A Genetic Model of NMDA Receptor Function in Cell Death
细胞死亡中 NMDA 受体功能的遗传模型
基本信息
- 批准号:8433815
- 负责人:
- 金额:$ 26.05万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2013
- 资助国家:美国
- 起止时间:2013-02-01 至 2017-01-31
- 项目状态:已结题
- 来源:
- 关键词:AffectAlcohol abuseAlcoholsAlzheimer&aposs DiseaseAnimal ModelAntibodiesApoptosisApoptoticAutophagocytosisBacteriaBindingBinding ProteinsBiochemicalBiochemical GeneticsBiological AssayBiological MetamorphosisBiological ModelsBuffersCalciumCell DeathCell Death Signaling ProcessCellsCessation of lifeComplementDataDependenceDevelopmentDiseaseDissectionDominant-Negative MutationDown-RegulationDrosophila genusDrosophila melanogasterEventExcisionGenesGeneticGenetic ModelsGlutamate ReceptorHomologous GeneHumanInjuryMAP Kinase GeneMalignant NeoplasmsMeasuresMediatingMedicalMitochondriaModelingMolecular ProfilingMutateN-Methyl-D-Aspartate ReceptorsNerve TissueNervous system structureNeurodegenerative DisordersNeurotoxinsOrganParkinson DiseasePathway interactionsPhospho-Specific AntibodiesPhosphotransferasesProcessProtein DephosphorylationProtein Tyrosine PhosphataseProteinsPublic HealthRNARNA InterferenceReceptor ActivationReceptor Protein-Tyrosine KinasesReceptor SignalingResearchRoleSalivary GlandsSignal PathwaySignal TransductionSiteStaining methodStainsStrokeSystemTestingTransgenic AnimalsTransgenic OrganismsTumor Suppressor ProteinsTyrosineUp-Regulationaging populationalcohol exposurebaseflyin vivo Modelinsightneuron lossnovelpreventprotein functionpublic health relevancereceptorreceptor functionresearch studyresponsetherapeutic developmenttooluptake
项目摘要
DESCRIPTION (provided by applicant): Excitatory cell death is responsible for the loss of nervous tissue in neurodegenerative disorders such as Alzheimer's or Parkinson's disease and after insults to the nervous system, for instance through stroke or alcohol abuse. This type of cell death is caused by an overstimulation of calcium uptake through glutamate receptors, in particular the N-methyl-D-aspartate receptor (NMDAR). Despite the medical importance of excitatory cell death, a coherent model of the events downstream of receptor activation has been slow to emerge. Progress has been hampered by the complexity of cell death under pathological conditions and the lack of an accessible animal model that allows a genetic dissection of the process. This project is based on the discovery that NMDAR is required for non-pathological programmed cell death in the genetic model organism Drosophila melanogaster. Death of the larval salivary glands during metamorphosis, which constitutes an important model system in cell death research, fails when NMDAR activity is reduced. This opens up the possibility to study the role of NMDAR in cell death using the powerful tools and approaches of model organism research. As a first result of this research, the protein tyrosine phosphatase Ptpmeg was identified as a putative partner of NMDAR in cell death control. Salivary glands deficient in Ptpmeg do not die and carry a lowered calcium load. Cell death control by Ptpmeg is a novel function that likely extends to its human homologs, which are dysregulated in cancers and suspected to be tumor suppressors. The broad objective of this project is to determine how NMDAR and Ptpmeg cooperate in cell death control and how the two proteins affect steps in the evolutionarily conserved cell death program. Specifically, the project will (1) use genetic approaches in transgenic animals to determine how NMDAR and Ptpmeg cooperate in regulating intracellular Ca2+ and cell death. Co- immunolocalization and pull-down assays will be used to examine physical interactions between NMDAR and Ptpmeg. Staining with a phospho-specific antibody will reveal the expression profile of the tyrosine-phosphorylated form of NMDAR and its dependence on Ptpmeg. (2) The role of Ca2+ in cell death will be examined in transgenic animals using a Ca2+-buffer protein. Downstream targets of NMDAR signaling will be identified by analyzing the effects of reduced NMDAR activity on Ca2+-dependent cell clearance pathways and markers of apoptosis and autophagy. (3) Biochemical and genetic assays will be used to identify steps in apoptotic and Ca2+-signaling pathways that are affected by a lack of Ptpmeg. Together, the results of the proposed research are likely to bring to light evolutionarily conserved, basic mechanisms of cell death control by NMDAR and Ptpmeg. Thus, the project will help understand the function of these proteins under pathological conditions and aid in the development of therapeutic measures.
