Regulation of Erythropoiesis by the VDR Nuclear Receptor Transcription Factor

VDR 核受体转录因子对红细胞生成的调节

• 批准号: 9260873
• 负责人: Margaret H Baron
• 金额: $ 38.14万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-06 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9260873
• 关键词: Adult; BFU-E; Binding; Binding Sites; Biological Assay; Bone Marrow; CFU-E; Calcitriol; Calcium Binding; Cell Cycle; Cell Line; Cell Lineage; Cell Nucleus; Cell Proliferation; Cells; Cholecalciferol; Clinical; Complex; DNA Binding; Development; Dexamethasone; Embryo; Erythroblasts; Erythrocyte Transfusion; Erythrocytes; Erythroid; Erythroid Progenitor Cells; Erythropoiesis; Fetal Liver; Gene Targeting; Genes; Genetic; Genetic Transcription; Glucocorticoids; Goals; Growth; Human; Individual; Leukemic Cell; Ligands; Liquid substance; Methylcellulose; Molecular; Molecular Analysis; Molecular Conformation; Mus; Myelogenous; Nuclear Receptors; Pathway interactions; Phenotype; Phosphotransferases; Population; Production; Proteins; Proto-Oncogene Protein c-kit; Publishing; RNA; Receptor Activation; Receptor Signaling; Recruitment Activity; Regulation; Reverse Transcriptase Polymerase Chain Reaction; Role; Signal Pathway; Signal Transduction; Spleen; Stem cells; Stress; Subfamily lentivirinae; Time; Transcript; Transcriptional Regulation; Vitamin D; Vitamin D3 Receptor; base; bone; c-myc Genes; erythroid differentiation; knock-down; loss of function; next generation; novel; novel strategies; novel therapeutics; progenitor; public health relevance; rab GTP-Binding Proteins; response; self-renewal; small hairpin RNA; stem cell differentiation; transcription factor; transcriptome sequencing; virtual

 DESCRIPTION (provided by applicant): Red blood cell progenitors undergo self-renewing divisions prior to the commitment switch to erythroid differentiation but the pathways that regulate erythroid progenitor growth are still largely unknown. In a computational search for genes expressed in definitive (adult) but not primitive (embryonic) red cell lineages, we identifie the nuclear receptor transcription factor VDR, which is activated by binding to its ligand, vitamin D3 (1,25(OH)2D3). Real-time RT-PCR analysis indicated that Vdr is expressed in definitive erythroid progenitors from mouse fetal liver and bone marrow and is downregulated during erythroid maturation. Structural studies have shown that VDR activation by the vitamin D3 ligand results in significant conformational changes that stabilize the protein and induce its translocation into the nucleus, where it recruits coregulatory complexes. The VDR signaling pathway has been studied mostly in bone. The regulation of erythropoiesis by this pathway has been essentially unexplored; published studies were performed almost entirely in leukemic cell lines (not normal primary cells). We find that vitamin D3 stimulates the growth of erythroid progenitors (BFU-E and CFU-E) from mouse fetal liver and bone marrow. Not only the numbers but also the size of the BFU-E colonies is increased when the VDR pathway is activated. The CD71low subset of c-Kit+ fetal liver progenitors, which contains BFU-E, is the most sensitive to activation of VDR. Progenitors cultured with vitamin D3 differentiate normally. Vitamin D3 can partially substitute for dexamethasone (a glucocorticoid) in progenitor cultures, suggesting a possible role in stress erythropoiesis. We hypothesized that VDR activates or represses genes in developing erythroid progenitors in combination with erythroid transcription factors (TFs) such as Gata, Klf1/Eklf, and Scl/tal and that DNA binding sites for VDR may cluster with binding sites for erythroid TFs. A computational approach based on this hypothesis was used to identify candidate VDR target genes in erythroid progenitors. We have begun to confirm the vitamin D3 responsiveness of some of these genes using real-time RT-PCR. The candidates include genes known to function in erythropoiesis (c-Myc, Gata2) and others that were not (Calcium binding kinase Camk1d; N-Myc; Mlx-interacting protein, a bHLH transcription factor; Grtp1, GTPase Rab activator). We hypothesize that the VDR pathway controls erythroid progenitor cell proliferation and/or survival, at least in part through transcriptional regulation of target genes. The goal of this project is to elucidate the molecular mechanisms by which vitamin D regulates red blood cell development.

 描述(申请人提供)：红血球祖细胞在承诺转变为红系分化之前经历自我更新分裂，但调节红系祖细胞生长的途径在很大程度上仍不清楚。在对确定的(成人)但不是原始的(胚胎)红细胞谱系中表达的基因的计算机搜索中，我们鉴定了核受体转录因子VDR，它通过与其配体维生素结合而激活 D3(1，25(OH)2D3)。实时荧光定量RT-PCR分析表明，VDR在小鼠胎肝和骨髓的红系祖细胞中表达，在红系成熟过程中表达下调。结构研究表明，维生素D3配体激活VDR会导致显著的构象变化，稳定蛋白质并诱导其移位到核中，在那里它招募共调节复合体。目前对VDR信号通路的研究主要集中在骨骼。这一途径对红细胞生成的调控基本上还未被探索；已发表的研究几乎全部在白血病细胞系(而不是正常原代细胞)中进行。我们发现维生素D3能刺激小鼠胎肝和骨髓红系祖细胞(BFU-E和CFU-E)的生长。当VDR途径被激活时，BFU-E集落的数量和大小都会增加。含有BFU-E的c-Kit+胎肝祖细胞的CD71低亚群对VDR的激活最为敏感。用维生素D3培养的祖细胞分化正常。在祖细胞培养中，维生素D3可以部分替代地塞米松(一种糖皮质激素)，这表明在应激性红细胞生成中可能起到了作用。我们推测VDR与GATA、KLF1/EKLF和SCL/TAL等红系转录因子(TF)一起激活或抑制红系祖细胞发育过程中的基因，VDR的DNA结合部位可能与红系转录因子的结合部位聚集。基于这一假设的计算方法被用来在红系祖细胞中识别候选的VDR靶基因。我们已经开始使用实时RT-PCR来确认其中一些基因对维生素D3的反应性。候选基因包括已知在红细胞生成中起作用的基因(c-Myc，Gata2)和其他不起作用的基因(钙结合激酶Camk1d；N-Myc；MLX相互作用蛋白，一种bHLH转录因子；Grtp1，GTP酶Rab激活剂)。我们假设VDR途径控制红系祖细胞的增殖和/或存活，至少部分是通过对靶基因的转录调控。 该项目的目标是阐明维生素D调节红细胞发育的分子机制。