The Role of Myc in ChREBP-dependent, Glucose-stimulated Gene Expression

Myc 在 ChREBP 依赖性、葡萄糖刺激的基因表达中的作用

• 批准号: 8422995
• 负责人: DONALD K. SCOTT
• 金额: $ 27.61万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-10-01 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8422995
• 关键词: Binding; Binding Proteins; Carbohydrates; Cell Nucleus; Cells; ChIP-seq; Complex; Data; Diabetes Mellitus; Disease; Environment; Epigenetic Process; Event; Failure; Fasting; Gene Expression; Gene Expression Profile; Gene Targeting; Genes; Genetic Transcription; Glucose; Glycolysis; Hepatic; Insulin Resistance; Lipids; Liver; Malignant Neoplasms; Measures; Mediating; Metabolic; Metabolic Diseases; Metabolic syndrome; Metabolism; Molecular; Molecular Conformation; Nucleic Acid Regulatory Sequences; Perception; Phenotype; Physiological; Play; Polymerase; Post-Translational Protein Processing; Process; Proteomics; Publishing; Pyruvate Kinase; Recruitment Activity; Response Elements; Reverse Transcriptase Polymerase Chain Reaction; Role; Structure of beta Cell of islet; Testing; Tissues; Transactivation; Transcriptional Activation; Warburg Effect; Work; activating transcription factor; blood glucose regulation; c-myc Genes; cell growth regulation; chromatin immunoprecipitation; detection of nutrient; domain mapping; feeding; glucose output; glucose tolerance; in vivo; insight; ketogenesis; lipid biosynthesis; oxidation; prevent; promoter; response; therapy design; transcription factor

DESCRIPTION (provided by applicant): The ability of cells to sense and respond to changes in their metabolic environment is crucial to preventing the pathophysiological consequences of metabolic syndrome and diabetes. Cellular adaptation to changes in metabolic fuel availability includes coordinated changes in patterns of gene expression, mediated by nutrient- sensing transcription factors, and this process is disrupted in diabetes, with cells trapped in the fasted state despite abundant metabolic fuel. The purpose of the present proposal is to understand the relationship between two transcription factors involved in nutrient sensing and the hepatic fasted-to-fed transition, the glucose-sensing transcription factor carbohydrate response element binding protein (ChREBP), and Myc. ChREBP mediates transcriptional activation of glucose-responsive target genes in metabolically relevant tissues such as the liver and pancreatic beta cells. Glucose stimulates translocation of ChREBP to the nucleus, and once ChREBP is bound to target genes, additional poorly defined molecular events, dependent on glucose, must take place for transactivation. Dysregulated Myc, and recently ChREBP, have both been implicated in the increased glycolysis (Warburg effect) associated with transformation in cancer. Our published and Preliminary Data demonstrate that Myc is required for ChREBP-dependent glucose-stimulated gene expression, is recruited to regulatory regions of glucose-responsive genes, and is necessary for the recruitment of ChREBP and other activating transcription factors of the glucose response. We hypothesize that Myc works in a larger context to allow transition between metabolic phenotypes, e.g., the fasted and fed states. Our challenge, reflected in this proposal is to understand more completely the molecular relationship between Myc and ChREBP within the context of the physiological roles they play in glucose homeostasis, so that we can better understand how to manipulate them therapeutically in disease states. Our overarching hypothesis is that Myc is required for the ChREBP-dependent glucose response, and for transitions between metabolic phenotypes. Specific Aim 1) is to determine the physiological, metabolic, and molecular consequences of depleting Myc activity in the liver. Using a CRE-Lox approach, we will determine the effect of hepatic Myc depletion on the fasting-to-fed transition by measuring gene expression with RT-PCR and SOLID SAGE; by measuring ChREBP and other transcription factor occupancies by ChIP and by ChIP-seq; and by measuring glycolysis, lipogenesis, lipid storage and oxidation, ketogenesis, and glucose output, as well as whole body and hepatic insulin resistance and glucose tolerance. Specific Aim 2 is to determine how Myc promotes ChREBP-dependent glucose-activated gene transcription. We will use a proteomic approach to test if glucose alters the posttranslational modification of Myc; we will determine if Myc alters the epigenetic environment of glucose responsive genes in a glucose-dependent manner; and will map the domains of Myc required for the glucose-specific functions of Myc. Accomplishing these Aims will provide the framework for understanding the normal physiological relationship between Myc and ChREBP, and will allow informed design of therapies for metabolic disorders such as metabolic syndrome and diabetes, and may well provide insights towards the metabolic perturbations of cancer.

描述（由申请人提供）：细胞感知和响应代谢环境变化的能力对于预防代谢综合征和糖尿病的病理生理后果至关重要。细胞对代谢燃料可用性变化的适应包括由营养敏感转录因子介导的基因表达模式的协调变化，而这一过程在糖尿病中被破坏，尽管代谢燃料充足，但细胞仍处于禁食状态。本提案的目的是了解两个参与营养感知和肝脏快速向摄食转变的转录因子，葡萄糖感知转录因子碳水化合物反应元件结合蛋白（ChREBP）和Myc之间的关系。ChREBP介导代谢相关组织（如肝脏和胰腺β细胞）中葡萄糖应答靶基因的转录激活。葡萄糖刺激ChREBP向细胞核的易位，一旦ChREBP与靶基因结合，必须发生依赖于葡萄糖的其他不明确的分子事件才能进行易动。失调的Myc和最近的ChREBP，都与癌症转化相关的糖酵解增加（Warburg效应）有关。我们发表的和初步的数据表明，Myc是ChREBP依赖性葡萄糖刺激基因表达所必需的，它被募集到葡萄糖应答基因的调控区域，并且是募集ChREBP和其他葡萄糖应答的激活转录因子所必需的。我们假设Myc在更大的背景下起作用，允许代谢表型之间的转换，例如，禁食和进食状态。我们的挑战，反映在本提案中，是更全面地了解Myc和ChREBP在葡萄糖稳态中所起的生理作用背景下的分子关系，以便我们能够更好地了解如何在疾病状态下进行治疗。我们的首要假设是，Myc是chrebp依赖性葡萄糖反应和代谢表型之间转换所必需的。具体目的1)是确定肝脏中Myc活性消耗的生理、代谢和分子后果。使用CRE-Lox方法，我们将通过RT-PCR和SOLID SAGE测量基因表达来确定肝脏Myc消耗对禁食到进食过渡的影响；通过ChIP和ChIP-seq检测ChREBP和其他转录因子的占用率；通过测量糖酵解，脂肪生成，脂质储存和氧化，生酮，葡萄糖输出，以及全身和肝脏胰岛素抵抗和葡萄糖耐量。特异性目的2是确定Myc如何促进chrebp依赖性葡萄糖激活基因转录。我们将使用蛋白质组学方法来测试葡萄糖是否会改变Myc的翻译后修饰；我们将确定Myc是否以葡萄糖依赖的方式改变葡萄糖反应基因的表观遗传环境；并绘制Myc的葡萄糖特异性功能所需的Myc结构域。实现这些目标将为理解Myc和ChREBP之间的正常生理关系提供框架，并将允许对代谢综合征和糖尿病等代谢紊乱的治疗进行知情设计，并可能为癌症的代谢扰动提供见解。