NOVEL REGULATORS OF LATE STAGE BONE MARROW ERYTHROBLAST DEVELOPMENT

晚期骨髓成红细胞发育的新型调节因子

基本信息

批准号：
8117782
负责人：
DON Michael WOJCHOWSKI
金额：
$ 21.7万
依托单位：
MAINEHEALTH
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-08-01 至 2013-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8117782
关键词：
Acquired Immunodeficiency Syndrome Adhesions Adult Affect Aging Anemia Attention Binding Bone Marrow CBL gene CD34 gene Cell Communication Cell Polarity Cell surface Cells Cellular Structures Cellular biology Complex Coupled Development Disease Employee Strikes Erythro Erythroblasts Erythrocytes Erythroid Erythroid Cells Erythropoiesis Event GATA1 gene Gene Expression Profile Genetic Growth and Development function Health Hour Human Inherited Investigation Island Kidney Diseases Knock-out Knockout Mice Maps Mass Spectrum Analysis Mediating Mediator of activation protein Molecular Molecular Mechanisms of Action Mus Myeloproliferation Nature Pathway interactions Physiological Population Process Production Proteomics Regulation Reticulocytes Role Serum Sickle Cell Anemia Staging Stromal Cells System T-Cell Development TRAF6 gene Testing Transcript Transgenic Mice Work base beta-1 Globin cancer therapy cell growth chemotherapy clinically relevant cohort in vivo insight mouse model novel progenitor public health relevance selective expression tissue oxygenation tumorigenesis ubiquitin ligase ubiquitin-protein ligase

项目摘要

DESCRIPTION (provided by applicant): Late stage erythroblast development is emerging as a sharply regulated process. New insight into key regulators has been won via aggressive genetic and cell biology approaches as recently applied to primary erythroid cell systems (with attention paid to the step-wise nature of erythroblast formation). Examples include Bcl11a and TR2/TR4 repressors of beta-globins (1, 2); miR-150, miR-144/451 as erythro-megakaryocytic transcript decay factors (3-5); and EMP plus ICAM4 as erythroblastic island bridging factors (6, 7). In ongoing studies of murine bone marrow (BM) erythropoiesis, the PI has observed a striking in vivo expansion capacity for a selective cohort of late-stage BM erythroblasts, specifically a KitnegCD71highTer119neg "stage E2" population. Towards characterizing this dynamic new cohort, we've optimized serum-free systems for murine BM erythroblast development, and have purified "stage E2" erythroblasts, their "stage-E1" KitposCD71highTer119neg progenitors, and "stage-E3" KitnegCD71highTer119pos progeny. Upon profiling each, we discovered two E3 ubiquitin ligase adaptors to be selectively expressed at high, sustained levels in late-stage murine and human BM erythroblasts (and subject to GATA1 regulation), TRIB3 pseudokinase and SPRY1. We've further constructed conditional Trib3-/- and Spry1-/- mouse models - and in each have observed selectively skewed erythropoiesis at steady-state; and markedly compromised erythropoiesis during anemia. In Aim #1, we now propose to define specific stages, and physiological conditions, during which TRIB3 and SPRY1 regulate erythroblast development. Effects on cellular pathways also will be investigated; and co-IP plus mass spectrometry approaches will be used to identify TRIB3 and SPRY1 complexed E3 ubiquitin ligases plus coupled erythroid targets. In Aim #2, we will apply transcriptome and phosphoproteome approaches to test the hypothesis that the above "stage E2" erythroblasts are dynamically advantaged in their expansion capacities via stage-specific expression of cell surface adhesion factors that mediate their selective interactions with a supporting BM stromal cell component. Studies promise to provide new insight into factors that (dys)regulate late-stage erythroblast and red cell production; and may uncover rational targets for new anti-anemia agents (as urgently needed in chemotherapy contexts)(8-10). PUBLIC HEALTH RELEVANCE: Hour-to-hour bone marrow production of red blood cells (RBC) is essential for tissue oxygenation. Faltered RBC formation results in health-compromising anemia as associated with cancer therapies, renal disease, aging, AIDS, and inherited RBC disorders (eg, sickle cell anemia). Proposed work promises to discover and characterize novel regulators of erythroblast and RBC production as potentially drugable E3 ligase complexes; and as a new RBC progenitor niche in bone marrow with unique expansion potential.

描述（由申请人提供）：后期的红细胞开发正在成为一个严格的监管过程。由于最近应用于原发性红细胞系统的侵略性遗传和细胞生物学方法，已经对关键调节剂进行了新的了解（注意红细胞形成的逐步性质）。示例包括β-球蛋白的BCL11A和TR2/TR4阻遏物（1，2）； miR-150，miR-144/451作为红细胞核细胞转录衰减因子（3-5）； EMP加ICAM4作为红细胞岛桥接因子（6，7）。在正在进行的鼠骨髓（BM）红细胞生成的研究中，PI观察到选择性阶段的BM后期红细胞，特别是Kitnegcd71Highter11 Highter119neg“ E2 e2 e2”的人群，观察到了体内膨胀能力引人注目。为了表征这种动态的新同类，我们为鼠BM粉红细胞的开发优化了无血清系统，并纯化了“ E2” e2'rythrosblasts，它们的“阶段 - E1” Kitposcd71Highter11 Highter119neg祖细胞和“ e3 stage-e3” kitnegcd71 Hightery119pos squorenyy。每个分析后，我们发现了两个E3泛素连接酶适配器在晚期和人类BM成年细胞（并受GATA1调节），Trib3 pseudokinase和spry1中以高持续水平的选择性表达。我们进一步构建了有条件的Trib3 - / - 和Spry1 - / - 小鼠模型 - 并且在稳态下观察到了选择性偏斜的红细胞生成。并在贫血期间明显损害了红细胞生成。在AIM＃1中，我们现在建议定义特定的阶段和生理条件，在此期间，Trib3和Spry1调节了红细胞的发育。还将研究对细胞途径的影响； Co-IP加质谱法将用于识别Trib3和Spry1复合的E3泛素连接酶以及耦合的红细胞靶标。在AIM＃2中，我们将采用转录组和磷蛋白组方法来检验以下假设：上述“ E2”红细胞通过阶段特异性表达在其扩展能力中动态优势，这些细胞表面粘附因子可以介导其选择性相互作用，从而介导其选择性相互作用，从而与支持BM Stromal stromal stromal细胞组件相互作用。研究有望提供有关（DYS）调节晚期红细胞和红细胞生产的因素的新见解；并可能发现新的抗血症药物的合理靶标（在化学疗法环境中迫切需要）（8-10）。公共卫生相关性：红细胞每小时的骨髓产生（RBC）对于组织氧合至关重要。步履蹒跚的RBC形成导致疾病促进性贫血与癌症疗法，肾脏疾病，衰老，艾滋病和遗传性RBC疾病有关（例如，镰状细胞贫血）。拟议的工作有望发现和将红细胞和RBC产生的新型调节剂视为潜在的可吸毒E3连接酶配合物。作为具有独特膨胀潜力的骨髓中的新RBC祖细胞生态位。