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Opioid Peptide Synthesizing Enzymes

阿片肽合成酶

基本信息

批准号：
10163827
负责人：
IRIS LINDBERG
金额：
$ 34.5万
依托单位：
UNIVERSITY OF MARYLAND BALTIMORE
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-07-15 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10163827
关键词：
Address Affect Binding Binding Proteins Biochemical Biological Body Weight C-terminal Cell Culture Techniques Cell model Cells Cellular biology Child Clustered Regularly Interspaced Short Palindromic Repeats Collaborations Corticotropin Defect Desire for food Development Diabetes Mellitus Disease Distress Dominant-Negative Mutation Drug Addiction Drug abuse Eating Endocrine Energy Metabolism Engineering Environment Enzymes Exhibits Genes Genetic Polymorphism Genotype Half-Life Heterozygote High Fat Diet Homeostasis Homozygote Human Human Biology Hypothalamic structure Impairment Interruption Investigation Knockout Mice Laboratories Learning LoxP-flanked allele Mediating Modeling Molecular Chaperones Molecular Conformation Mus Mutant Strains Mice Mutation Neurons Neuropeptides Obesity Opioid Opioid Peptide Pathway interactions Peptide Hydrolases Peptide Signal Sequences Peptides Phenotype Physiological Play Predisposition Pro-Opiomelanocortin Process Production Prohormone Convertase Property Proprotein Convertase 1 Proprotein Convertase 2 Protein Precursors Proteins Rewards Risk Factors Role Satiation Signal Transduction Stress Testing Tissues Variant Wild Type Mouse Work alpha-Melanocyte stimulating hormone anorexic base beta-Endorphin cost endoplasmic reticulum stress energy balance enzyme pathway feeding glucose metabolism hormonal signals in vivo inhibitor/antagonist insight lipid metabolism mouse model mutant neuronal circuitry novel obesity risk pain signal pediatric patients peptide A relating to nervous system stressor trafficking

项目摘要

The prohormone convertases PC1/3 and PC2, encoded by the genes PCSK1 and PCSK2 respectively, are the endoproteolytic enzymes responsible for the liberation of opioid-active peptides from larger precursor proteins. Prohormone convertases play important roles not only in opioid peptide-mediated pain signaling but also function in many other neuronal circuits, including in reward pathways and in hypothalamic circuits involved in feeding and energy homeostasis. For example, both rare and common variations in PCSK1 function as major risk factors for human obesity, potentially due to deficiencies in hypothalamic peptidergic processing. In collaboration with clinicians who have identified children with novel mutations in PCSK1, we have recently determined that mutant human and mouse PC1/3 proteins are subject to targeting defects which are likely to result in hypothalamic proteostatic stress. Based on our prior finding that mouse PC1/3 proteins oligomerize during synthesis, we propose that dominant-negative interactions play a major role in human PC1/3 heterozygote obesity phenotypes, affecting precursor processing to bioactive peptides involved in satiety signaling. We propose that external stressors will exacerbate even mild forms of PC1/3 conformational distress, impairing C-terminal cleavage of PC1/3 to the smaller, more active forms. These processes will ultimately converge to strongly impair precursor processing, eg. proopiomelanocortin cleavage to beta-endorphin, ACTH, and most importantly, to the anorexic peptide α- MSH. In the present proposal we will use CRISPR-engineered cell models to elucidate the cell biology and precursor processing efficacy of three human PC1/3 variants and mutants known to be strongly associated with increased risk of obesity. Secondly, we will create mouse models of two common human PCSK1 obesity mutants, and a third model of a rare but highly impaired mutant, to extend findings made in cell culture to actual secretory tissues, and to identify the specific physiologic alterations which underlie the PCSK1-mediated obesity phenotype. Lastly, we will test our hypothesis that processing deficits in proopiomelanocortin-synthesizing neurons underlie the obesity phenotype by selectively eliminating Pcsk1 expression in proopiomelanocortin-expressing cells using a floxed Pcsk1 null mouse model. The results of these studies will illuminate the biosynthetic mechanisms controlling hypothalamic peptide production that contribute to human susceptibility to a variety of diseases, from obesity to reward pathways in drug addiction.

激素原转化酶 PC1/3 和 PC2 分别由基因 PCSK1 和 PCSK2 编码，是负责从较大前体中释放阿片活性肽的内切蛋白水解酶蛋白质。激素原转化酶不仅在阿片肽介导的疼痛信号传导中发挥重要作用而且还在许多其他神经元回路中发挥作用，包括奖赏通路和下丘脑参与喂养和能量稳态的电路。例如，罕见和常见的变异 PCSK1 是人类肥胖的主要危险因素，可能是由于下丘脑缺陷所致肽能加工。与已发现具有新突变的儿童的临床医生合作在 PCSK1 中，我们最近确定突变的人类和小鼠 PC1/3 蛋白会受到针对可能导致下丘脑蛋白静态应激的缺陷。根据我们之前的发现由于小鼠 PC1/3 蛋白在合成过程中寡聚，我们提出显性负相互作用在人类 PC1/3 杂合子肥胖表型中发挥重要作用，影响前体加工参与饱腹感信号传导的生物活性肽。我们认为外部压力甚至会加剧轻度形式的 PC1/3 构象窘迫，损害 PC1/3 的 C 端裂解，使其变得更小、更多活跃的形式。这些过程最终将集中到严重损害前体加工，例如。阿黑皮质素原裂解为 β-内啡肽、ACTH，最重要的是，裂解为厌食肽 α- MSH。在本提案中，我们将使用 CRISPR 工程细胞模型来阐明细胞生物学和已知密切相关的三种人类 PC1/3 变体和突变体的前体加工功效肥胖风险增加。其次，我们将创建两种常见人类PCSK1的小鼠模型肥胖突变体，以及罕见但高度受损的突变体的第三种模型，以扩展细胞中的发现培养实际的分泌组织，并识别潜在的特定生理变化 PCSK1 介导的肥胖表型。最后，我们将检验我们的假设，即处理缺陷阿黑皮质素原合成神经元通过选择性消除 Pcsk1 成为肥胖表型的基础使用 floxed Pcsk1 null 小鼠模型在阿片黑皮素原表达细胞中进行表达。结果这些研究将阐明控制下丘脑肽产生的生物合成机制导致人类对多种疾病的易感性，从肥胖到药物奖励途径瘾。