Nutrient Flux and Development

养分流动与发育

• 批准号: 8741418
• 负责人: Michael Krause
• 金额: $ 47.83万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8741418
• 关键词: Acetylglucosamine; Affect; Aging; Alleles; Animal Model; Animals; Antibodies; Biological; C. elegans genome; Caenorhabditis elegans; Cells; Chromatin; Collaborations; Complex; Development; Diabetes Mellitus; Disease; Enzymes; Excision; Fertility; Gene Expression Profile; Gene Expression Regulation; Genes; Genetic Models; Genomics; Glucose; Heme; Hexosamines; Homeostasis; Human; Insulin; Knock-out; Ligands; Link; Longevity; Longitudinal Studies; Maintenance; Mammals; Maryland; Mediating; Metabolic Diseases; Metabolism; National Institute of Diabetes and Digestive and Kidney Diseases; Nematoda; Non-Insulin-Dependent Diabetes Mellitus; Nuclear Hormone Receptors; Nuclear Pore; Nutrient; Nutritional; Organism; Pathway interactions; Phosphotransferases; Physiology; Pore Proteins; Process; Proteins; RNA; RNA Interference; Regulation; Role; Signal Pathway; Signal Transduction; Single Nucleotide Polymorphism; Stress; System; Testing; Time; Toxic effect; Universities; blood glucose regulation; chromatin immunoprecipitation; detection of nutrient; fly; heme a; insight; insulin signaling; interest; man; mutant; novel; peptide O-linked N-acetylglucosamine-beta-N-acetylglucosaminidase; pressure; prevent; promoter; reproductive; response; sensor; transcription factor

A dynamic cycle of O-linked N-acetylglucosamine (O-GlcNAc) addition and removal acts on nuclear pore proteins, transcription factors, and kinases to modulate cellular signaling cascades. This nutrient sensing hexosamine signaling pathway is conserved from nematodes to man. A single nucleotide polymorphism in the human O-GlcNAcase gene is linked to type 2 diabetes, suggesting that perturbation of this pathway results in disease. In collaboration the Hanover lab (NIDDK), we showed that the C. elegans genome encodes the two evolutionarily conserved enzymes that mediate O-GlcNAc cycling, with the genes called ogt-1 and oga-1. We previously characterized knockout alleles of ogt-1 and oga-1 genes. Using a combination of genomic expression arrays and chromatin immunoprecipitation (ChIP) we are looking for genes that respond to nutrient flux differently in the mutants with the hope of identifying pathways of importance. The expression analysis has revealed widespread de-regulation of gene expression in the mutants, identify affected pathways including longevity and aging. We have tested these pathways in the mutants and find alteration in function that are consistent with the gene expression patterns we observe. From the ChIP studies, we have identified a discrete number of genes associated with O-GlcNAcylated proteins. These associations are pronounced at the promoters of the genes and show some overlap with ChIP signals using RNA PolII antibodies. We are currently investigating the functional roles, if any, of these restricted O-GlcNAc chromatin marks. These marks have the potential to link nutritional flux in the cell directly to gene regulation, offering a novel insight into the role of O-GlcNAc cycling in animal physiology and development. In a variety of organisms, including worms, flies, and mammals, glucose homeostasis is maintained by insulin-like signaling in a robust network of opposing and complementary signaling pathways. The hexosamine signaling pathway, terminating in O-linked-N-acetylglucosamine (O-GlcNAc) cycling, is a key sensor of nutrient status and has been genetically linked to the regulation of insulin signaling in Caenorhabditis elegans. During the past year, we have demonstrated that O-GlcNAc cycling and insulin signaling are both essential components of the C. elegans response to glucose stress. A number of insulin-dependent processes were found to be sensitive to glucose stress, including fertility, reproductive timing, and dauer formation, yet each of these differed in their threshold of sensitivity to glucose excess. Our findings suggest that O-GlcNAc cycling and insulin signaling are both required for a robust and adaptable response to glucose stress, but these two pathways show complex and interdependent roles in the maintenance of glucoseinsulin homeostasis. In collaboration with the Hamza Lab (University of Maryland), we continue to use the C. elegans system to explore genes required for proper heme sensing and homeostasis. Further analysis of 288 heme responsive genes (hrgs) by RNAi mediated knockdown in a heme sensing strain has revealed additional genes required for proper heme homeostasis. Our study provides insights into metazoan regulation of organismal heme homeostasis. In collaboration with the Kostrouch and Kostrouchova Labs (Charles University, Prague) we have continued are long-term studies of the many nuclear hormone receptors (nhrs) in C. elegans. The current year project focused on a nhr that was found to have an interesting role in development. This information adds to our general understanding of nhr function and provides insights into the biological pressures in nematodes that have led to a huge expansion of this class of ligand-regulated transcription factors.

