Cellular and Molecular Mechanisms of SH2B1 Mutations That Cause Profound Childhood Obesity

摘要 肥胖是一个主要的健康问题，与代谢综合征、糖尿病和 心脏病。因此，肥胖需要从细胞、分子和社会三个层面深入了解其原因。 生物体水平。在一个令人兴奋的新发展中，鉴定出与SH2B1相关的人类突变 与严重的儿童肥胖症有关的支架蛋白SH2B1是体重的关键调节因子，在- 苏林的敏感性和行为。我们对SH2B1如何调节神经的理解存在一个根本性的空白 维持能量平衡的回路，以及人类肥胖SH2B1突变是如何破坏这一回路的 并导致肥胖。我们的长期目标是识别新的关键信号蛋白和/或基因调控 受SH2B1调控的事件，对于建立和维持对NOR- 雄性摄食行为和能量平衡，可靶向治疗肥胖症，胰岛素- 抵抗力和/或不适应行为。将使用原代神经元、新的小鼠模型和培养细胞 为了检验SH2B1对神经回路的建立和维持至关重要的中心假设 对正常的进食行为和能量平衡很重要。从力学上讲，SH2B1是一种支架。 加强质膜上的信号通路和向细胞核循环的Tein都是必需的 用于调节基因转录和轴突生长。人类疾病的突变会损害这些基因的一个子集 海绵。其具体目的是：1)确定SH2B1和SH2B1调控的神经营养配体信号通路； 2)确定神经突起生长所需的核SH2B1是如何生长的， 增强基因表达，而人类突变削弱这种增强；以及3)定义 SH2B1在下丘脑神经元的神经回路形成和转录中参与能量平衡。这 研究具有创新性，因为：1)SH2B1最近被认为是一种人类肥胖基因；2)新发现的SH2B1是一种肥胖基因。 SH2B1基因突变为研究肥胖的原因提供了强有力的工具；3)协调和 支架蛋白在质膜间移动对神经营养配体的综合反应 4)提出的许多技术和小鼠模型都是尖端的，包括- 用CRISPR/Cas9技术删除或编辑Sh2b1，小鼠模型研究SH2B1对神经元的影响 投射，研究下丘脑LepRb神经元小群体内的基因表达，以及 独一无二的Sh2b1P322S小鼠，能够研究人类SH2B1突变对完整小鼠和 孤立的神经元。这项拟议的研究意义重大，因为它将提供对细胞 SH2B1和人类突变影响神经元功能的分子机制包括 这些调节体重，以及复杂的，多蛋白质为基础的信号是如何协调之间的Plas- MA膜受体和细胞核。这一见解将促进我们对神经元功能和 确定肥胖、胰岛素抵抗和/或适应不良行为的潜在治疗目标。

ABSTRACT Obesity is a major health problem associated with a significant increase in metabolic syndrome, diabetes, and heart disease. Thus, obesity demands an in-depth understanding of its causes at the cellular, molecular and organismal level. In an exciting new development, identification of human mutations in SH2B1 that associate with profound childhood obesity implicate the scaffold protein SH2B1 as a critical regulator of body weight, in- sulin sensitivity and behavior. There is a fundamental gap in our understanding of how SH2B1 regulates neural circuitry that maintains energy homeostasis and how human obesity mutations in SH2B1 disrupt that circuitry and contribute to obesity. Our long-term goal is to identify novel key signaling proteins and/or gene regulatory events that are regulated by SH2B1, critical for establishing and maintaining neural circuits important for nor- mal feeding behavior and energy balance, can be targeted for therapeutic intervention for obesity, insulin- resistance and/or maladaptive behavior. Primary neurons, novel mouse models and cultured cells will be used to test the central hypothesis that SH2B1 is crucial for the establishment and maintenance of neural circuits important for normal feeding behavior and energy balance. Mechanistically, SH2B1 serves as a scaffold pro- tein that enhances signaling pathways at the plasma membrane and cycles to the nucleus, both are required for regulating gene transcription and neurite outgrowth. Human disease mutations impair a subset of these re- sponses. The specific aims are: 1) Determine neurotrophic ligand signaling pathways regulated by SH2B1 and impaired by the human mutations; 2) Determine how nuclear SH2B1, which is required for neurite outgrowth, enhances gene expression and the human mutations impair that enhancement; and 3) Define the role for SH2B1 in neural circuit formation and transcription in hypothalamic neurons implicated in energy balance. This research is innovative because: 1) SH2B1 was recently implicated as a human obesity gene; 2) newly identi- fied SH2B1 mutations provide powerful tools to study the causes of obesity; 3) the concept of coordinating an integrated response to neurotrophic ligands by movement of scaffold proteins between the plasma membrane and the nucleus is novel; and 4) many of the proposed techniques and mouse models are cutting edge, includ- ing CRISPR/Cas9 technology to delete or edit Sh2b1, mouse models to study the effect of SH2B1 on neuronal projections, studying gene expression within the small population of LepRb neurons in the hypothalamus, and the unique Sh2b1P322S mouse that enables study of the impact of a human SH2B1 mutation in intact mice and isolated neurons. The proposed research is significant because it will provide critical insight into the cellular and molecular mechanisms by which SH2B1 and the human mutations affect the function of neurons, including those that regulate body weight, and how complex, multi-protein based signals are coordinated between plas- ma membrane receptors and the nucleus. This insight will advance our understanding of neuron function and identify potential new therapeutic targets for obesity, insulin resistance and/or maladaptive behavior.