Characterization of a Novel Erythropoietin Signaling Pathway

新型促红细胞生成素信号通路的表征

• 批准号: 8150951
• 负责人: Adam N. Goldfarb
• 金额: $ 30.49万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-30 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8150951
• 关键词: Address; Adverse effects; Anemia; Anemia due to Chronic Disorder; Binding; CD34 gene; Calcium/calmodulin-dependent protein kinase; Cardiovascular system; Cell Death; Cells; Chimera organism; Chronic Disease; Clinical; Clinical Trials; Congestive Heart Failure; Dependence; Dose; Elements; Engineering; Erythrocytes; Erythroid; Erythroid Cells; Erythroid Progenitor Cells; Erythropoiesis; Erythropoietin; Erythropoietin Receptor; Functional disorder; Genetic Transcription; HDAC5 gene; Heart Diseases; Heart Hypertrophy; Hematology; Histone Deacetylase; Histone Deacetylase Inhibitor; Human; In Vitro; Investigation; JAK2 gene; Maps; Mediator of activation protein; Modeling; Mus; Mutagenesis; Myocardial; Myocardium; NIH Program Announcements; Pathway interactions; Patients; Phosphorylation; Phosphotransferases; Physiological; Play; Production; Prolactin; Pronormoblasts; Protein Isoforms; Proto-Oncogene Protein c-kit; Publishing; Regulation; Repression; Resistance; Resources; Role; Signal Pathway; Signal Transduction; Stem Cell Factor; Therapeutic Studies; Tyrosine; base; cardiovascular disorder risk; cytokine; design; erythroid differentiation; gain of function; human GATA1 protein; improved; in vivo; loss of function; mutant; novel; progenitor; programs; protein kinase D; public health relevance; research study; small hairpin RNA; transcription factor

DESCRIPTION (provided by applicant): The soluble cytokine erythropoietin (Epo) plays a central role in physiologic erythropoiesis and is a mainstay in treatment of human anemias. Two major problems associated with Epo administration are treatment resistance and adverse side-effects. Treatment resistance usually occurs at the level of the target cells, the erythroid progenitors. The most prominent adverse side-effects of Epo consist of cardiovascular complications, particularly congestive heart failure (CHF). Although Epo signaling has been studied for several decades, the mechanistic basis for these important clinical problems remains unknown. Furthermore, the basic question of how Epo signaling promotes erythroid differentiation remains itself unresolved. It has been known for over a decade that Epo somehow regulates the function of the master regulator of erythroid gene transcription, GATA-1. Activation of GATA-1 in the absence of Epo signaling causes cell death without differentiation. Although several models have been proposed for Epo regulation of GATA-1, no mechanism has been established. This proposal provides evidence for Epo regulation of GATA-1 through a protein kinase D/class IIa histone deacetylase (PKD/HDAC) pathway, building on previous observations of GATA-1 binding and repression by class IIa HDACs. This evidence, obtained in both G1ER and human erythroid progenitors, includes the activation of PKD by Epo and the blockade of Epo-driven erythroid differentiation through PKD inhibition. Further evidence consists of the enhancement of erythroid Epo responsiveness through HDAC5 knockdown and elimination of GATA-1 dependence on Epo signaling by an HDAC inhibitor. Importantly, class IIa HDAC signaling pathways have been implicated in the pathophysiology of cardiac hypertrophy and CHF, wherein class IIa HDAC phosphorylation by PKD or CaMK causes inappropriate activation of the myogenic transcription factor MEF2. Thus Epo activation of PKD may promote both GATA-1-driven erythropoiesis and MEF2-driven CHF. In this proposal, we will determine the mechanism for Epo activation of PKD, examining the structural requirements of the EpoR and its coregulator, c-Kit. In addition, the essential EpoR downstream mediators, JAK2 and PLC3, will be analyzed for their contributions to PKD activation. We will also examine whether Epo adminstration induces PKD activation in vivo in myocardium and, if so, which EpoR-associated elements contribute to this signaling. These experiments will thus delineate the upstream components of a novel Epo signaling pathway involved in programming of erythropoiesis and potentially in induction of CHF. The information gained from these studies will establish a conceptual framework for probing mechanisms of clinical Epo resistance and for design of pharmacologic agents to reverse Epo resistance. Of equal importance, these experiments will mechanistically address the unresolved issue of Epo's association with cardiovascular complications. This proposal is responsive to the program announcement "Stimulating Hematology Investigation: New Endeavors" (PAS-10-046). One specific topic listed in this announcement is "Non-erythroid Expression and Function of Erythropoietin Receptors." A significant component of this project will be devoted to analysis of myocardial PKD activation by Epo administration in mice with wild type and mutant EpoR, as well as in mice with wild type and mutant c-Kit. Another topic listed is "Anemia of Inflammation and Chronic Disease." Improved understanding of normal erythroid EpoR signaling is a critical first step toward understanding the mechanisms of Epo resistance associated with these anemias. PUBLIC HEALTH RELEVANCE: Erythropoietin (Epo) is critical for normal red cell production and is administered as a treatment to numerous patients with anemia. A published, large-scale clinical trial has shown that giving patients sufficient doses of Epo to completely correct anemia, rather than partially correct anemia, confers an increased risk of cardiovascular disease, particularly congestive heart failure (CHF). Despite the tremendous amount of resources invested in Epo therapy and research, relatively little is understood about how it may promote red cell production as well as heart disease. In this proposal, a new signaling pathway, that can potentially explain both the beneficial and deleterious effects of Epo, will be characterized.

