enzymes, however their potential to modify human DNA has not been evaluated. The long-term objective of this proposal is to evaluate whether F. nucleatum DNA methyltransferase enzymes are able to access and aberrantly methylate colon cell DNA, potentially giving rise to the epimutations observed in CRC development. This will be tested with three specific aims. The first aim will experimentally evaluate F. nucleatum methyltransferase activity in vitro, by ectopically expressing F. nucleatum DNA methyltransferase enzymes in CRC cell lines and evaluating their ability to enter the nucleus, methylate DNA, as well as determine the locations of DNA methylation. The second aim will take advantage of the microbiome core facilities at Lerner Research Institute to preform bacterial co-culture experiments using F. nucleatum that have been genetically modified to contain tagged-DNA methyltransferase enzyme, to test whether F. nucleatum DNA methyltransferase can access and methylate colon cell DNA. The third aim is to detect F. nucleatum activity in vivo, by searching for F. nucleatum R-M motif enrichment at CRC hypermethylated loci, using our labs published MBD-seq data and the TCGA CRC cohort. Results could reveal a novel paradigm in host-microbiome interactions, as direct epigenetic crosstalk between microbial and host cells has not been fully explored. However, if true, this new paradigm will transform studies of bacterial-associated cancers and open new therapeutic avenues for treatment and risk-management.

项目摘要：DNA 甲基化在各种癌症中均失调。虽然有大量关于跨肿瘤类型异常 DNA 甲基化的位置和功能的知识，我们非常重视缺乏了解这种异常的 DNA 甲基化在癌症形成过程中是如何形成的。结直肠癌症（CRC）是世界上第三大常见的癌症类型，与身体结构的改变密切相关。 DNA 甲基化，然而，DNA 甲基化/去甲基化途径基因的突变很少见，不能解释了过度的 DNA 高甲基化。大约 1.5 公斤的细菌驻留在我们的肠道内，并已被表明其在 CRC 的发展中发挥着重要作用。最近的证据表明这些细菌可以通过诱导异常 DNA 高甲基化来改变结肠细胞表观基因组，尽管该机制促进这一点还没有被完全理解。其中一种微生物，具核梭杆菌，能够侵入结肠上皮细胞，与结直肠癌的发展以及结肠上皮细胞的高甲基化密切相关肿瘤抑制基因。大约 90% 的细菌，包括具核梭杆菌，都含有 DNA 甲基转移酶作为其限制性修饰系统的组成部分；防御机制旨在通过甲基化 DNA 中的特定基序来保护其基因组免受病毒入侵区分自己的DNA和入侵的病毒DNA。迄今为止，已鉴定出超过 3500 个不同的 R-M 基序，因此，我们的肠道微生物组充满了微生物基因组修饰酶，但是它们的修饰潜力人类 DNA 尚未得到评估。该提案的长期目标是评估 F. nucleatum 是否 DNA 甲基转移酶能够接触结肠细胞 DNA 并异常甲基化，从而可能产生 CRC 发育中观察到的表突变的增加。这将通过三个具体目标进行测试。第一个目标将通过异位表达具核梭杆菌甲基转移酶活性来进行体外实验评估。 CRC细胞系中的有核DNA甲基转移酶并评估其进入细胞核的能力，甲基化 DNA，并确定 DNA 甲基化的位置。第二个目标将利用勒纳研究所的微生物组核心设施使用 F. 经基因改造含有标记 DNA 甲基转移酶的有核杆菌，以测试 F. nucleatum DNA甲基转移酶是否可以接触结肠细胞DNA并将其甲基化。第三个目标是通过搜索 CRC 超甲基化处的 F. nucleatum R-M 基序富集来检测 F. nucleatum 体内活性位点，使用我们实验室发布的 MBD-seq 数据和 TCGA CRC 队列。结果可能揭示一种新的范式在宿主-微生物组相互作用中，微生物和宿主细胞之间的直接表观遗传串扰尚未被研究充分探索。然而，如果这是真的，这种新范式将改变细菌相关癌症和为治疗和风险管理开辟新的治疗途径。

项目成果

期刊论文数量（2）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Processing and cryopreservation of human ureter tissues for single-cell and spatial transcriptomics assays.

用于单细胞和空间转录组学分析的人类输尿管组织的处理和冷冻保存。

DOI：
发表时间：
2022-12-16
期刊：
影响因子：
0
作者：
Fink, Emily E;Sona, Surbhi;Lee, Byron H;Ting, Angela H
通讯作者：
Ting, Angela H

Single-cell and spatial mapping Identify cell types and signaling Networks in the human ureter.

DOI：
10.1016/j.devcel.2022.07.004
发表时间：
2022-08-08
期刊：
Developmental cell
影响因子：
11.8
作者：
Fink, Emily E.;Sona, Surbhi;Tran, Uyen;Desprez, Pierre-Emmanuel;Bradley, Matthew;Qiu, Hong;Eltemamy, Mohamed;Wee, Alvin;Wolkov, Madison;Nicolas, Marlo;Min, Booki;Haber, Georges-Pascal;Wessely, Oliver;Lee, Byron H.;Ting, Angela H.
通讯作者：
Ting, Angela H.