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: 8331642
• 负责人: Adam N. Goldfarb
• 金额: $ 5.6万
• 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 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8331642
• 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）在生生性红细胞生成中起核心作用，是治疗人类贫血的主要药物。与Epo给药相关的两个主要问题是治疗耐药性和不良副作用。治疗耐药通常发生在靶细胞水平，即红系祖细胞。Epo最突出的不良副作用包括心血管并发症，特别是充血性心力衰竭（CHF）。虽然促生成素信号已经研究了几十年，但这些重要临床问题的机制基础仍然未知。此外，Epo信号如何促进红细胞分化的基本问题本身仍未解决。十多年来，人们已经知道Epo以某种方式调节红系基因转录的主调控因子GATA-1的功能。在缺乏Epo信号的情况下，GATA-1的激活导致细胞无分化死亡。虽然已经提出了几种关于Epo调控GATA-1的模型，但尚未建立机制。这一建议为Epo通过蛋白激酶D/ IIa类组蛋白去乙酰化酶（PKD/HDAC）途径调控GATA-1提供了证据，建立在先前IIa类HDAC对GATA-1结合和抑制的观察基础上。在G1ER和人红细胞祖细胞中都获得了这一证据，包括Epo激活PKD和通过PKD抑制阻断Epo驱动的红细胞分化。进一步的证据包括通过HDAC抑制剂敲低HDAC5和消除GATA-1对Epo信号的依赖，增强红细胞的Epo反应性。重要的是，IIa类HDAC信号通路与心脏肥大和CHF的病理生理有关，其中IIa类HDAC被PKD或CaMK磷酸化会导致肌源性转录因子MEF2的不适当激活。因此，Epo激活PKD可能促进gata -1驱动的红细胞生成和mef2驱动的CHF。在本提案中，我们将确定Epo激活PKD的机制，检查EpoR及其协同调节因子c-Kit的结构要求。此外，我们还将分析必不可少的EpoR下游介质JAK2和PLC3对PKD激活的作用。我们还将研究Epo给药是否会诱导心肌中PKD的激活，如果是的话，哪些Epo相关元素有助于这种信号传导。因此，这些实验将描述一种新的Epo信号通路的上游成分，该通路涉及红细胞生成的编程，并可能诱导CHF。从这些研究中获得的信息将为探索临床促生成素耐药机制和设计逆转促生成素耐药的药物建立一个概念框架。同样重要的是，这些实验将从机制上解决促红细胞生成素与心血管并发症的关联这一尚未解决的问题。本提案是响应计划公告“刺激血液学研究：新的努力”（PAS-10-046）。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红细胞表达和功能”。本项目的一个重要组成部分将致力于分析野生型和突变型EpoR小鼠以及野生型和突变型c-Kit小鼠中Epo给药对心肌PKD的激活。另一个主题是“炎症性贫血和慢性疾病”。提高对正常红细胞EpoR信号的理解是理解与这些贫血相关的Epo耐药机制的关键的第一步。