MINING THE YEAST GENOME FOR ZINC REGULATED GENES

锌是一种必需的微量营养素，在真核细胞内被大量利用，但尽管锌的相对重要性，我们对真核生物如何适当地代谢这种离子知之甚少。酿酒葡萄球菌的全基因组测序已经完成，将作为研究锌代谢遗传学的模式生物。此外，锌依赖性转录激活因子（ZAP1）在这种生物体中也被鉴定出来，它负责调节锌体内平衡的基因。本研究的目的是利用全基因组分析来确定ZAP1调控子。为了完成这项任务，将使用DNA微阵列技术来确定整个酵母基因组在不同条件下的比较表达水平，例如高锌和低锌浓度。初步的微阵列实验显示，在锌缺乏的情况下，超过200个基因的转录水平显著改变。这项提议的总体设计是重复和完善这些研究，以获得更可靠的数据，并确定真正受ZAP1调控的基因，而不是简单地对锌缺乏的继发性影响敏感。一旦确定了ZAP1靶基因，将通过锌响应性的独立构象，然后通过对所述基因的缺失分析来确定其功能，选择几个最有趣的基因进行进一步的遗传研究。

Zinc is an essential micronutrient that is abundantly utilized inside eukaryotic cells, but despite the relative importance of zinc, very little is known about how eukaryotic organisms properly metabolize this ion. S. cerevisiae will be used as a model organism to study the genetics of zinc metabolism, since the entire genome of this organism has been sequenced. Also a zinc dependent transcriptional activator (ZAP1) has been identified in this organism that is responsible for regulating genes in zinc homeostasis. The purpose of this study will be to use genome wide analysis to define a ZAP1 regulon. To accomplish this task, DNA microarray technology will be used that allows for the determination of comparative expression levels of the entire yeast genome under different conditions, such as high and low zinc concentrations. Preliminary microarray experiments revealed that over 200 genes have significantly altered transcription levels under zinc deficiency. The general design of this proposal is to repeat and refine these studies to obtain more reliable data as well as to identify genes that are truly ZAP1 regulated rather than simply sensitive to the secondary effects of zinc deficiency. Once ZAP1 target genes have been identified several of the most interesting will be selected for further genetic study by independent conformation of the zinc responsiveness then by deletional analysis of said genes to determine their function.