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

基本信息

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
项目类别：
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

项目摘要

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（由适用提供）：红细胞祖细胞在承诺切换到红细胞分化之前经历自我更新的部门，但是调节红细胞祖细胞生长的途径仍然在很大程度上未知。在对以确定性（成人）但不是原始（胚胎）红细胞谱系表达的基因的计算搜索中，我们鉴定 D3（1,25（OH）2d3）。实时RT-PCR分析表明，VDR在小鼠胎儿肝脏和骨髓的确定红细胞祖细胞中表达，在红细胞成熟过程中被下调。结构研究表明，维生素D3配体的VDR激活会导致显着的构象变化，从而稳定蛋白质并诱导其转运到核中，在该细胞核中募集了核调节络合物。 VDR信号通路主要是在骨骼中研究的。通过这一途径对红细胞生成的调节本质上是出乎意料的。已发表的研究几乎完全在白血病细胞系（不是正常的原代细胞）中进行。我们发现，维生素D3刺激小鼠胎儿肝脏和骨髓的红斑祖细胞（BFU-E和CFU-E）的生长。当激活VDR途径时，不仅数字，而且BFU-E菌落的大小也会增加。包含BFU-E的C-KIT+胎儿肝祖细胞的CD71LOW子集对VDR的激活最敏感。用维生素D3培养的祖细胞正常区分。维生素D3可以在祖细胞培养物中部分代替地塞米松（一种糖皮质激素），这表明在胁迫红细胞生成中可能起作用。我们假设VDR激活或反映了与红细胞转录因子（TFS）（例如GATA，KLF1/EKLF和SCL/TAL）结合使用的红细胞祖细胞中的基因，以及VDR的DNA结合位点可能会与Erythroid TFS的结合位点聚集。基于此假设的计算方法用于鉴定红细胞祖细胞中的候选VDR靶基因。我们已经开始使用实时RT-PCR确认其中一些基因的维生素D3反应性。候选者包括已知在红细胞生成（C-MYC，GATA2）和其他没有的基因（钙结合激酶CAMK1D; N-MYC; N-MYC； MLX相互作用蛋白，BHLH转录因子； GRTP1，GRTP1，GTPase RAB激活剂）。我们假设VDR途径控制红细胞祖细胞的增殖和/或存活，至少部分通过靶基因的转录调节。该项目的目的是阐明维生素D调节红细胞发育的分子机制。