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p53-Regulation in Liver Regeneration

p53-肝脏再生的调节

基本信息

批准号：
8281681
负责人：
Michelle Ann Barton
金额：
$ 32.07万
依托单位：
UNIVERSITY OF TX MD ANDERSON CAN CTR
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-06-10 至 2013-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8281681
关键词：
Address Affinity Chromatography Aging Apoptosis Area Attenuated Binding Biochemical C-terminal Cell Cycle Cell Cycle Arrest Cell physiology Cells Chemicals Chromatin Complement Development Disease Epitopes Excision Family member Financial compensation Goals Growth Hepatocyte Investigation Knock-in Mouse Knowledge Liver Liver Regeneration Masks Mass Spectrum Analysis Mediating Methodology Modification Molecular Natural regeneration Normal Cell Organogenesis Partial Hepatectomy Pathway interactions Peptides Physiological Post-Translational Protein Processing Process Protein p53 Proteome Regulation Research Signal Transduction Pathway Tissues Tumor Suppressor Proteins Tumor-Derived Work chromatin immunoprecipitation genome-wide analysis innovation mouse model mutant protein complex response senescence stem tissue regeneration

项目摘要

DESCRIPTION (provided by applicant): Renewed organogenesis of the fully differentiated liver is stimulated in response to surgical removal (partial hepatectomy, PHx) or destruction of liver tissue by chemicals or disease. An induced network of signal transduction pathways triggers re-entry of hepatocytes (followed by non-parenchymal cells) into cell cycle, proliferation and compensatory growth, which terminates at a precisely regulated endpoint. We hypothesize that critical functions of the p53 tumor suppressor protein, in promoting cell cycle arrest and apoptosis, must be blocked or attenuated in response to partial hepatectomy. Our goal is to define the mechanisms of this regulatory process. The wealth of information, regarding regulation and functions of p53, stems primarily from studies of tumor-derived cells, cells under long term, continuous culture, and/or cells expressing dysfunctional, mutant or exogenous p53. Although evidence continues to mount that p53 acts in normal cells during development, aging and senescence, major gaps in our knowledge exist regarding mechanisms of endogenous p53-regulation. Tumor suppressor p53 is expressed at low levels, which are tightly controlled by multiple pathways. This offers a considerable challenge to biochemical purification and mechanistic analyses of endogenous p53, especially in normal cells. We have developed a new mouse model to address these gaps in our knowledge. The mouse model, created by knock-in methodology, expresses endogenous p53 fused in-frame with a C-terminal epitope-tag (TAP-p53). TAP or Tandem Affinity Purification is a proven means of purifying low-abundance, large protein complexes under relatively physiological conditions. TAP-p53 purification and subsequent peptide analyses by mass spectrometry will be used to define p53-protein interactions (the p53 "proteome") and to determine post-translational modifications of p53. Epitope-tagging of p53 further facilitates studies of p53-chromatin interactions by chromatin immunoprecipitation (ChIP) and genome-wide analysis of p53-targets (ChIP-chip). We will use these and other approaches to establish the ground state of p53 function and regulation in normal hepatic cells and to determine how these functions may be altered during liver regeneration. The proposed research will address how p53 functions in normal, differentiated cells, an area generally overlooked in investigations of tumor suppressor activities. This work will further molecular understanding of tissue regeneration with a long-term goal of understanding how the surveillance status of the p53-network may be temporally controlled to facilitate cellular renewal and tissue regeneration. PROJECT NARRATIVE: These studies are focused on understanding how a fully differentiated tissue circumvents regulation and surveillance by tumor suppressor p53 to regenerate itself in response to partial hepatectomy. We believe that p53, which normally stops growth and proliferation, is temporarily blocked from functioning during regeneration and then restored to normal capacities when regeneration is finished. How p53 functions in normal cells and during regeneration is essentially unknown.

描述(由申请人提供)：完全分化的肝脏的器官再生是在手术切除(部分肝切除，PHX)或因化学物质或疾病破坏肝组织时被刺激的。诱导的信号转导网络触发肝细胞(随后是非实质细胞)重新进入细胞周期、增殖和代偿性生长，最终终止于精确调控的终点。我们假设P53肿瘤抑制蛋白在促进细胞周期停滞和细胞凋亡方面的关键功能必须被阻断或减弱，以回应部分肝切除。我们的目标是定义这一监管过程的机制。关于p53调控和功能的丰富信息主要来自对肿瘤来源的细胞、长期持续培养的细胞和/或表达功能障碍的、突变的或外源性p53的细胞的研究。虽然越来越多的证据表明P53在正常细胞的发育、衰老和衰老过程中起作用，但我们对内源性P53调控机制的认识存在重大差距。抑癌基因P53的表达水平较低，受到多条途径的严格调控。这对内源性P53的生化纯化和机理分析提出了相当大的挑战，尤其是在正常细胞中。我们已经开发了一种新的鼠标模型来解决我们知识中的这些空白。通过敲入方法建立的小鼠模型表达内源性P53与C末端表位标签(TAP-P53)融合在一起。TAP或串联亲和纯化是一种在相对生理条件下纯化低丰度、大蛋白质复合体的成熟方法。TAP-P53的纯化和随后的质谱分析将用于确定P53与蛋白质的相互作用(P53“蛋白质组”)，并确定P53的翻译后修饰。P53的表位标记进一步促进了通过染色质免疫沉淀(CHIP)和全基因组分析P53靶标(CHIP-CHIP)来研究P53-染色质的相互作用。我们将使用这些方法和其他方法来建立正常肝细胞中P53功能和调控的基本状态，并确定这些功能在肝脏再生过程中可能发生的变化。这项拟议的研究将解决P53如何在正常、分化的细胞中发挥作用，这是研究肿瘤抑制活性时通常忽视的一个领域。这项工作将进一步加深对组织再生的分子理解，长期目标是了解如何在时间上控制P53网络的监视状态，以促进细胞更新和组织再生。项目简介：这些研究的重点是了解完全分化的组织如何绕过肿瘤抑制基因P53的调控和监视，从而在部分肝切除后自我再生。我们认为，通常阻止生长和增殖的P53在再生过程中被暂时阻止功能，然后在再生完成后恢复到正常能力。P53在正常细胞和再生过程中是如何发挥作用的基本上是未知的。

项目成果

期刊论文数量（6）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

p53 regulates a mitotic transcription program and determines ploidy in normal mouse liver.

p53 调节有丝分裂转录程序并确定正常小鼠肝脏中的倍性。

DOI：
10.1002/hep.26233
发表时间：
2013-05
期刊：
HEPATOLOGY
影响因子：
13.5
作者：
Kurinna, Svitlana;Stratton, Sabrina A.;Coban, Zeynep;Schumacher, Jill M.;Grompe, Markus;Duncan, Andrew W.;Barton, Michelle Craig
通讯作者：
Barton, Michelle Craig

Integrative genomics: liver regeneration and hepatocellular carcinoma.