Molecular Regulation of Ureagenesis

尿素生成的分子调控

基本信息

批准号：
7570626
负责人：
Ljubica Morizono Caldovic
金额：
$ 10.76万
依托单位：
CHILDREN'S RESEARCH INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-03-01 至 2011-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7570626
关键词：
Acute Ammonia Animals Biochemistry Bioinformatics Blood Brain Cessation of life Chronic Cleaved cell Cognitive Coma Complement Defect Diet Dietary Proteins Disease Drosophila genus Enzymes Genes Goals Hepatocyte Homologous Gene Intake Intervention Knowledge Liver Liver Failure Liver Mitochondria Mentors Messenger RNA Mitochondria Modeling Molecular Molecular Biology Mus N acetyl L glutamate Nitrogen Organism Patients Peptide Hydrolases Pharmaceutical Preparations Physiological Play Process Production Proteins Proteolytic Processing Proteomics RNA Regulation Research Personnel Research Project Grants Research Training Role System Testing Time Tissues Training Urea Variant Waste Products career development experience fatty acid metabolism hepatic ureagenesis nitrogen metabolism prevent protein profiling research study response statistics therapeutic target tool urea cycle

项目摘要

DESCRIPTION (provided by applicant): This is a revised career development proposal, aimed at understanding molecular regulation of the urea cycle. The urea cycle functions in the liver to convert neuro-toxic ammonia into non-toxic urea. One potential strategy to alleviate the effects of high blood ammonia is to increase patients' innate ability to produce urea by raising the expression of urea cycle enzymes. Such interventions require an understanding of the regulation of ureagenesis at the molecular level. The activities of all urea cycle enzymes coordinately change in response to varying nitrogen loads. However, it appears that different mechanisms regulate gene expression of individual urea cycle genes. The goal of this project is to understand the regulation of ureagenesis at the molecular level. The specific aims of this project are: 1) To determine the functional and structural changes in liver NAGS in response to changing dietary protein intake. My hypothesis is that the ratio of two NAGS variants in the liver mitochondria is determined by the organism's nitrogen load. Furthermore, changes of the ratio of two NAGS variants reflect a post- translational mechanism for regulation of hepatic ureagenesis in response to varying nitrogen load. This hypothesis will be tested by measuring the relative amounts of the two NAGS variants in the livers from mice exposed to different nitrogen load, and identification of mitochondrial protease(s) capable of processing NAGS. 2) To identify signaling pathways that respond to changes in nitrogen loads using transcriptional profiling. My hypothesis is that change in nitrogen loads lead to changes in expression levels of signaling molecules that result in either activation or repression of urea cycle genes. This hypothesis will be tested by measuring changes of mRNA levels in response to varying nitrogen load by transcriptional profiling of RNA using Affymetrix microarrays. 3) To identify proteins whose amounts and post- translational modifications change in response to changing nitrogen loads. My hypothesis is that changes in nitrogen loads triggers changes in amounts and/or post-translational modifications of proteins in one or more signaling pathways, resulting in change of expression and activity of urea cycle enzymes. This hypothesis will be tested by measuring changes in protein levels and their modifications in response to varying nitrogen loads using two- dimensional gel electrophoresis, densitometry and mass spectrometry.

描述（由申请人提供）：这是一项修订的职业发展建议，旨在了解尿素周期的分子调节。尿素周期在肝脏中起作用，将神经毒性氨转化为无毒尿素。减轻高血氨作用的一种潜在策略是通过提高尿素周期酶的表达来提高患者生产尿素的能力。这种干预措施需要了解分子水平的尿素发生的调节。所有尿素周期酶的活性随着氮负荷的不同而协调变化。然而，看来不同的机制调节了单个尿素周期基因的基因表达。该项目的目的是了解分子水平的尿素发生的调节。该项目的具体目的是：1）确定肝脏NAG的功能和结构变化，以响应不断变化的饮食蛋白质摄入。我的假设是，肝线粒体中两个NAG变体的比率取决于生物体的氮负荷。此外，两个NAG变体的比率变化反映了响应不同氮负荷的肝尿素发生后的转化后机制。该假设将通过测量暴露于不同氮负荷的小鼠的肝脏中两个NAG变体的相对量，并鉴定能够处理NAGS的线粒体蛋白酶的鉴定。 2）确定使用转录分析对氮负荷变化做出反应的信号通路。我的假设是，氮负荷的变化会导致信号分子的表达水平变化，从而导致尿素循环基因的激活或抑制。该假设将通过使用Affymetrix微阵列对RNA的转录分析来测量mRNA水平的变化来测量mRNA水平的变化。 3）确定蛋白质的量和翻译后修饰会随着氮负荷的变化而变化。我的假设是，氮负荷的变化触发了一种或多种信号通路中蛋白质的量和/或翻译后修饰的变化，从而导致尿素循环酶的表达和活性的变化。该假设将通过测量蛋白质水平的变化及其修饰，以响应使用两维凝胶电泳，光密度测定法和质谱法对变化的氮负荷进行修饰。