Establishing the role and mechanisms of LSD1 during megakaryocytic and erythroid fate commitment

建立LSD1在巨核细胞和红细胞命运决定过程中的作用和机制

• 批准号: 10605993
• 负责人: Evrett Thompson
• 金额: $ 5.02万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10605993
• 关键词: ATAC-seq; Adult; Anemia; Benign; Biological Models; Blood; Blood Cells; Blood Platelets; Bone Marrow; CD34 gene; Candidate Disease Gene; Cell Culture Techniques; Cell Differentiation process; Cell Lineage; Cell Therapy; Cells; Chimeric Proteins; Chromatin; Computer Analysis; Data; Data Set; Disease; Enhancers; Epigenetic Process; Erythrocytes; Erythroid; Erythroid Progenitor Cells; Erythropoiesis; Gene Expression; Gene Targeting; Genes; Genetic; Genetic Transcription; Genome; Genomics; Goals; Guide RNA; Hematopoiesis; Hematopoietic; Hematopoietic stem cells; Human; Individual; Investigation; KDM1A gene; Lead; Lysine; Maintenance; Malignant - descriptor; Mediating; Megakaryocytes; Megakaryocytopoieses; Messenger RNA; Modification; Mus; Myelogenous; Pathology; Pathway interactions; Play; Population; Process; Production; Publishing; Research; Research Personnel; Role; Series; Site; Specific qualifier value; Testing; Training; candidate identification; career; cell fate specification; cell type; demethylation; endonuclease; granulocyte; hematopoietic differentiation; histone modification; inhibitor; insight; monocyte; peripheral blood; progenitor; programs; promoter; stem; stem cells; transcriptome sequencing

PROJECT SUMMARY/ABSTRACT Hematopoiesis is responsible for producing the varied cell types found in blood. In adults, this process primarily takes place in the bone marrow and is characterized by successive rounds of differentiation beginning with the hematopoietic stem cell through to lineage commitment. This process involves progressive narrowing of lineage potency as progenitors eventually commit to the production of a single cell lineage. Disruption of hematopoiesis can lead to benign and malignant pathologies. We focus on the bipotent megakaryocytic-erythroid progenitor (MEP), which has the potential to differentiate into a lineage committed erythroid progenitor (ErP) or a lineage committed megakaryocytic progenitor (MkP). MEP fate specification has been studied to a limited extent, uncovering only a small number of influences that contribute to this process. One understudied aspect of human MEP fate specification is the role of epigenetics. KDM1A (LSD1) is known to be important for erythroid maturation, but its role in MEP fate specification is unknown. Preliminary data using primary human cells show that inhibition of LSD1 in primary human ErP results in the ability of erythroid committed progenitors to undergo granulocytic-monocytic and megakaryocytic commitment. In contrast, LSD1 inhibition does not affect MkP commitment to the megakaryocytic fate. This suggests that LSD1 only antagonizes alternative lineage potential during erythropoiesis, and not megakaryopoiesis, even though LSD1 mRNA is expressed at similar levels in MEP, ErP, and MkP. The goal of this proposal is to establish the mechanism by which LSD1 promotes erythroid commitment while silencing alternative myeloid and megakaryocytic lineage potential. I will achieve these goals utilizing a mix of functional (cell culture), genetic and epigenetic approaches to determine where and how LSD1 regulates gene expression and fate specification in MEP, ErP, and MkP. Computational analyses will determine candidate gene targets, and potential genomic sites to force epigenetic modifications using different fusion proteins based on the catalytically inactive Cas9 endonuclease (dCas9). These results will establish the epigenetic mechanisms that govern erythroid lineage commitment mediated by LSD1. They will also shed light on the distinct lineage-specific epigenetic mechanisms that mediate and maintain fate specification in hematopoiesis.

项目概要/摘要 造血作用负责产生血液中发现的各种细胞类型。对于成年人来说，这个过程主要是 发生在骨髓中，其特征是从 造血干细胞直至谱系定型。这个过程涉及谱系的逐渐缩小 祖细胞最终致力于产生单细胞谱系的效力。造血破坏 可导致良性和恶性病变。我们专注于双能巨核细胞-红系祖细胞 （MEP），有可能分化成谱系定型红系祖细胞（ErP）或谱系 定型巨核细胞祖细胞（MkP）。 MEP 命运规范的研究范围有限， 仅揭示了促成这一过程的一小部分影响。人类尚未被研究的一个方面 MEP命运规范是表观遗传学的作用。众所周知，KDM1A (LSD1) 对于红系细胞很重要 成熟，但其在 MEP 命运规范中的作用尚不清楚。使用原代人类细胞的初步数据显示 抑制原代人 ErP 中的 LSD1 会导致红系定型祖细胞经历 粒细胞-单核细胞和巨核细胞的承诺。相反，LSD1 抑制不会影响 MkP 对巨核细胞命运的承诺。这表明 LSD1 仅拮抗替代谱系潜力 尽管 LSD1 mRNA 在红细胞生成期间而非巨核细胞生成期间以相似的水平表达 MEP、ErP 和 MkP。该提案的目标是建立LSD1促进红细胞生成的机制 承诺，同时沉默替代骨髓和巨核细胞谱系潜力。我将实现这些目标 利用功能（细胞培养）、遗传和表观遗传方法的组合来确定 LSD1 的位置和方式 调节 MEP、ErP 和 MkP 中的基因表达和命运规范。计算分析将确定 候选基因靶标和潜在的基因组位点，以使用不同的融合来强制进行表观遗传修饰 基于催化失活 Cas9 核酸内切酶 (dCas9) 的蛋白质。这些结果将确定 LSD1 介导的控制红系谱系定型的表观遗传机制。他们也将照亮 介导和维持命运规范的独特谱系特异性表观遗传机制 造血作用。