Progressive DNA Hypomethylation as a Measure of Mitotic History and Potential Contributor to Replicative Senescence.

进行性 DNA 低甲基化作为有丝分裂历史的衡量标准和复制衰老的潜在贡献者。

• 批准号: 10672187
• 负责人: PETER W LAIRD
• 金额: $ 59.7万
• 依托单位: VAN ANDEL RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-30 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10672187
• 关键词: Acceleration; Aging; Anatomy; Attention; Attenuated; Automobile Driving; Behavior; Biological Clocks; Biology; Calibration; Cell Aging; Cell Culture Techniques; Cell Cycle; Cell Cycle Inhibition; Cell Line; Cell Lineage; Cell division; Cells; Chromatin; Chromosome Mapping; Chronology; Circadian Rhythms; Complex; Contact Inhibition; CpG Islands; CpG dinucleotide; Cues; Cytosine; DNA; DNA Maintenance; DNA Methylation; DNA Modification Methylases; DNA methyltransferase inhibition; DNMT3B gene; DNMT3a; Data; Degenerative Disorder; Dependence; Development; Disease; Elasticity; Epigenetic Process; Fibroblasts; Gene Expression; Genome; Genomics; Health; Histones; Human; Individual; Intervention; Link; Longevity; Maintenance; Malignant Neoplasms; Measures; Metabolic; Methylation; Mitosis; Mitotic; Modeling; Molecular; Outcome; Phenotype; Physiological; Process; Recording of previous events; Reporting; Research; Resistance; Role; Serum; Techniques; Telomerase; Testing; Time; Tissues; cell type; deprivation; design; genomic locus; human disease; human tissue; inhibitor; malignant phenotype; molecular clock; overexpression; premalignant; premature; prevent; senescence; tool; ubiquitin-protein ligase

PROJECT SUMMARY / ABSTRACT Cellular and molecular alterations that accumulate during cell division are considered to be contributors to aging phenotypes. Changes in epigenetic marks, including DNA methylation, have been widely documented in aging. This has spurred the development of epigenetic molecular clocks, which rely primarily on replication- independent gain of methylation at CpG islands. As molecular clocks tuned to chronological time across different tissue types with varying mitotic rates and histories, these clocks are not directly linked to mitotic cell division. In contrast, we have found that loss of DNA methylation at lamina-attached, late-replicating regions of the genome is closely tied to apparent mitotic history. We propose to use primary human cell culture to experimentally validate that mitosis is a driver of hypomethylation, and to explore the underlying mechanisms and consequences of this hypomethylation, to define genomic and chromatin features driving hypomethylation, and use this data to construct a mitotic molecular clock. In Specific Aim 1 we propose to use primary human cell culture to provide experimental evidence for the contribution of mitosis to PMD hypomethylation, and to disentangle the time-dependency and mitosis- dependency of the phenomenon using cell cycle inhibition. We will also investigate whether enhancing maintenance methylation machinery is able to counteract the progressive loss of DNA methylation. In Specific Aim 2 we propose to investigate whether DNA hypomethylation contributes to replicative senescence or associated phenotypes in primary human cell culture. We will investigate whether inhibition of maintenance methylation accelerates senescence, and whether progressive hypomethylation is extended in telomerase- immortalized primary human fibroblasts, contributing to premalignant phenotypes associated with such immortalization. We will also investigate whether accelerated senescence by progerin expression or supra- physiologic O2 leads to accelerated DNA hypomethylation. In Specific Aim 3 we will characterize the genomic and chromatin features that influence individual CpG hypomethylation rates. We will measure the rates of DNA hypomethylation of individual CpGs in six different primary human cell types and use this information to identify cell-type-specific, as well as universal genomic and chromatin drivers of hypomethylation. Finally, we will use elastic net regression on the assembled data to construct cell-type-specific and universal epigenetic mitotic clocks in Specific Aim 4. The preliminary studies strongly support the concept and feasibility of a DNA hypomethylation-based mitotic clock. The outcome of this proposed research could have important impacts on our understanding of the contribution of widespread hypomethylation to aging phenotypes, with potential implications for aging interventions. The availability of accurate molecular clocks specifically designed to measure mitotic history would provide a valuable molecular tool to characterize cells and tissues in human health and disease.

项目摘要/摘要 在细胞分裂过程中积累的细胞和分子变化被认为是导致 衰老表型。表观遗传标记的变化，包括dna甲基化，已经在 衰老。这刺激了表观遗传分子时钟的发展，这种时钟主要依赖于复制- CpG岛上甲基化的独立获得。当分子时钟调谐到时间跨度时 不同的组织类型有不同的有丝分裂速率和病史，这些时钟与有丝分裂细胞没有直接联系 组织。相反，我们发现DNA甲基化的丢失在连接到椎板的晚期复制区域 基因组与明显的有丝分裂史密切相关。我们建议使用原代人类细胞培养来 实验验证有丝分裂是低甲基化的驱动因素，并探索其潜在的机制 以及这种低甲基化的后果，以确定驱动低甲基化的基因组和染色质特征， 并使用这些数据来构建有丝分裂分子时钟。 在具体目标1中，我们建议使用原代人类细胞培养，为 有丝分裂对PMD低甲基化的贡献，并解开时间依赖性和有丝分裂- 依赖现象使用细胞周期抑制。我们还将调查是否增强了 维持甲基化的机制能够抵消DNA甲基化的逐渐丧失。具体而言 目的研究DNA低甲基化是否导致复制性衰老或 原代人类细胞培养中的相关表型。我们将调查是否阻止维持 甲基化加速衰老，而端粒酶是否延长了进行性低甲基化- 永生化的原代人类成纤维细胞，导致与此相关的癌前表型 永垂不朽。我们还将调查是否通过孕激素表达或超... 生理性氧气导致DNA低甲基化加速。在特定的目标3中，我们将描述基因组 染色质特性会影响个体的CpG低甲基化速率。我们将测量DNA的速率 在六种不同的原代人类细胞类型中单个CPGS的低甲基化并利用这一信息识别 细胞类型特异性，以及普遍的基因组和染色质低甲基化驱动因素。最后，我们将使用 弹性网络回归用于构建特定细胞类型和普遍存在的表观遗传学有丝分裂 4.初步研究有力地支持了DNA的概念和可行性 以低甲基化为基础的有丝分裂时钟。这项拟议研究的结果可能对以下方面产生重要影响 我们对广泛的低甲基化对衰老表型的贡献的理解，具有潜力 对老龄化干预的影响。专门设计的精确分子时钟的可用性 测量有丝分裂历史将为描述人类细胞和组织的特征提供有价值的分子工具 健康和疾病。