Quantitative Proteomic Reconfiguration in Induction of Neuroprotection against St

诱导神经保护作用的定量蛋白质组重构

• 批准号: 8269878
• 负责人: AN ZHOU
• 金额: $ 17.69万
• 依托单位: MOREHOUSE SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-01 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8269878
• 关键词: Anabolism; Antibodies; Bioinformatics; Biological; Brain; Chemistry; Complex; Development; Drosophila genus; Elements; Epigenetic Process; Exposure to; Gene Expression; Gene Proteins; Genes; Genetic Transcription; Genomics; Goals; Histones; Immunoprecipitation; In Vitro; Injury; Ischemia; Ischemic Brain Injury; Ischemic Stroke; Label; Mass Spectrum Analysis; Measures; Mediating; Metabolic; Modeling; Molecular; Molecular Biology; Planet Mars; Polycomb; Property; Protein Biosynthesis; Proteins; Proteome; Proteomics; Regulation; Repressor Proteins; Resistance; Rodent; Science; Signal Transduction; Spectrophotometry; Stroke; Techniques; Testing; Therapeutic; Transcription Process; in vivo; knock-down; neuroprotection; new therapeutic target; novel; preconditioning; protein function; response; stoichiometry; therapeutic target; tool

DESCRIPTION (provided by applicant): Brief "preconditioning" ischemia results in "tolerance" to subsequent ischemic brain injury. The genomic signature of the tolerant brain is transcriptional suppression. The translational elements of this protein synthesis-dependent phenomenon of tolerance have been unknown. However, we now show the tolerance proteome (Stapels et al. Science Signaling, Mar 2010). The proteome is enriched in histone proteins and remarkably in Polycomb group (PcG) proteins whose molecular functions are those of transcriptional suppression. Thus a mechanism of induction of transcriptional suppression, characterizing tolerance, may have been discovered. This discovery implicates epigenetic gene repressor proteins (PcGs) as responsible for the process of transcription suppression resulting in ischemic tolerance. Further, we show that PcGs, previously known as regulators of segmentation during development in drosophila, have a novel neuroprotective function in brain. Our initial studies in ischemic-tolerance in vivo and in vitro show that the development of ischemic tolerance is dependent upon the expression of PcG proteins: knock down ablates tolerance, and over-expression produces tolerance. Further, we now have preliminary evidence in ischemic tolerance for opposing changes of Trithorax group (TrxG) proteins, the gene activator and antagonist of PcG proteins. Accordingly we will investigate PcG/TrxG epigenetic regulation to determine their effector properties upon the tolerance mechanism. We offer the following aims: Specific Aim 1. To establish in absolute molar numbers, the stoichiometry of proteins that comprise PcG/TrxG epigenetic regulator complexes at the onset of ischemic injury and the onset of ischemic tolerance. Quantitative MS analyses will be performed on brain proteins immunoprecipitated with specific antibodies against selected PcG or TrxG proteins. Specific Aim 2. To establish the identity of newly synthesized proteins in the brain at the onset of ischemic injury and the onset of ischemic tolerance. We will employ the Click technique to metabolically label the newly synthesized protein, isolate them, and quantify them with quantitative MS analyses. Bioinformatics of the nascent proteomes will be established with the assistance of bioinformatic tools.

描述(申请人提供)：短暂的“预适应”脑缺血导致对随后的脑缺血损伤的“耐受性”。耐受脑的基因组特征是转录抑制。这种依赖蛋白质合成的耐受现象的翻译成分尚不清楚。然而，我们现在展示了耐受蛋白质组(Stapels等人)。科学信号，2010年3月)。蛋白质组富含组蛋白，尤其是聚梳组蛋白，其分子功能是转录抑制。因此，可能已经发现了一种诱导转录抑制的机制，即表征耐受性的机制。这一发现表明，表观基因抑制蛋白(PCGS)参与了导致缺血耐受的转录抑制过程。此外，我们还证明了PCGS在果蝇发育过程中具有新的神经保护功能，以前被认为是果蝇发育过程中分段的调节器。我们在体内和体外对缺血耐受的初步研究表明，缺血耐受的发展依赖于PcG蛋白的表达：下调导致耐受，过度表达产生耐受。此外，我们现在有初步证据表明，对于PcG蛋白的基因激活剂和拮抗剂TrxG蛋白的变化，我们对缺血耐受有初步的研究。因此，我们将研究PcG/TrxG的表观遗传调控，以确定它们在耐受机制上的效应特性。我们提供了以下目标：1.以绝对摩尔数建立组成PcG/TrxG表观遗传调节复合体的蛋白质在缺血损伤和缺血耐受开始时的化学计量比。将对免疫沉淀的大脑蛋白进行定量的MS分析，这些蛋白与选定的PcG或TrxG蛋白有特定的抗体。具体目的2.确定脑内新合成的蛋白质在脑缺血损伤和脑缺血耐受开始时的同一性。我们将使用Click技术对新合成的蛋白质进行代谢性标记，分离它们，并用定量MS分析对它们进行定量。在生物信息学工具的帮助下，将建立新生蛋白质组的生物信息学。