Co-repressors SMRT and NCoR1 regulate UGT1A1 gene expression

共阻遏物 SMRT 和 NCoR1 调节 UGT1A1 基因表达

基本信息

批准号：
8761224
负责人：
Robert H Tukey
金额：
$ 33.79万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-01 至 2018-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8761224
关键词：
Animal Model Benign Bilirubin Birth Blood ChIP-seq Chromatin Complex Development Environmental Risk Factor Epigenetic Process Event Gastrointestinal tract structure Gene Expression Gene Expression Profile Gene Expression Regulation Gene Targeting Genes Genetic Hepatic Hepatic Tissue Hepatocyte Human Hyperbilirubinemia In Vitro Interruption Intestines Knock-out Lead Life Link Liver Location Measures Metabolic Metabolism Modification Mus Mutation NCOR1 gene Neonatal Neuraxis Neurologic Dysfunctions Newborn Infant Nuclear Receptors Pharmaceutical Preparations Play Proteins Publishing Receptor Inhibition Recovery Regulation Repression Repressor Proteins Role Series Serum Small Interfering RNA Syndrome Testing Therapeutic Agents Tissues Toxic effect Transcriptional Regulation UDP-Glucuronosyltransferase 1A1 UGT1A1 gene Vision base design drug metabolism early onset gastrointestinal gene repression human NCOR1 protein in vivo Model model design neonatal death neonatal hyperbilirubinemia novel pregnane X receptor prevent public health relevance receptor binding research study transcriptome sequencing

项目摘要

DESCRIPTION (provided by applicant): The characterization of the regulatory events leading to developmental control of the human UGT1A1 gene and the early onset of neonatal hyperbilirubinemia is the central focus of this application. While generally benign, the occurrence of hyperbilirubinemia can lead to central nervous system toxicity, even when extreme measures to prevent its duration and accumulation have been implemented. The underlying mechanisms associated with this well-known syndrome are currently unknown. We will demonstrate in preliminary studies using novel animal models that the onset of neonatal hyperbilirubinemia is a highly coordinated event that involves tissue specific repression of the UGT1A1 gene. In humanized UGT1 (hUGT1) mice, neonatal mice develop severe levels of serum bilirubin, resulting from repression of the liver and intestinal UGT1A1 gene. In a series of experiments designed to examine the impact of PXR and CAR on regulation of the genes encoded by the UGT1 locus, we prepared hUGT1 mice that were deficient in PXR (hUGT1/Pxr-/-) and CAR (hUGT1/Car-/-). From these studies, it was discovered that the deletion of PXR resulted in de-repression of UGT1A1 gene expression in liver tissue, a result that led to accelerated metabolism of serum bilirubin as shown by reduction of bilirubin during neonatal development. Since the nuclear receptors (NRs) such as PXR are targets for co- repressors SMRT and NCoR1, we undertook experiments to examine the role of these repressor proteins on developmental expression of the UGT1A1 gene. We have discovered that in primary neonatal hUGT1 hepatocytes, knockout of SMRT by siRNA leads to induction of UGT1A1 gene expression, potentially linking SMRT repression with the actions of PXR in liver. These preliminary findings will be the basis for experiments performed in Specific Aim 1 which will examine the impact of liver specific knockout of SMRT on UGT1A1 gene expression in hUGT1 mice. NCoR1, on the other hand, is highly specific for repressing UGT1A1 gene expression in intestinal tissue during development. Targeted deletion of NCoR1 in intestinal tissue of hUGT1 mice (hUGT1/NCor1GI¿GI mice) leads remarkably to the complete reversal of neonatal hyperbilirubinemia, with dramatic induction of intestinal UGT1A1 gene expression. Experiments outlined in Specific Aim 2 will focus on the identification of the NR involved in NCoR1 repression of intestinal UGT1A1 gene expression. In Specific Aim 3, studies will focus on the epigenetic changes that occur in liver and intestine as a result of SMRT and NCoR1 deletion in these tissues, respectively. These changes will focus on the location of SMRT/NCoR/NR binding and the chromatin changes across the UGT1 locus in liver and intestine as characterized by ChIP-seq and RNA-seq analysis. We will be describing novel mechanisms leading to developmental control of the UGT1A1 gene and its implications toward neonatal hyperbilirubinemia.

描述（通过应用程序提供）：调节事件的表征导致对人类UGT1A1基因的控制以及新生儿高胆红素血症的早期发作是该应用的中心重点。尽管通常良性，但高胆红素血症的发生也会导致中枢神经系统毒性，即使采取了极端的措施，以防止其持续时间和积累。目前尚不清楚与该众所周知的综合征相关的基本机制。我们将在初步研究中使用新型动物模型证明，新生儿高胆红素血症的发作是一个高度协调的事件，涉及UGT1A1基因的组织特异性表达。在人源化的UGT1（HUGT1）小鼠中，新生儿小鼠会出现严重水平的血清胆红素，这是由于肝脏和肠道UGT1A1基因的表达而产生的。 PXR和CAR对UGT1基因座编码的基因调节的影响，我们准备了Hugt1小鼠，这些小鼠缺乏PXR（HUGT1/PXR - / - ）和CAR（HUGT1/CAR - / - ）。从这些研究中，发现PXR的缺失导致肝组织中UGT1A1基因表达的抑制，这导致血清胆红素的代谢加速，如新生儿发育过程中胆红素的还原所示。由于核接收器（NR）（例如PXR）是共抑制剂SMRT和NCOR1的靶标，因此我们进行了实验，以检查这些复制蛋白在UGT1A1基因发育表达中的作用。我们发现，在原发性新生儿HUGT1肝细胞中，siRNA敲除SMRT会导致诱导UGT1A1基因表达，可能将SMRT表达与肝脏中PXR的作用联系起来。这些初步发现将是在特定目标1中进行的实验的基础，该实验将研究SMRT的肝脏特异性敲除对HUGT1小鼠中UGT1A1基因表达的影响。另一方面，NCOR1高度特异性对于在发育过程中抑制肠道组织中的UGT1A1基因表达。在Hugt1小鼠（HUGT1/NCOR1GI小鼠）肠道组织中，NCOR1的靶向缺失显着导致新生儿高胆红素血症的完全逆转，并具有巨大的肠道UGTT1A1基因表达的巨大诱导。特定目标2中概述的实验将集中于识别参与NCOR1肠道ugt1a1基因表达的NR。在特定的目标3中，研究将重点放在这些时机中SMRT和NCOR1删除的结果上发生的表观遗传变化。这些变化将集中于SMRT/NCOR/NR结合的位置，以及以ChIP-Seq和RNA-Seq分析为特征的UGT1基因座的染色质变化。我们将描述导致对UGT1A1基因的发育控制及其对新生儿高胆红素血症的影响。