COPPER HOMEOSTATIC MECHANISMS IN YEAST

描述：(申请人摘要)铜是一种必需的，但潜在的 有毒的营养物质。存在调节细胞内环境平衡的机制 铜离子的浓度，从而保持铜的平衡，并将 铜离子的有害作用。金属传感系统是 金属离子动态平衡。来自这些系统的信号通常会导致特定的 金属--动态平衡中蛋白质生物合成的调节。在……里面 在酵母中，铜的调控包括铜诱导和铜诱导 铜介导的转录抑制。这笔赠款的重点是机制 酵母细胞通过它来感知细胞内铜离子的水平并调节 以铜依赖的方式表达基因。表达了一系列的 编码酵母中铜离子吸收的产物的基因是特异的 被铜离子抑制。我们证明了基因的铜衰减 表达是通过铜介导的抑制反式激活发生的 MAC1转录因子的活性。对其作用机制的阐明 铜介导的MAC1抑制是这一提议的一个主要焦点。 MAC1中的激活结构域活性映射到包含两个富含半胱氨酸的区域 序列基序。需要检验的一个假设是，铜(I)与 MAC1激活结构域中富含CyS的基序是 铜-MAC1功能的抑制。实验概述是为了辨别是否 抑制通过分子内或分子间过程发生。一个 MAC1研究的显著特点是，例子有限 转录激活功能的调节。超额条件 环境中的铜离子导致a基因的诱导表达 不同的基因子集，其产物具有保护作用。 ACE1介导铜诱导酵母基因表达 转录因子。ACE1的铜激活需要铜(I)离子 存在于细胞核内。我们假设特定的核中的铜(I) 转运蛋白的存在类似于已知的铜转运蛋白 线粒体和分泌途径。我们的目标是鉴定基因 铜激活ACE1所需的产物。如果一位候选人 核铜转运体被确认，我们想要确定它是否也 参与铜(I)的核转运，调节MAC1功能。

DESCRIPTION: (applicant's abstract) Copper is an essential, but potentially toxic nutrient. Homeostatic mechanisms exist to regulate the cellular concentration of copper ions, thus maintaining Cu balance and minimizing the deleterious effects of Cu ions. Metal sensory systems are one facet of metal ion homeostasis. Signals from these systems often result in specific metal-regulation of biosynthesis of proteins involved in homeostasis. In yeast, copper regulation consists of both Cu-mediated induction and Cu-mediated inhibition of transcription. This grant focuses on mechanisms by which yeast cells sense the intracellular level of Cu ions and regulate gene expression in a Cu-dependent manner. The expression of a number of genes encoding products involved in Cu ion uptake in yeast is specifically inhibited by Cu ions. We demonstrated that Cu-attenuation of gene expression occurs through Cu-mediated repression of the transactivation activity of the Mac1 transcription factor. Elucidation of the mechanism of Cu-mediated repression of Mac1 is a major focus of this proposal. Activation domain activity in Mac1 maps to a region containing two Cys-rich sequence motifs. One hypothesis to be tested is that Cu(I) binding to the Cys-rich motifs in the Mac1 activation domain is an initial event in Cu-repression of Mac1 function. Experiments are outlined to discern whether repression occurs through an intramolecular or intermolecular process. A significant feature of the Mac1 studies is that there are limited examples of regulation of transcriptional activation function. Conditions of excess copper ions in the environment result in the induced expression of a different subset of genes whose products have protective roles. Cu-induction of gene expression in yeast is mediated by the Ace1 transcription factor. Cu-activation of Ace1 requires Cu(I) ions to be present within the nucleus. We postulate that a specific nuclear Cu(I) transporter exists analogous to known Cu transporters specific for the mitochondrion and the secretory pathway. Our objective is to identify gene products that are required for Cu-activation of Ace1. If a candidate nuclear Cu transporter is identified, we want to determine whether it also participates in nuclear transport of Cu(I) for regulation of Mac1 function.