Building a unified framework for understanding bacterial gene regulation and chromosomal architecture

建立理解细菌基因调控和染色体结构的统一框架

• 批准号: 9892610
• 负责人: Lydia Freddolino
• 金额: $ 6.78万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9892610
• 关键词: ATAC-seq; Architecture; Area; Bacteria; Bacterial Antibiotic Resistance; Bacterial Chromosomes; Bacterial Genes; Bacterial Genome; Behavior; Binding; Binding Sites; Bioinformatics; Biotechnology; DNA-Binding Proteins; DNA-Protein Interaction; Data Set; Escherichia coli; Eukaryota; Food; Gene Expression Regulation; Genetic Transcription; Growth; Heterochromatin; Impairment; Infection; Investigation; Logic; Maps; Molecular; Molecular Biology; Organism; Orphan; Physiological; Play; Process; Proteins; Role; Signal Transduction; Site; Source; Stress; Study models; Technology; Therapeutic; Time; Transcriptional Regulation; Virulence; combat; experimental study; follow-up; global health; improved; innovation; interest; prevent; protein profiling; tool; transcription factor

Transcriptional regulation via protein-DNA interactions plays an important role in the regulatory networks of all known organisms. Bacterial regulatory networks are now an especially fruitful target for detailed investigation: as antibiotic-resistant bacteria continue to emerge as a global health threat, new and innovative approaches to either preventing virulence or impairing bacterial growth are required. As our ability to predict and exploit bacterial behavior for therapeutic purposes hinges on our understanding of the logic behind their regulatory networks, it is of great utility to fully map those networks and the molecular mechanisms underlying them. Several challenges, both old and newly recognized, stand in the way of a comprehensive understanding of regulatory logic, even in well-studied models such as Escherichia coli. Progress in mapping bacterial regulatory networks has in general been slow, requiring a steady march of mapping binding sites of one transcription factor (TF) at a time. Even when such experiments are done, they can typically be performed only under a handful of physiological conditions, and thus may miss key contributions of a transcription factor in responding to specific environmental triggers. In addition, contrary to prevailing dogma over the last several decades, we and others have recently gathered substantial evidence that bacterial chromosomes are in fact not universally accessible to transcription, but rather, that they are packaged by densely protein occupied heterochromatin-like regions that we refer to as EPODs, which influence both overall chromosomal architecture and transcriptional regulation in particular. Progress in the area of fully charting bacterial regulation of transcription via DNA binding proteins thus simultaneously requires more efficient coverage of transcription factor space and an improved understanding of the role of larger-scale protein occupancy in gene regulation. We have optimized a technology referred to as IPODHR for overall profiling of protein occupancy on bacterial genomes, similar to the signal provided by ATAC-seq in eukaryotes. Building on IPODHR data sets as a cornerstone, we are pursuing several highly innovative and efficient approaches to expand our understanding of bacterial regulatory networks: Massively parallel profiling of TF occupancy. Tracking IPODHR signal across known TF binding sites, in tandem with appropriate bioinformatic analysis, provides occupancy information on dozens of known TFs in a single experiment. We will utilize this technology to profile TF binding under a broad range of conditions. Identification of orphan TFs. IPODHR profiles enable us to identify active regulatory sites under conditions of interest, and identify the responsible TFs through follow-up experiments and bioinformatics. Regulatory roles and molecular biology of EPODs. IPODHR has revealed the presence of EPODs across a wide range of bacterial taxa, and we will determine the full impact of EPODs on condition-dependent gene regulation, and the molecular mechanisms through which these regions are established.

通过蛋白质-DNA相互作用进行的转录调控在ALL的调控网络中起着重要作用 已知的生物。细菌调控网络现在是一个特别有成效的详细研究目标： 随着抗药性细菌继续成为全球健康威胁，新的创新方法 要么要防止毒力，要么要阻止细菌生长。因为我们预测和开发的能力 细菌用于治疗目的的行为取决于我们对其调控背后的逻辑的理解 对于网络来说，充分绘制这些网络及其背后的分子机制是非常有用的。 一些挑战，既有旧的，也有新认识的，阻碍了全面的 对调控逻辑的理解，即使是在研究得很好的模型中，如大肠杆菌。测绘进展 细菌调控网络总体上是缓慢的，需要稳步绘制结合位点的图谱 一次一个转录因子(Tf)。即使当这样的实验完成后，它们通常也可以进行 仅在极少数生理条件下，因此可能会错过转录因子的关键作用 在对特定的环境触发因素做出反应时。此外，与过去几年流行的教条相反 几十年来，我们和其他人最近收集了大量证据，表明细菌染色体实际上是 并不是普遍可以转录，而是由密集占据的蛋白质包装而成 异染色质样区，我们称之为EPOD，它影响整个染色体 尤其是建筑和转录调控。全面绘制细菌调控图领域的进展 因此，通过DNA结合蛋白进行转录的同时需要更有效地覆盖转录 因子空间和对更大范围的蛋白质占据在基因调控中的作用的更好的理解。 我们优化了一种名为IPODHR的技术，用于全面分析蛋白质占有率 细菌基因组，类似于真核生物中atac-seq提供的信号。在IPODHR数据集基础上构建为 作为基石，我们正在寻求几种高度创新和高效的方法来扩大我们的 了解细菌调控网络： 大规模并行分析特遣部队占有率。跨已知转铁蛋白结合位点跟踪IPODHR信号，在 与适当的生物信息学分析相结合，提供有关数十个已知TF的占用信息 单次实验。我们将利用这项技术在广泛的条件下分析TF结合。 孤儿TF的身份识别。IPODHR配置文件使我们能够在以下条件下识别活跃的监管站点 兴趣，并通过后续实验和生物信息学确定负责任的信托基金。 EPODS的调节作用和分子生物学。IPODHR揭示了EPOD在整个 广泛的细菌分类群，我们将确定EPODS对条件依赖基因的全面影响 调控，以及建立这些区域的分子机制。