METALLOREGULATORY TRANSCRIPTION FACTORS IN YEAST

铜（Cu）作为电子转移中间体的能力 使其成为多种酶的重要辅助因子， 基本的生物化学功能;然而，同样的“属性”也是 负责其强大的细胞毒性。因此，至关重要的是 生物体维持一种严格调节的铜感应机制， 铜的基本和毒性水平之间的微妙平衡。研究 该提案中概述的目标是描述一个新成员的特点， 铜金属调节转录因子（CuMRTF）家族，命名为 MRTF-X已知的酵母CuMRTF家族成员ACE 1、AMT和MAC 1 直接感应铜并通过激活金属的转录来响应 解毒基因称为金属硫蛋白在高浓度铜， 和一种称为FRE 1的铜、铁还原基因， 在低Cu浓度下的亲和力Cu运输。的部分序列 MRTF-X的氨基末端110个氨基酸是44%和53%相同的， 与AMT 1和ACE 1相同区域的相似性分别为65%和68 其参与Cu依赖性DNA结合活性。在这 应用，结构，功能，表达，亚细胞定位， 并且MRTF-X的靶DNA序列将通过基因缺失来表征 诱变，DNA结合试验，北方印迹分析，体内基因 表达测定和间接免疫荧光显微术。本研究 将提供关于参与《公约》的机制的重要信息， 维持正常铜含量的基因的转录调节 体内平衡

The ability of copper (Cu) to act as an electron transfer intermediate makes it an important co-factor in a variety of enzymes that perform essential biochemical functions; however, this same' property is also responsible for its potent cytotoxicity. Thus, it is critical that organisms maintain a tightly regulated Cu sensing mechanism to maintain the delicate balance between essential and toxic levels of Cu. The studies outlined in this proposal are aimed at characterizing a new member of the Cu Metalloregulatory Transcription Factor (CuMRTF) family, designated MRTF-X. The known members of the yeast CuMRTF family, ACE1, AMT, and MAC1 directly sense Cu and respond by activating the transcription of metal detoxification genes known as metallothioneins at high Cu concentrations, and a Cu,Fe reducing gene known as FRE1 which is required for high affinity Cu transport at low Cu concentrations. The partial sequence of the amino terminal 110 amino acids of MRTF-X is 44% and 53% identical and 65% and 68% similar to the same region in AMT1 and ACE1, respectively which is involved in a Cu-dependent DNA binding activity. In this application, the structure, function, expression, subcellular location, and target DNA sequences of MRTF-X will be characterized by gene deletion mutagenesis, DNA binding assays, Northern blot analysis, in vivo gene expression assays and indirect immunofluorescence microscopy. This study will provide important information on the mechanisms involved in the transcriptional regulation of the genes that maintain normal Cu homeostasis.