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甲基化以各种形式的癌症失调。虽然有实质性关于跨肿瘤类型异常DNA甲基化的位置和功能的知识，我们严重缺乏了解如何在癌症形成过程中建立这种异常的DNA甲基化。结直肠癌症（CRC）是全球第三大癌症类型，与改变有关然而，DNA甲基化，DNA甲基化/脱甲基化途径的突变基因很少，不能解释了夸张的DNA高甲基化。大约1.5公斤的细菌居住在我们的肠道中，已经证明在CRC的发展中起着重要作用。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，尽管促进的机制这仍然没有完全理解。一种这样的微生物，fusobacterium nucleatum，能够入侵结肠上皮细胞，与CRC的发展密切相关，以及肿瘤抑制基因。大约90％的细菌，包括核细菌，含有DNA 甲基转移酶是其限制性修饰系统的组成部分；防御机制旨在通过在其DNA中甲基甲基特定基序来保护其基因组免受入侵病毒的影响区分自己和入侵的病毒DNA。迄今为止，已经确定了> 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DNA尚未评估。该提议的长期目标是评估F. nucleatum是否 DNA甲基转移酶能够访问并异常甲基甲酸盐结肠DNA，有可能给出在CRC发育中观察到的表述。这将通过三个特定目标进行测试。第一个目标将通过异位表达F实验评估体外甲基转移酶活性的F. CRC细胞系中的核DNA甲基转移酶酶，并评估其进入核的能力，甲基化DNA，并确定DNA甲基化的位置。第二个目标将利用 Lerner Research Institute的微生物组核心设施使用F进行了细菌共培养实验。经过遗传修饰以包含标记的DNA甲基转移酶的核，以测试 F. nucleatum DNA甲基转移酶是否可以访问和甲基化结肠细胞DNA。第三个目标是通过在CRC高甲基化的F. nubleatum R-M基序富集来检测体内核活性。 Loci，使用我们的实验室发布的MBD-Seq数据和TCGA CRC队列。结果可能揭示了一种新颖的范式在宿主 - 微生物组相互作用中，由于微生物和宿主细胞之间的直接表观遗传串扰尚未完全探索。但是，如果是真的，这种新范式将改变对细菌相关癌症和为治疗和风险管理开放新的治疗途径。

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）

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.

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