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: 8577575
• 负责人: DONALD K. SCOTT
• 金额: $ 24.62万
• 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/8577575
• 关键词: Role; Myc; ChREBP; dependent; Glucose

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)：细胞对代谢环境变化的感知和反应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。细胞对代谢燃料供应的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，这一过程在糖尿病患者中被破坏，尽管有丰富的代谢燃料，但细胞仍处于禁食状态。本研究的目的是了解参与营养感知的两个转录因子--葡萄糖敏感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在代谢相关的组织，如肝脏和胰腺的β细胞中，介导葡萄糖反应靶基因的转录激活。葡萄糖刺激ChREBP的转位到细胞核，一旦ChREBP与靶基因结合，依赖于葡萄糖的其他不明确的分子事件必须发生才能反式激活。调节失调的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。我们已发表的和初步的数据表明，Myc是依赖ChREBP的葡萄糖刺激的基因表达所必需的，被招募到葡萄糖反应基因的调节区，并对ChREBP和其他激活葡萄糖反应的转录因子的招募是必需的。我们假设Myc在更大的背景下工作，以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，反映在这一提议中，是更全面地了解Myc和ChREBP之间的分子关系，从它们在血糖稳态中所起的生理作用出发，以便我们能够更好地理解如何在疾病状态下对它们进行治疗。我们的主要假设是，Myc是依赖ChREBP的葡萄糖反应所必需的，也是代谢表型转换所必需的。具体目的1)是确定肝脏中Myc活性枯竭的生理、代谢和分子后果。使用CRE-Lox方法，我们将通过RT-PCR和Solid SAGE检测基因表达，通过CHIP和CHIP-SEQ检测ChREBP和其他转录因子的占有率，通过测量糖酵解、脂肪生成、脂肪储存和氧化、酮合成和葡萄糖输出，以及全身和肝脏的胰岛素抵抗和葡萄糖耐量，来确定肝脏Myc耗竭对空腹到喂养过渡的影响。具体目的2是确定Myc如何促进ChREBP依赖的葡萄糖激活基因的转录。我们将使用蛋白质组学方法来测试葡萄糖是否改变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；并将绘制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方案进行知情设计，并很可能为癌症的代谢扰动提供见解。