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Control of Erythropoiesis by the Oxygen Sensor PHD2

通过氧传感器 PHD2 控制红细胞生成

基本信息

批准号：
9751846
负责人：
FRANK S LEE
金额：
$ 36万
依托单位：
UNIVERSITY OF PENNSYLVANIA
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：
2015-09-21 至 2021-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9751846
关键词：
Address Altitude Amino Acid Substitution Anemia Attenuated Binding Binding Sites Catalytic Domain Cells Chelating Agents Chronic Client Dependence Disease EPO gene Erythrocytes Erythroid Erythropoiesis Erythropoietin Receptor Event Gene Expression Genes Genetic Transcription Genetic study Heat-Shock Proteins 90 Homeostasis Hydroxylation Hypersensitivity Hypoxia Hypoxia Inducible Factor Kidney Knock-in Mouse Knowledge Left Mediating Modeling Modification Molecular Mus Mutagenesis Mutate Myelogenous Nature Oxygen Pathway interactions Procollagen-Proline Dioxygenase Protein Isoforms Proteins Reaction Red Cell Mass result Regulation Response Elements Role Signaling Protein Stem cells Tertiary Protein Structure Testing Therapeutic Tissues Translating Up-Regulation VHL protein Variant Zinc Zinc Fingers base bone deaf experimental study flexibility kidney cell novel progenitor receptor recruit sensor transcription factor

项目摘要

PROJECT SUMMARY/ABSTRACT A key event in transducing changes in oxygen concentration to changes in red cell mass is Prolyl Hydroxylase Domain protein 2 (PHD2)-catalyzed hydroxylation of the transcription factor, Hypoxia Inducible Factor-2α (HIF-2α). When oxygen is plentiful, constitutive hydroxylation of HIF-2α by PHD2 targets the former for degradation. When oxygen is lacking, PHD2 activity is attenuated, leading to the stabilization of HIF-2α and the activation of the ERYTHROPOIETIN (EPO) gene in the kidney. Circulating EPO then binds to the EPO receptor on red cell progenitors to expand red cell mass. Our understanding of PHD2 is still at a very early stage. PHD2 contains two evolutionarily conserved domains, a prolyl hydroxylase domain that catalyzes the hydroxylation reaction, and a predicted zinc finger domain. Very little is known about the latter, which, it may be noted, is lacking in the other two mammalian PHD paralogues. We have recently found that the zinc finger domain, which is of the Myeloid Nervy Deaf (MYND) type, binds with high stringency to a Pro-Xaa-Leu-Glu motif that is present in select cochaperones of the HSP90 pathway, such as p23. The HIF-α's (of which HIF-1α and HIF-2α are the major isoforms) are client proteins of the HSP90 pathway, and this therefore leads to a model in which p23 recruits PHD2 to HSP90 to promote HIF-α hydroxylation. Thus, the zinc finger of PHD2 is proposed to serve a positive regulatory role. Key unanswered questions are whether the zinc finger of PHD2 is actually of the MYND-type, and whether its function is relevant to erythropoiesis and more generally, oxygen homeostasis. We propose to address these questions by pursuing the following Specific Aims. In Specific Aim 1, we will conduct point mutagenesis studies on the zinc finger of PHD2 to determine if residues that might be predicted to be functional important based on homology to known MYND zinc fingers are indeed important for interaction with the Pro-Xaa-Leu-Glu motif of p23. In Specific Aim 2, we will characterize a knockin mouse line in which two predicted zinc chelating residues have been mutated (C36S/C42S) in order to ablate zinc finger function. We will examine this mouse for dysregulation of erythropoiesis, and cross it with Hif-1α and Hif-2α heterozygous deficient mice to determine Hif-α dependency. In Specific Aim 3, we will determine whether erythroid progenitors obtained from these mice display hypersensitivity to Epo, which would point to an Epo-independent role for this zinc finger that is distinct from the regulation the Epo gene itself. We will also examine whether there is a broader upregulation of Hif target genes in tissues of these mice. Collectively, we anticipate that these studies will provide critical information for PHD2, a key signaling protein that regulates red cell mass.

项目总结/文摘