Metal Homeostatic Mechanisms in Yeast

酵母中的金属稳态机制

• 批准号: 6575831
• 负责人: Dennis R. Winge
• 金额: $ 22.47万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-12-15 至 2007-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6575831
• 关键词: DNA binding protein; Saccharomyces cerevisiae; alternatives to animals in research; biological signal transduction; copper; fluorescence resonance energy transfer; gene expression; genetic promoter element; green fluorescent proteins; homeostasis; intermolecular interaction; iron; matrix assisted laser desorption ionization; metal metabolism; microorganism culture; microorganism metabolism; molecular cloning; polymerase chain reaction; transcription factor; yeast two hybrid system; yeasts

DESCRIPTION (provided by applicant): This application focuses on homeostatic coupling of intracellular copper (Cu) and iron (Fe) levels to gene expression in the yeast Saccharomyces cerevisiae. Metalloregulation of Cu uptake genes is controlled by the Mac1 transcriptional activator. Metalloregulation of Fe uptake genes is controlled by two related activators, Aft1 and Aft2. These three factors are transcriptional activators in metal-deficient cells, but specifically inhibited in metal-replete cells. The major objectives are to elucidate the mechanisms of Cu ion sensing by Mac1 and Fe ion sensing by Aft1 and Aft2. Many interconnections exist between Fe and Cu homeostasis in yeast. In the past grant cycle we provided evidence suggestive that Mac1 senses Cu ions within the yeast nucleus through direct binding of Cu(I) ions by Mac1. Cu(I) binding appears to induce an intramolecular interaction between the N-terminal DNA binding domain and C-terminal transactivation domain. Studies are proposed to verify direct sensing of Cu and to map the intramolecular interface that inhibits both DNA binding and transactivation. An important goal is to identify other yeast proteins important in either Cu shuttling to the nucleus or Cu inhibition of Mac1. Fe-homeostasis in yeast is regulated by Aft1 and Aft2. The mechanism of Fe inhibition of Aft1 and Aft2 function is unknown. The significance of these studies is that S. cerevisiae is the best model eukaryotic system to understand the mechanism of Fe sensing by a transcription regulator. We propose to determine whether Fe sensing occurs through direct Fe binding, Fe-mediated post-translational modification or through another protein? Aft1 and Aft2 exhibit limited functional redundancy in yeast. We propose to determine whether each exhibits a distinct physiological function. Mac1 and Aftl/Aft2 are excellent model eukaryotic systems to study nutritional control of gone expression. Information gleaned on the mechanism of Cu- and Fe-regulation may be applicable to other species.

描述（由申请人提供）：此申请的重点是细胞内铜（CU）和铁（Fe）水平的稳态耦合，以酿酒酵母中的基因表达。 Cu摄取基因的金属调节受到MAC1转录激活剂的控制。 Fe摄取基因的金属调节由两个相关激活剂AFT1和AFT2控制。这三个因素是金属缺陷细胞中的转录激活剂，但在金属复发细胞中特别抑制。 主要目标是阐明Mac1和Fe Ion Seensing通过AFT1和AFT2传感的Cu离子传感的机制。 Fe和Cu稳态之间存在许多互连。在过去的赠款周期中，我们提供了证据表明，Mac1通过MAC1直接结合Cu（I）离子在酵母核中感知Cu离子。 Cu（i）结合似乎诱导了N末端DNA结合结构域和C末端反式激活结构域之间的分子内相互作用。提出了研究以验证CU的直接感测，并绘制抑制DNA结合和反式激活的分子内界面。 一个重要的目标是鉴定其他在CU穿梭至核或MAC1抑制Cu的酵母蛋白。 酵母中的Fe Homeostasis受AFT1和AFT2的调节。 Fe抑制AFT1和AFT2功能的机制尚不清楚。 这些研究的意义在于，酿酒酵母是了解转录调节剂Fe感应机制的最佳模型真核系统。 我们建议通过直接Fe结合，Fe介导的翻译后修饰或通过另一种蛋白质来确定Fe感测？ AFT1和AFT2在酵母中表现出有限的功能冗余。 我们建议确定每个表现出不同的生理功能。 MAC1和AFTL/AFT2是出色的模型真核系统，用于研究对消失表达的营养控制。 有关Cu-和Fe-调节机制的信息可能适用于其他物种。