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，但人们对真核生物如何正确代谢这种离子知之甚少。S.酿酒酵母将被用作研究锌代谢遗传学的模式生物，因为这种生物的整个基因组已经测序。此外，锌依赖性转录激活因子（ZAP 1）已被确定在这种生物体中，是负责调节基因的锌稳态。本研究的目的是使用全基因组分析来定义ZAP 1调节子。为了完成这一任务，将使用DNA微阵列技术，该技术允许在不同条件下（例如高锌浓度和低锌浓度）测定整个酵母基因组的比较表达水平。初步的微阵列实验显示，超过200个基因在缺锌的情况下显著改变了转录水平。这项建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，以及确定真正的ZAP 1调节的基因，而不仅仅是对锌缺乏的继发性影响敏感。一旦ZAP 1靶基因被鉴定，将选择几个最感兴趣的基因用于进一步的遗传研究，通过锌响应性的独立构象，然后通过所述基因的缺失分析以确定它们的功能。

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.