Towards a comprehensive protein interactome network in Schizosaccharomyces pombe

描述(由申请人提供)： 跨多个物种的基因组序列的戏剧性增长正在成倍增加预测的具有未知功能的蛋白质的数量。传统的“一次一个”表征因子的方法将始终是必不可少的，但它有助于对这些蛋白质之间的功能联系有一个全局的看法，以帮助产生特定的假设。研究蛋白质功能的一个重要方法是能够确定哪些其他蛋白质与其相关。在这个建议中，我们的目标是在模式生物S.pombe中系统地产生蛋白质-蛋白质相互作用(PPI)数据。除了发芽酵母外，裂解酵母S.pombe拥有所有真核模式生物中最容易操纵的基因组，但它在进化上与酿酒酵母截然不同。我们的目标是使用一个双管齐下的平台：酵母双杂交方法和共亲和纯化-质谱仪(AP/MS)方法来生成分裂酵母PPI图谱。这项工作汇集了两个人(Yu和Krogan)，他们共同工作，使用AP/MS策略在任何有机体(酿酒酵母)中生成了迄今为止最全面的共同复合体PPI图(Krogan等人，自然，2006年)。此外，最近，Yu还在萌芽酵母中生成了迄今为止最全面的双杂交数据集(Yu等人，《科学》，2008年)。因此，我们认为，在这两个小组之间，我们拥有实验和生物信息学专业知识，可以对分裂酵母中的蛋白质相互作用组进行全面的分析。从纯进化论的观点来看，由于这两种酵母菌大约在3.3亿-4.2亿年前分化，因此通过比较这两种生物的相互作用组网络将获得有价值的信息。此外，对S.pombe中相互作用网络的分析将提供从对S.cerevisiae进行的研究中无法获得的新信息。例如，在其复杂的着丝粒结构、纺锤体结构对有丝分裂进入的限制、冗余和低效的复制起点、组蛋白甲基化和染色质异染色质蛋白对基因的调控、RNAi途径对基因和转座子的调控、端粒结合蛋白以及大多数基因中内含子的存在等因素，庞贝葡萄球菌比酿酒酵母更接近后生动物。因此，这个项目不仅将对分裂酵母领域产生巨大影响，还将推动进化研究，并将允许审问 存在于高等有机体中但不存在于萌芽酵母中的生物系统。

DESCRIPTION (provided by applicant): The dramatic increase of genome sequences across multiple species is exponentially increasing the number of predicted proteins with unknown function. The traditional "one at a time approach" of characterizing factors will always be essential, but it helps to be able to have a global view of the functional connections between and among these proteins in order to aid in the generation of specific hypotheses. One important way to study protein function is to be able to ascertain which other proteins are associated with it. In this proposal, we aim to systematically generate a protein-protein interaction (PPI) data in the model organism, S. pombe. Next to budding yeast, the fission yeast S. pombe has the most easily manipulatable genome of any eukaryotic model organism, and yet it is evolutionarily distinct from S. cerevisiae. We aim to generate the fission yeast PPI map using a two-pronged platform: yeast two-hybrid methodology and co-affinity purification followed by mass spectrometry (AP/MS) approach. This work brings together two individuals (Yu and Krogan) who worked together to generate the most comprehensive co-complex PPI map to date in any organism (S. cerevisiae) using the AP/MS strategy (Krogan et al., Nature, 2006). Furthermore, more recently, Yu generated the most comprehensive two-hybrid dataset to date, also in budding yeast (Yu et al., Science, 2008). Therefore, we argue that, between the two groups, we have the experimental and bioinformatic expertise to carry out a comprehensive analysis of the protein interactome in fission yeast. From a purely evolutionary standpoint, valuable information would be obtained by comparing the interactome networks from both organisms since these two yeasts diverged around 330-420 million years ago. Additionally, analysis of interaction networks in S. pombe will provide novel information that cannot be obtained from studies performed with S. cerevisiae. For example, in its large complex centromere structure, the restriction of spindle construction to mitotic entry, redundant and inefficient origins of replication, gene regulation by histone methylation and chromodomain heterochromatin proteins, gene and transposon regulation by the RNAi pathway, telomere binding proteins, and the presence of introns in most genes, S. pombe is more similar to metazoans than is S. cerevisiae. Therefore, this project will not only have a huge impact on the fission yeast field but also will fuel evolutionary studies and will allow for interrogation of biological systems that are present in higher organisms but not in budding yeast.