Transcriptional Control During Erythropoiesis

红细胞生成过程中的转录控制

• 批准号: 10053139
• 负责人: Marjorie Carole Brand
• 金额: $ 72.66万
• 依托单位: INSTITUTE FOR SYSTEMS BIOLOGY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-16 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10053139
• 关键词: Adult; Automobile Driving; Benchmarking; Binding; Biological Assay; Blood; Cell Fate Control; Cell model; Cell physiology; Cells; ChIP-seq; Chromatin; Complex; Computer Analysis; Dancing; Data; Disease; Erythrocytes; Erythroid; Erythroid Cells; Erythropoiesis; FLI1 gene; Gene Expression; Genes; Genetic Transcription; Genomics; Goals; Health; Hematopoietic stem cells; Hemoglobin; Human; Joints; Knowledge; Mass Spectrum Analysis; Measurement; Measures; Messenger RNA; Methodology; Methods; Mission; Modeling; Monitor; Nature; Outcome; Patients; Pharmacology; Population; Process; Production; Proteins; Proteome; Proteomics; Proxy; Public Health; Reaction; Regulation; Research; Role; Specific qualifier value; System; Technology; Testing; Time; Transcript; Transcriptional Regulation; Umbilical Cord Blood; United States National Institutes of Health; Validation; Vision; Work; base; beta Globin; beta Thalassemia; comparative; design; dosage; experimental study; fetal; genome-wide; human model; improved; in vivo; innovation; network models; novel; novel therapeutic intervention; novel therapeutics; progenitor; programs; temporal measurement; transcription factor; transcriptome; transcriptomics

Erythropoiesis is a dynamic process governed by quantitative changes in the relative levels of transcription fac- tors (TFs). Due to the current paucity of quantitative data on the proteins that constitute the transcriptional regu- latory network (TRN), most models of erythropoiesis are based primarily on mRNA measurements and do not typically consider changes in the protein levels of specific TFs. This significantly limits the understanding of erythropoiesis and other transcriptionally regulated processes such as ß-globin expression, ultimately impinging on the capacity to correct hemoglobin disorders. The long-term goal is to decipher the TRN that controls eryth- ropoiesis in health and disease. The objective of this proposal is to significantly expand our TRN model for cell fate decision during erythropoiesis by integrating dynamic bulk and single cell TF protein abundance measure- ments with other transcription-relevant -omics data. The central hypothesis is that the relative protein levels of TFs are critical parameters in the establishment of proper gene expression programs during the continuum of differentiation, and that erythropoiesis is driven by graded changes in the relative amounts of specific combina- tions of TFs. The rationale is that integration of the dynamic and quantitative nature of the TF proteome into an expanded TRN of erythropoiesis will result in a model with improved predictive power which will serve as a benchmark for healthy erythropoiesis against which to compare erythroid-related disease states, and will facili- tate the identification of pharmacological agents to restore normal erythropoiesis. Three specific aims have been designed: 1) Absolute quantification of the TF proteome during erythropoiesis; 2) Determine how gradual changes in the abundance of multiple TFs in single cells initiate and progressively reinforce cell fate decisions along the erythroid trajectory; and 3) Computational analysis, modeling and validation of the erythropoiesis TRN. For the first aim, a high throughput quantitative mass spectrometry (MS) approach will be used to measure absolute levels of the TF proteome during ex vivo erythropoiesis of HSPCs derived from healthy donors. For the second aim, complementary CyTOF and targeted-MS proteomic approaches will be combined to estimate TF protein abundances in single cells, with other single cell –omics technologies to measure changes in gene ex- pression and TF genomic binding during ex vivo erythropoiesis. Under the third aim, TRN models of erythropoi- esis will be built utilizing measurements of TF protein abundances, and other transcription-relevant –omics data. Functional validation will be performed for TFs that have been implicated in transcriptional control during eryth- ropoiesis based on our recent results. The approach is innovative because it uses a novel combination of single cell and bulk proteomics methodologies to quantify large numbers of TFs during erythropoiesis in primary human cells and uses the data for integrative TRN modeling. The proposed research is significant because it will illumi- nate complex regulatory processes that control erythropoiesis. Ultimately, such knowledge has the potential to guide the design of new therapeutics to re-establish proper ß-globin expression in ß-thalassemic patients.

红细胞生成是一个动态过程，受转录因子相对水平的定量变化控制。 托尔（TF）。由于目前缺乏构成转录调控蛋白的定量数据 实验室网络 (TRN)，大多数红细胞生成模型主要基于 mRNA 测量，而不是 通常考虑特定转录因子的蛋白质水平的变化。这极大地限制了人们的理解 红细胞生成和其他转录调节过程，例如 β-珠蛋白表达，最终影响 纠正血红蛋白紊乱的能力。长期目标是破译控制红细胞的TRN 健康和疾病中的细胞生成。该提案的目标是显着扩展我们的细胞 TRN 模型 通过整合动态体积和单细胞 TF 蛋白丰度测量来决定红细胞生成过程中的命运 与其他转录相关的组学数据进行比较。中心假设是相对蛋白质水平 转录因子是在连续过程中建立适当的基因表达程序的关键参数。 分化，并且红细胞生成是由特定组合的相对量的分级变化驱动的 TF 的蒸发散。基本原理是将 TF 蛋白质组的动态和定量性质整合到一个 红细胞生成的扩展 TRN 将产生一个具有改进预测能力的模型，该模型将作为 健康红细胞生成的基准，用于比较红细胞相关疾病状态，并将促进 tate 鉴定恢复正常红细胞生成的药物。制定了三个具体目标 设计：1）红细胞生成过程中TF蛋白质组的绝对定量； 2）确定渐进的程度 单细胞中多个转录因子丰度的变化启动并逐渐强化细胞命运决定 沿着红系轨迹； 3) 红细胞生成 TRN 的计算分析、建模和验证。 对于第一个目标，将使用高通量定量质谱 (MS) 方法来测量 来自健康供体的 HSPC 离体红细胞生成过程中 TF 蛋白质组的绝对水平。对于 第二个目标，将结合互补的 CyTOF 和靶向 MS 蛋白质组学方法来估计 TF 单细胞中的蛋白质丰度，以及其他单细胞组学技术来测量基因表达的变化 离体红细胞生成过程中的抑制和 TF 基因组结合。在第三个目标下，红细胞生成的 TRN 模型 esis 将利用 TF 蛋白丰度的测量和其他转录相关的组学数据来构建。 将对红细胞期间参与转录控制的 TF 进行功能验证。 ropiesis 基于我们最近的结果。该方法是创新的，因为它使用了单一的新颖组合 用于量化原代人类红细胞生成过程中大量 TF 的细胞和本体蛋白质组学方法 细胞并使用数据进行综合 TRN 建模。拟议的研究意义重大，因为它将阐明 控制红细胞生成的复杂调节过程。最终，这些知识有可能 指导新疗法的设计，以在 β 地中海贫血患者中重新建立适当的 β-珠蛋白表达。