Biochemical Analysis of a p53 Isoform that Accelerates Mammalian Aging

加速哺乳动物衰老的 p53 异构体的生化分析

• 批准号: 8016662
• 负责人: Dylan J Taatjes
• 金额: $ 5.97万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-01 至 2012-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8016662
• 关键词: Affect; Age; Aging; Animals; Biochemical; Cells; Chimeric Proteins; DNA Binding; Disease; Future; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic Transcription; HCT116 Cells; Heart; Human; Insecta; Insulin-Like Growth Factor I; Knock-in Mouse; Lead; Length; Link; Longevity; MDM2 gene; Malignant Neoplasms; Mammals; Messenger RNA; Modeling; Molecular; Mus; Phenotype; Physiological Processes; Play; Premature aging syndrome; Process; Protein Isoforms; Protein p53; Proteins; Refractory; Regulation; Relative (related person); Repression; Research; Role; Series; Staging; System; Testing; Time; Transcription Repressor/Corepressor; Transcriptional Activation; Transcriptional Regulation; Tumor Suppression; Tumor Suppressor Proteins; Work; age related; aged; base; cancer prevention; design; dimer; human disease; insight; mutant; overexpression; prevent; public health relevance; reconstitution; research study; response; transcription factor

DESCRIPTION (provided by applicant): The p53 tumor suppressor plays critical roles in both cancer-prevention and aging. Activation of p53 response is important for its tumor suppressor function; conversely, shutting off active p53 is critical to prevent premature aging. This dual role of p53 in cancer and aging is perhaps best demonstrated by the activity of a naturally-occurring isoform of p53 called Np53 (a.k.a. 40p53). When over-expressed in mice (together with wild-type p53), Np53 causes hyper-activation of p53 target genes; although this results in enhanced tumor suppression, the mice also age rapidly and die prematurely. The hyper-active p53 response caused by Np53 has been linked to transcription activation. Because p53 binds DNA and activates transcription as a tetramer, the formation of p53/ Np53 hetero-tetramers is believed to cause the enhanced gene expression and accelerated aging. However, the molecular mechanism by which this occurs is largely unexplored. In this proposal, we outline a series of experiments that will begin to define the molecular mechanisms by which Np53 functions within the context of the p53 tetramer. Central to this work is a detailed biochemical analysis of mixed p53/ Np53 tetramers in the context of a native p53 target gene using a reconstituted human transcription system. In addition, we will identify which p53 target genes are differentially impacted by p53/ Np53 in human cells by performing mRNA analysis of 28 known p53 target genes. We will focus specifically on aging-related p53 targets in this screen to determine which might be altered most significantly by the Np53 isoform. Taken together, these biochemical and cell-based studies will begin to define the molecular mechanisms that enable the Np53 isoform to accelerate mammalian aging. PUBLIC HEALTH RELEVANCE: Our research analyzes the basic mechanisms of gene expression-what turns a gene "on" or "off" in a cell. Understanding this process is vital because proper regulation of gene expression is essential for virtually every major physiological process; furthermore, breakdown in this regulation is a hallmark of human disease, most notably cancer. In this proposal we will examine the function of an improperly activated protein called p53. Understanding how p53 functions, particularly in its abnormal, "hyper-active" state, lies at the heart of the cancer and aging problem because the form of p53 we are studying shows enhanced ability to prevent cancer yet concomitantly shortens lifespan. The precise mechanisms by which this occurs are not at all understood and will be explored in this study. We anticipate that the information accumulated by our efforts will identify new strategies for controlling the transcriptional activity of p53.

描述（由申请人提供）：p53肿瘤抑制剂在预防癌症和衰老中都起着关键作用。 p53反应的激活对于其肿瘤抑制功能很重要。相反，关闭主动P53对于防止过早衰老至关重要。 p53在癌症和衰老中的这种双重作用也许最好通过p53的自然疾病同工型的活性来证明NP53（又称40p53）。当在小鼠中过表达（与野生型p53一起）时，NP53会导致p53靶基因过度激活。尽管这导致肿瘤抑制增加，但小鼠也迅速变老并过早死亡。由NP53引起的超活动p53响应与转录激活有关。由于p53结合DNA并激活转录作为四聚体，因此据信p53/ np53杂种四聚体的形成会导致增强的基因表达和加速衰老。但是，发生这种情况的分子机制在很大程度上没有探索。在此提案中，我们概述了一系列实验，这些实验将开始定义NP53在p53四聚体背景下起作用的分子机制。这项工作的核心是使用重构的人类转录系统在天然p53靶基因的背景下对混合p53/ NP53四聚体进行详细的生化分析。此外，我们将通过对28个已知p53靶基因进行mRNA分析来确定人类细胞中p53/ np53差异影响哪些p53靶基因。我们将专门关注与衰老相关的p53目标，以确定NP53同工型可能最大程度地改变了哪些目标。综上所述，这些生化和基于细胞的研究将开始定义能够加速哺乳动物衰老的NP53同工型的分子机制。 公共卫生相关性：我们的研究分析了基因表达的基本机制 - 细胞中的基因“ ON”或“ OFF”。理解这一过程至关重要，因为对基因表达的适当调节对于几乎每个主要的生理过程都是必不可少的。此外，该法规中的细分是人类疾病的标志，最著名的是癌症。在此提案中，我们将研究一种称为p53的不当激活蛋白质的功能。了解p53的功能，尤其是在其异常，“活跃”状态下的功能，是癌症和衰老问题的核心，因为我们研究的p53形式显示出增强的预防癌症能力，同时缩短了寿命。根本没有理解发生这种情况的确切机制，并将在本研究中探索。我们预计我们的努力所积累的信息将确定控制p53转录活动的新策略。