O-连接N-乙酰氨基葡萄糖(O-GlcNAc)的添加和移除作用于核孔蛋白、转录因子和激酶的动态循环，调节细胞信号级联反应。这种感知营养的氨基己糖信号通路从线虫到人类都是保守的。人类O-GlcNAcase基因的单核苷酸多态与2型糖尿病有关，这表明这一途径的扰动导致了疾病。 在汉诺威实验室(NIDDK)的合作下，我们发现线虫基因组编码了两种进化上保守的酶，它们介导了O-GlcNAc循环，基因被称为OGT-1和OGA-1。我们以前描述了OGT-1和OGA-1基因的敲除等位基因。利用基因组表达阵列和染色质免疫沉淀(CHIP)的组合，我们正在寻找在突变体中对营养通量做出不同反应的基因，希望确定重要的途径。表达分析表明，突变体中的基因表达普遍下调，确定了影响基因表达的途径，包括长寿和衰老。我们已经在突变体中测试了这些途径，发现功能上的变化与我们观察到的基因表达模式一致。从芯片研究中，我们已经确定了一些与O-GlcNacylated蛋白质相关的基因。这些关联在基因的启动子上明显存在，并与使用RNA PolII抗体的芯片信号显示出一些重叠。我们目前正在研究这些限制性O-GlcNAc染色质标记的功能作用。这些标记有可能将细胞内的营养流量直接与基因调控联系起来，为O-GlcNAc循环在动物生理和发育中的作用提供了新的见解。 在包括蠕虫、苍蝇和哺乳动物在内的各种生物体中，葡萄糖的稳态是由 胰岛素样信号在一个由相反和互补的信号通路组成的强大网络中。这个 氨基己糖信号通路终止于O-连接N-乙酰氨基葡萄糖(O-GlcNAc)循环，是营养状态的关键感受器，在基因上与秀丽线虫胰岛素信号的调节有关。在过去的一年里，我们已经证明O-GlcNAc循环和胰岛素信号都是线虫对葡萄糖胁迫反应的重要组成部分。许多胰岛素依赖的过程被发现对葡萄糖压力敏感，包括生育、生殖时机和Dauer形成，但这些过程中的每一个对葡萄糖过量的敏感性阈值都不同。我们的发现表明，O-GlcNAc循环和胰岛素信号都是对葡萄糖应激做出强健和适应性反应所必需的，但这两条途径在维持葡萄糖胰岛素稳态方面显示出复杂和相互依赖的作用。 在与哈姆扎实验室(马里兰大学)的合作下，我们继续使用线虫系统来探索适当的血红素感应和动态平衡所需的基因。在一个血红素敏感菌株中，通过RNAi介导的敲除，进一步分析了288个血红素反应基因(Hrgs)，发现了正确的血红素动态平衡所需的额外基因。我们的研究为后生动物对生物体内血红素动态平衡的调节提供了见解。 在与Kostrouch和Kostrouchova实验室(查尔斯大学，布拉格)的合作下，我们继续对线虫中的许多核激素受体(Nhr)进行长期研究。今年的项目重点是一项被发现在发展中具有有趣作用的国家卫生条例。这些信息增加了我们对NHR功能的总体理解，并提供了对线虫中的生物压力的见解，这些压力导致了这类配体调节的转录因子的巨大扩张。