Dissecting CEBPB Function with Synthetic Biology and Imaging

用合成生物学和影像学剖析 CEBPB 功能

• 批准号: 10345006
• 负责人: Timothee Lionnet
• 金额: $ 64.68万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-01 至 2026-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10345006
• 关键词: Acetylation; Address; Adverse effects; Age; Aging; Binding; Biological Assay; Biology; Biophysical Process; Caloric Restriction; Cell Nucleus; Cells; Chromatin; Complex; Crowding; Custom; DNA Binding Domain; DNA Library; Diet; Diffusion; Dimensions; Dimerization; Dominant-Negative Mutation; Engineering; Exhibits; FRAP1 gene; Fluorescence Resonance Energy Transfer; Genes; Genetic Transcription; Goals; Health; Health Benefit; Heterodimerization; Heterogeneity; Image; Imaging Device; In Vitro; Individual; Knock-out; Libraries; Light; Longevity; Measures; Mediating; Mediator of activation protein; Messenger RNA; Metabolic; Modification; Molecular; Monitor; Nuclear; PPARG gene; Pathway interactions; Pharmacology; Phase; Phenotype; Phosphorylation; Play; Post-Translational Protein Processing; Protein Isoforms; Proteins; Recording of previous events; Reporter; Reporter Genes; Reporting; Role; Signal Transduction; System; Technology; Testing; Time; Tissues; Touch sensation; Transcription Coactivator; Transcription Repressor; Transcriptional Activation; Transcriptional Regulation; Translating; Tumorigenicity; age effect; anti aging; base; biophysical model; design; dimer; epigenetic memory; improved; interest; lipid biosynthesis; mTOR Inhibitor; mTOR inhibition; promoter; quantitative imaging; recruit; scaffold; senescence; side effect; single molecule; small molecule; synthetic biology; therapeutic target; tool; transcription factor

Project Summary Caloric restriction and reduced mTOR signaling mitigate the adverse effects of aging. They do not translate into effective anti-aging therapies though, because diets are difficult to maintain and mTOR inhibitors exhibit undesirable side effects. The transcription factor CEBPB drives many of the undesirable effects of aging downstream of the TOR pathway. The levels of its short isoform called LIP increase with age; loss of LIP increases lifespan and provides health benefits, while expression of LIP alone increase tumorigenicity. Reprogramming CEBPB and LIP therefore holds great promise as a tool to control aging. Pioneering studies indicated that LIP activates transcription of its target genes, while the canonical long isoform of CEBPB, LAP, is a transcription repressor. Recent evidence draws a more complex picture: - both isoforms are extensively modified by phosphorylation, acetylation (and many others), and those modifications impact their activity; - dimerization of CEBPB with other factors impacts its function; - finally, LIP rescues CEBPB knock-out, suggesting that LIP is not simply a dominant negative inert isoform. Understanding how these regulatory dimensions are integrated by CEBPB isoforms is a pre-requisite to develop technologies able to reprogram aging. Here, we deploy synthetic biology approaches in order to dissect how the CEBPB isoforms, their dimerization partners and post-translational modifications impact the nuclear dynamics of CEBPB and its transcription regulation activity. We also develop synthetic tools to visualize and perturb the LAP:LIP ratio in individual cells. Being able to perturb aging regulators in single cells will enable measuring the contributions of cell-autonomous aging phenotypes and those driven by signals between cells. It will also provide precise tools to dissect one of the common hallmarks of aging: increased heterogeneity of expression.

项目摘要 热量限制和MTOR信号降低减轻衰老的不良影响。他们不翻译 不过 不良副作用。转录因子CEBPB驱动衰老的许多不良影响 TOR路径的下游。其短同工型的水平称为唇部随着年龄的增长；嘴唇的损失 增加寿命并提供健康益处，而单独唇部表达会增加肿瘤性。 因此，重编程CEBPB和LIP具有控制衰老的工具的巨大希望。 开创性研究表明，唇部激活其靶基因的转录，而规范长 CEBPB的同工型是转录阻遏物。最近的证据提出了一个更复杂的图片： - 两种同工型都通过磷酸化，乙酰化（以及许多其他）进行了广泛的修饰，而这些同工型 修改会影响其活动； - CEBPB与其他因素的二聚化会影响其功能； - 最后，嘴唇营救了CEBPB的淘汰赛，这表明唇部不仅是主导的负惰性 同工型。 了解这些法规维度是如何通过CEBPB同工型整合的，这是前提的先决条件 开发能够重新编程的技术。 在这里，我们采用合成生物学方法，以剖析CEBPB同工型如何 二聚合作伙伴和翻译后修改会影响CEBPB及其ITS的核动力学 转录调控活性。 我们还开发了合成工具来可视化和扰动单个细胞中的膝盖：唇部比。能够 单个细胞中的敏捷衰老调节剂将能够测量细胞自主衰老的贡献 表型和由细胞之间信号驱动的表型。它还将提供精确的工具来剖析其中一种 衰老的常见标志：表达的异质性增加。