描述(由申请人提供):兴奋性细胞死亡是神经退行性疾病(如阿尔茨海默病或帕金森病)中神经组织损失的原因,并且在神经系统受到损伤(例如通过中风或酒精滥用)后。这种类型的细胞死亡是由通过谷氨酸受体,特别是N-甲基-D-天冬氨酸受体(NMDAR)的钙摄取的过度刺激引起的。尽管兴奋性细胞死亡的医学重要性,一个连贯的模型下游的受体激活的事件一直缓慢出现。由于病理条件下细胞死亡的复杂性以及缺乏一种可以对该过程进行遗传解剖的动物模型,进展受到阻碍。该项目基于以下发现:NMDAR是遗传模式生物黑腹果蝇(Drosophila melanogaster)中非病理性程序性细胞死亡所必需的。在变态过程中的幼虫唾液腺的死亡,这构成了一个重要的模型系统在细胞死亡的研究,失败时,NMDAR活性降低。这开辟了使用模式生物研究的强大工具和方法来研究NMDAR在细胞死亡中的作用的可能性。作为本研究的第一个结果,蛋白酪氨酸磷酸酶Ptpmeg被鉴定为细胞死亡控制中NMDAR的推定伴侣。唾液腺缺乏Ptpmeg不会死亡,并携带降低的钙负荷。Ptpmeg的细胞死亡控制是一种新的功能,可能延伸到其人类同源物,这些同源物在癌症中失调,并被怀疑是肿瘤抑制因子。该项目的主要目标是确定NMDAR和Ptpmeg如何在细胞死亡控制中合作,以及这两种蛋白质如何影响进化保守的细胞死亡程序中的步骤。具体而言,该项目将(1)在转基因动物中使用遗传方法来确定NMDAR和Ptpmeg如何在调节细胞内Ca 2+和细胞死亡中合作。将使用共免疫定位和下拉测定来检查NMDAR和Ptpmeg之间的物理相互作用。用磷酸特异性抗体染色将揭示NMDAR的酪氨酸磷酸化形式的表达谱及其对Ptpmeg的依赖性。(2)将使用Ca 2+缓冲蛋白在转基因动物中检查Ca 2+在细胞死亡中的作用。将通过分析降低的NMDAR活性对Ca 2+依赖性细胞清除途径以及凋亡和自噬标志物的影响来鉴定NMDAR信号传导的下游靶标。(3)生化和遗传分析将被用来确定的步骤,细胞凋亡和Ca 2+信号通路的影响,缺乏Ptpmeg。总之,拟议研究的结果可能会揭示NMDAR和Ptpmeg控制细胞死亡的进化保守的基本机制。因此,该项目将有助于了解这些蛋白质在病理条件下的功能,并有助于开发治疗措施。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
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MICHAEL LEHMANN其他文献
MICHAEL LEHMANN的其他文献
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{{ truncateString('MICHAEL LEHMANN', 18)}}的其他基金
The control of growth and metabolism by effectors of the TOR signaling pathway
TOR 信号通路效应子对生长和代谢的控制
- 批准号:
7304845 - 财政年份:2007
- 资助金额:
$ 26.05万 - 项目类别:
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