Mechanisms of Glucose Signal Transduction in Yeast

酵母中葡萄糖信号转导机制

基本信息

批准号：
7254131
负责人：
Martin C Schmidt
金额：
$ 30.2万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
1991
资助国家：
美国
起止时间：
1991-07-01 至 2008-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7254131
关键词：
5&apos -AMP-activated protein kinase Alanine Alleles Binding Biochemical Genetics Biological Assay C-terminal Cells Client Co-Immunoprecipitations Complex Condition Discrimination Drug usage Enzymes Event GLC2 protein Gene Deletion Glucose Grant Homologous Gene In Vitro Length Mammalian Cell Maps Mass Spectrum Analysis Measures Messenger RNA Metformin Molecular Chaperones Molecular Conformation Mutagenesis Non-Insulin-Dependent Diabetes Mellitus Northern Blotting Numbers Nutrient Pathway interactions Phosphoric Monoester Hydrolases Phosphorylation Phosphorylation Site Phosphotransferases Play Post-Transcriptional Regulation Protein Kinase Proteins Regulation Research Personnel Role Role playing therapy Saccharomyces cerevisiae Saccharomycetales Signal Pathway Signal Transduction Site Stress Testing Ubiquitin Western Blotting Yeasts genetic analysis genetic regulatory protein in vivo member mutant novel programs protein protein interaction response upstream kinase

项目摘要

DESCRIPTION (provided by applicant): The Snf1 protein kinase plays a central role in nutrient stress signaling in the budding yeast, Saccharomyces cerevisiae. The mammalian homologue of Snf1, the AMP-activated protein kinase (AMPK) plays an equally important role in mammalian cells. Recently AMPK was shown to be the target of metformin, the most widely prescribed drug used to treat type 2 diabetes. We propose to study the regulation of this medically important enzyme in yeast. In the previous grant period we developed an assay for the phosphorylation state of the Snf1 activation loop. We showed that phosphorylation of Snf1 was a critical step for the activation of the Snf1 kinase and that the phosphorylation of Snf1 was catalyzed by a distinct upstream kinase. Recently, we were able to demonstrate that there are three upstream kinases (Pakl, Tos3 and Elm1) that are capable of activating Snf1 in vivo. The identification of the Snf1 activating kinases was highly significant because it immediately led to the identification of the mammalian kinase that activates AMPK. In the next grant period we propose to continue our studies of the regulation of the Snf1 kinase signaling pathway. We will focus on upstream events in this pathway and determine the mechanism that regulate the activity of the three Snf1-activating kinases. We will determine whether there are distinct forms of the Snf1 kinase complex that are specialized for response to different stresses. We will assess the regulatory significance of phosphorylation sites in the Snf1 kinase complex that we have mapped by mass spectrometry. Finally, biochemical and genetic analyses have identified a number of candidates for regulators of the Snf1 kinase signaling pathway. These candidates proteins will be studied to determine if they associate with and regulate the activity of the Snf1 kinase.

描述（由申请人提供）：Snf1 蛋白激酶在芽殖酵母（酿酒酵母）的营养应激信号传导中发挥核心作用。 AMP 激活蛋白激酶 (AMPK) 是 Snf1 的哺乳动物同源物，在哺乳动物细胞中发挥着同样重要的作用。最近，AMPK 被证明是二甲双胍的靶标，二甲双胍是治疗 2 型糖尿病最广泛使用的处方药物。我们建议研究酵母中这种医学上重要的酶的调节。在之前的资助期内，我们开发了一种针对 Snf1 激活环磷酸化状态的检测方法。我们发现 Snf1 的磷酸化是 Snf1 激酶激活的关键步骤，并且 Snf1 的磷酸化是由独特的上游激酶催化的。最近，我们能够证明三种上游激酶（Pakl、Tos3 和 Elm1）能够在体内激活 Snf1。 Snf1 激活激酶的鉴定非常重要，因为它立即导致了激活 AMPK 的哺乳动物激酶的鉴定。在下一个资助期内，我们建议继续研究 Snf1 激酶信号通路的调节。我们将重点关注该途径的上游事件，并确定调节三种 Snf1 激活激酶活性的机制。我们将确定是否存在专门针对不同压力做出反应的不同形式的 Snf1 激酶复合物。我们将评估通过质谱法绘制的 Snf1 激酶复合物中磷酸化位点的调控意义。最后，生化和遗传分析确定了许多 Snf1 激酶信号通路调节剂的候选者。将研究这些候选蛋白以确定它们是否与 Snf1 激酶活性相关并调节其活性。