描述（由申请人提供）：可溶性细胞因子促红细胞生成素（EPO）在生理红细胞生成中起着核心作用，并且是治疗人类贫血的中流型。与EPO给药相关的两个主要问题是治疗性和不良副作用。治疗耐药性通常发生在靶细胞水平，即红细胞祖细胞。 EPO的最突出的不良副作用包括心血管并发症，尤其是充血性心力衰竭（CHF）。尽管已经研究了EPO信号几十年，但这些重要的临床问题的机械基础仍然未知。此外，EPO信号如何促进红细胞分化的基本问题仍未解决。十多年来，EPO以某种方式调节了红细胞基因转录GATA-1的主调节剂的功能。在没有EPO信号传导的情况下，GATA-1的激活会导致细胞死亡而没有分化。尽管已经提出了几种模型来调节GATA-1，但尚未建立机制。 该建议提供了通过蛋白激酶D/IIA类组蛋白脱乙酰基酶（PKD/HDAC）途径对GATA-1调节的证据，这是基于先前观察到IIA HDACS的GATA-1结合和抑制作用的基础。在G1ER和人类红细胞祖细胞中获得的这一证据包括通过EPO激活PKD，以及通过PKD抑制而封锁了EPO驱动的红细胞分化。进一步的证据包括通过HDAC5敲低和消除HDAC抑制剂对EPO信号的依赖性来增强红系EPO响应能力。重要的是，IIA类HDAC信号通路已与心脏肥大和CHF的病理生理有关，其中PKD或CAMK的IIA类HDAC磷酸化导致肌源性转录因子MEF2的不适当激活。因此，PKD的EPO激活可能会促进GATA-1驱动的红细胞生成和MEF2驱动的CHF。 在此提案中，我们将确定PKD激活的EPO激活机制，以检查EPOR及其核心基因c-Kit的结构要求。此外，将分析基本的EPOR下游介质JAK2和PLC3，以分析其对PKD激活的贡献。我们还将检查EPO守卫在心肌体内是否诱导PKD激活，如果是的，则与EPOR相关的元件有助于这种信号。因此，这些实验将描绘出与促红细胞生成和诱导CHF进行编程的新型EPO信号传导途径的上游组成部分。从这些研究中获得的信息将建立一个概念框架，用于探测临床EPO耐药性的机制以及设计药理学剂以逆转EPO耐药性。同样重要的是，这些实验将机械地解决EPO与心血管并发症相关的尚未解决的问题。 该提案对计划公告“刺激血液学调查：新努力”（PAS-10-046）的响应敏感。本公告中列出的一个特定主题是“促红细胞生成素受体的非果蝇表达和功能”。该项目的一个重要组成部分将致力于分析具有野生型和突变EPOR的小鼠以及具有野生型和突变型C-KIT的小鼠中EPO给药的心肌PKD激活。列出的另一个主题是“炎症和慢性疾病的贫血”。对正常红细胞EPOR信号传导的理解的提高是了解与这些贫血相关的EPO抗性机制的关键第一步。 公共卫生相关性：促红细胞生成素（EPO）对于正常的红细胞生产至关重要，并作为对众多贫血患者进行治疗。一项已发表的大规模临床试验表明，给患者提供足够剂量的EPO，以完全纠正贫血，而不是部分纠正贫血，使心血管疾病的风险增加，尤其是充血性心力衰竭（CHF）。尽管在EPO疗法和研究中投入了大量资源，但关于如何促进红细胞生产以及心脏病的了解相对较少。在此提案中，将表征一个新的信号通路，可以解释EPO的有益和有害影响。