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感应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，但细胞仍陷入禁食状态。本提案的目的是了解参与营养感应和肝脏禁食到进食转变的两个转录因子（葡萄糖感应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 介导肝脏和胰腺 β 细胞等代谢相关组织中葡萄糖响应靶基因的转录激活。葡萄糖刺激 ChREBP 易位到细胞核，一旦 ChREBP 与靶基因结合，就必须发生依赖于葡萄糖的额外定义不明确的分子事件以进行反式激活。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。我们发表的初步数据表明，Myc 是 ChREBP 依赖性葡萄糖刺激基因表达所必需的，被招募到葡萄糖反应基因的调节区域，并且是招募 ChREBP 和其他葡萄糖反应激活转录因子所必需的。我们假设 Myc 在更大的背景下发挥作用，以允许代谢表型之间的转换，例如禁食和进食状态。该提案反映的我们面临的挑战是，在 Myc 和 ChREBP 在葡萄糖稳态中发挥的生理作用的背景下，更全面地了解它们之间的分子关系，以便我们能够更好地了解如何在疾病状态下对它们进行治疗。我们的总体假设是 Myc 是 ChREBP 依赖性葡萄糖反应以及代谢表型之间转变所必需的。具体目标 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 测量 ChREBP 和其他转录因子的占据；并通过测量糖酵解、脂肪生成、脂质储存和氧化、生酮和葡萄糖输出，以及全身和肝脏胰岛素抵抗和葡萄糖耐量。具体目标 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 是否以葡萄糖依赖性方式改变葡萄糖反应基因的表观遗传环境；并将绘制 Myc 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疗法进行明智的设计，并且很可能为癌症的代谢扰动提供见解。