Defining New Functions in DNA Replication and DNA Damage Response Genes

科学目标：在萌芽中的酿酒酵母中，全基因组的突变菌株已被广泛用于大规模阐明生物学功能。由此产生的信息已被用于在途径中放置基因，识别不同途径之间的交叉点，并将基因功能分配给新的蛋白质。这些研究的一个缺陷源于这些全基因组样本中突变试剂的性质，这些样本中的基因功能已因缺失或条件性耗尽而失活。由于许多(如果不是全部)蛋白质执行不止一种功能，这种突变具有潜在的多效性。一个潜在的解决方案是利用功能分离突变来消除蛋白质的单一生物功能。然而，在过去，即使对单基因来说，识别这类特殊的等位基因也是一个逻辑障碍。这一研究项目是基于一种新开发的大规模分离功能等位基因的策略，该策略将应用于一组涉及DNA复制和对DNA损伤的反应的相互关联的基因。由于这些基因中很少有突变被分离出来，这项研究应该定义DNA复制和DNA损伤反应途径中的新功能。更广泛的影响：该研究项目的科学目标与两个更广泛的影响密切相关。首先，由此产生的突变将被酵母社区广泛获得，特别关注将这些新试剂纳入全基因组系统分析。由功能分离错义突变构建的遗传网络可能会发现以前未被认识到的接口，这些接口在使用目前可用的突变株的先前系统分析中被遗漏了。其次，该项目将严重依赖初级研究人员(本科生和高中生)，他们将负责产生功能分离突变面板。这种经历将允许非常初级的研究人员在重要的研究项目中发挥核心作用，并向他们介绍指导在生物医学研究社区中发挥的关键作用。

Scientific goals: In the budding yeast S. cerevisiae, genome-wide collections of mutant strains have been employed extensively to elucidate biological function on a large scale. The resulting information has been used to place genes in pathways, to identify points of intersection between different pathways and to assign gene function to novel proteins. One flaw of these studies stems from the nature of the mutant reagents in these genome-wide collections, where gene function has been inactivated by either deletion or conditional depletion. Since many, if not all, proteins execute more than one function, such mutations are potentially pleiotropic. A potential solution is to employ separation-of-function mutations that eliminate a single biological function of a protein. However, identification of this particular class of alleles in the past has been a logistic hurdle even for single genes. This research project is based on a newly developed strategy for large-scale isolation of separation-of-function alleles which will be applied to a set of inter-related genes involved in DNA replication and response to DNA damage. Since very few mutations have been isolated for many of these genes, this research should define new functions in the DNA replication and DNA damage response pathways. Broader impacts: The scientific goals of this research project are closely intertwined with two broader impacts. First, the resulting mutations will be made widely available to the yeast community, with a particular focus on incorporation of these new reagents into genome-wide systems analysis. Genetic networks constructed from separation-of-function missense mutations are likely to uncover previously unappreciated interfaces that were missed in prior systems analysis which employed currently available mutant strains. Second, this project will rely heavily on entry-level researchers (undergraduate and high school students) who will be responsible for generating the panels of separation-of-function mutations. This experience will allow very junior researchers to play a central role in a significant research project and also introduce them to the critical function that mentoring plays in the biomedical research community.