Chronic Alcohol Effects on Transcriptional Regulation in Liver Regeneration

慢性酒精对肝脏再生转录调节的影响

• 批准号: 7587997
• 负责人: Rajanikanth Vadigepalli
• 金额: $ 22.21万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2011-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7587997
• 关键词: Acute; Address; Adult; Alcohol consumption; Alcohol dependence; Alcoholic Liver Diseases; Alcohols; Automobile Driving; Binding Sites; Bioinformatics; Biological; Biological Assay; Chemicals; Chronic; Cluster Analysis; Complex; DNA Binding; Data; Development; Event; Funding; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Hour; Impairment; Liver; Liver Regeneration; Liver diseases; Log-Linear Models; Mechanics; Mediating; Methods; Modeling; Morbidity - disease rate; Natural regeneration; Nature; Paint; Partial Hepatectomy; Pathway Analysis; Pattern; Process; Rattus; Regulation; Series; Site; System; Testing; Time; Tissues; Transcriptional Regulation; Viral; alcohol abuse therapy; alcohol effect; alcohol exposure; base; chromatin immunoprecipitation; combinatorial; feeding; in vivo; mortality; novel; promoter; regenerative; repaired; response; therapeutic target; transcription factor

DESCRIPTION (provided by applicant): The primary objective of this exploratory application is to delineate the changes in the nature and time course of transcriptional regulatory mechanisms underlying the effects of chronic alcohol exposure on the onset of liver regeneration regeneration. Alcoholic liver disease continues to be a major cause of morbidity and mortality in the USA and worldwide. Evidence suggests that ethanol consumption makes the liver more vulnerable to other challenges, thereby predisposing the liver to damage through the impairment of the normal repair capacity of the liver. This disruption of transcriptional regulatory mechanisms by chronic alcohol exposure remains largely unexplained. The present application is aimed at addressing this problem taking advantage of a unique synergistic opportunity presented by two recent advances: (1) availability of high-throughput gene expression time series data being obtained from regenerating rat liver in normal and chronic alcohol exposure conditions, and (2) availability of PAINT, a toolkit of sensitive bioinformatics methods for transcriptional regulatory network analysis (TRNA) for deriving hypotheses on key underlying transcription factors (TFs). The overall driving biological hypothesis of the present project is that the disruptive effects of chronic alcohol on the liver regeneration involve alterations in a focused set of TFs acting in a network to regulate system-wide downstream target genes. We propose to investigate this hypothesis in an integrated computational and experimental approach in two aims, respectively: (1) Identify and validate the 'baseline' transcriptional regulation underlying the onset of normal liver regeneration, and (2) Identify and validate the effects of chronic alcohol exposure on the changes in transcriptional regulation underlying the onset of liver regeneration. In each of the aims, we will employ a mixed-effects ANOVA model to analyze the time series gene expression data in order to identify differentially expressed genes and a subsequent cluster analysis to derive temporal expression patterns. We will perform TRNA using PAINT to hypothesize the candidate TFs with altered activity under chronic alcohol exposure in the onset of liver regeneration. We will validate a prioritized subset of the regulatory network using ELISA-based TF DNA-binding activity assays and chromatin immunoprecipitation followed by quantitative PCR and to assess quantitative changes in the in vivo occupancy of the candidate TFs at promoters of specific physiologically significant genes in the liver regeneration. In order to derive hypotheses on alcohol-perturbed combinatorial regulatory interactions, we will follow a log-linear modeling approach to compare the baseline and chronic alcohol-altered regulatory networks. A key deliverable of this project is a quantitative description of the chronic alcohol-dependent alterations of the activity of key TFs at specific promoter sites of physiologically significant genes in the onset of liver regeneration. Such a systems level understanding of alcohol-perturbed transcriptional regulatory mechanisms in regenerating liver will greatly aid in the development of therapeutic targets for ameliorating alcohol effects on progression to liver disease.

描述（由申请人提供）：本探索性申请的主要目的是描述慢性酒精暴露对肝脏再生发生影响的转录调节机制的性质和时间过程的变化。酒精性肝病仍然是美国和世界范围内发病率和死亡率的主要原因。有证据表明，乙醇的消耗使肝脏更容易受到其他挑战，从而通过损害肝脏的正常修复能力使肝脏容易受到损害。慢性酒精暴露对转录调控机制的破坏在很大程度上仍无法解释。本申请旨在利用最近两项进展提供的独特协同机会来解决这一问题：(1)从正常和慢性酒精暴露条件下的再生大鼠肝脏中获得高通量基因表达时间序列数据的可用性；(2)可获得PAINT，一种用于转录调控网络分析（TRNA）的敏感生物信息学方法工具箱，用于推导关键潜在转录因子（TFs）的假设。本项目的总体驱动生物学假设是，慢性酒精对肝脏再生的破坏性影响涉及在网络中调节全系统下游靶基因的一组重点tf的改变。我们建议通过综合计算和实验方法来研究这一假设，分别有两个目标：(1)确定并验证正常肝脏再生开始的“基线”转录调节，以及(2)确定并验证慢性酒精暴露对肝脏再生开始的转录调节变化的影响。在每个目标中，我们将采用混合效应方差分析模型来分析时间序列基因表达数据，以识别差异表达基因，并随后进行聚类分析以获得时间表达模式。我们将使用PAINT进行TRNA，以假设在肝脏再生开始时慢性酒精暴露下活性改变的候选tf。我们将使用基于elisa的TF dna结合活性测定和染色质免疫沉淀以及定量PCR来验证调控网络的优先子集，并评估肝脏再生中特定生理重要基因启动子处候选TF在体内占用的定量变化。为了得出关于酒精干扰组合调节相互作用的假设，我们将采用对数线性建模方法来比较基线和慢性酒精改变的调节网络。该项目的一个关键成果是定量描述肝脏再生开始时，生理上重要基因的特定启动子位点上关键tf活性的慢性酒精依赖性改变。这种对再生肝脏中酒精干扰的转录调控机制的系统水平的理解将极大地有助于开发治疗靶点，以改善酒精对肝脏疾病进展的影响。