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: 8168421
• 负责人: AN ZHOU
• 金额: $ 21.23万
• 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 2013-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8168421
• 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. PUBLIC HEALTH RELEVANCE: We show novel proteins discovered in brain made tolerant to ischemia (stroke). We hypothesize that they are epigenetic regulators proteins (i.e. the brains genes aren't changed but their response is changed). We will perform molecular biology analyses to test this hypothesis.

描述（由申请人提供）：短暂的“预处理”缺血导致对随后缺血性脑损伤的“耐受”。耐受性大脑的基因组特征是转录抑制。这种蛋白质合成依赖性耐受现象的翻译元件尚不清楚。然而，我们现在展示了耐受性蛋白质组（Stapels等人，Science Signaling，Mar 2010）。蛋白质组富含组蛋白，尤其是具有转录抑制功能的多梳组蛋白（Polycomb group，PcG）。因此，可能已经发现了一种以耐受为特征的转录抑制诱导机制。这一发现暗示表观遗传基因阻遏蛋白（PcGs）负责转录抑制过程，导致缺血耐受。此外，我们表明，PcGs，以前被称为调节器的分割在果蝇的发展过程中，有一个新的神经保护功能的大脑。我们在体内和体外对缺血耐受性的初步研究表明，缺血耐受性的发展依赖于PcG蛋白的表达：敲低消除耐受，过表达产生耐受。此外，我们现在有初步的证据，在缺血耐受性的三胸组（TrxG）蛋白，基因激活剂和拮抗剂的PcG蛋白的相反的变化。因此，我们将研究PcG/TrxG表观遗传调控，以确定其效应器的耐受机制。我们提供以下目标：具体目标1。以绝对摩尔数建立在缺血性损伤和缺血耐受开始时包含PcG/TrxG表观遗传调节剂复合物的蛋白质的化学计量。将对用针对所选PcG或TrxG蛋白的特异性抗体免疫沉淀的脑蛋白进行定量MS分析。具体目标2。确定缺血性损伤和缺血性耐受发生时脑中新合成蛋白的特性。我们将采用点击技术来代谢标记新合成的蛋白质，分离它们，并用定量MS分析对其进行定量。新生蛋白质组的生物信息学将在生物信息学工具的帮助下建立。 公共卫生相关性：我们展示了在大脑中发现的新蛋白质，这些蛋白质可以耐受缺血（中风）。我们假设它们是表观遗传调节蛋白（即大脑基因没有改变，但它们的反应发生了变化）。我们将进行分子生物学分析来验证这一假设。