Yeast Gene Ontology by Phenotypic Profiling

描述(申请人提供)：过去20年的研究表明，在面包酵母中，酿酒酵母的基本细胞过程和执行这些过程的蛋白质与人类细胞中的蛋白质有很大的相似之处。这为通过与酿酒酵母和其他模式生物的基因产物的同源性、相互作用伙伴或功能相似性来鉴定人类蛋白质的功能提供了重要的手段。然而，尽管贝克酵母是大约8年前第一个被完全测序的真核生物，但这种模式生物中仍有大约30%的基因功能未知。这项建议的目标是通过在全基因组范围内对精心选择的250种处理方法的敏感性进行筛选，并结合对表型谱的复杂分析，来确定大量这些未表征基因的功能。我们的表型分析技术包括将识别每个缺失菌株的20个核苷酸“条形码”与高密度寡核苷酸阵列杂交。由此产生的对任何处理的表型图谱是一个敏感性的连续体，允许对治疗的表型图谱进行更准确的比较分析，而不是将表型“分类”为敏感、正常和抗性的常见程序。我们已经用大约50种不同的处理证明了表型谱的等级聚类是通过功能途径的相似性来聚类基因的一种强大的方法。我们建议为生物界创建一个资源，允许个体研究人员询问我们的数据，以寻找关于特定基因功能的假说，此外，还允许社区成员提出新的治疗方法，以帮助分配单个基因的功能。除了向社区提供表型图谱数据和分析外，我们还将重点研究在DMA修复和染色质重塑中发现的新基因，并评估这些基因在维持基因组稳定方面的参与程度。

DESCRIPTION (provided by applicant): Research over the past 20 years has demonstrated that in Baker's yeast, S. cerevisiae, the basic cellular processes, and the proteins that carry them out, have strong similarities to those in human cells. This provides an important means of identifying the function of human proteins by their homologies, interacting partners, or similarities of function with the gene products of S. cerevisiae as well as other model organisms. However, despite the fact that Baker's yeast was the first eukaryote to be fully sequenced some eight years ago, there are still some 30% of the genes of this model organism whose function is unknown. The goal of this proposal is to identify the function of a large number of these uncharacterized genes by performing a genome-wide screen of the sensitivities to 250 carefully chosen treatments of strains with individual deletion of all non-essential genes, coupled with sophisticated analyses of the phenotypic profiles. Our phenotypic profiling technique involves hybridization of the PCR-amplified 20-nucleotide "barcodes" that identify each deletion strain, to high-density oligonucleotide arrays. The resulting phenotypic profile to any treatment is a continuum of sensitivity that allows more accurate comparative analyses of phenotypic profiles to the treatments than the common procedure of "binning" phenotypes into sensitive, normal, and resistant. We have demonstrated with some 50 different treatments that hierarchical clustering of the phenotypic profiles is a powerful method of clustering the genes by similarity of functional pathways . We propose to create for the biological community a resource that will allow individual investigators to interrogate our data for hypotheses as to the function of specific genes and, in addition, allow members of the community to suggest new treatments to aid in assigning function to individual genes. In addition to providing the phenotypic profiling data and analyses to the community, we will focus on new genes identified in DMA repair and chromatin remodeling and assess the extent to which these genes are involved in maintaining genome stability.