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急剧增加正在指数地增加具有未知功能的预测蛋白质的数量。传统的“一次一个”的表征因素的方法将始终是必不可少的，但它有助于能够有一个整体的观点，这些蛋白质之间的功能连接，以帮助在特定的假设的产生。研究蛋白质功能的一个重要途径是能够确定哪些其他蛋白质与之相关。粟球。在芽殖酵母旁边，是分裂酵母S。粟酒裂殖酵母在任何真核模式生物中具有最容易操作的基因组，但它在进化上与S.酿酒。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，然后通过质谱（AP/MS）的方法产生裂变酵母PPI地图。这项工作汇集了两个人（Yu和Krogan），他们一起工作，生成了迄今为止在任何生物体中最全面的共复合物PPI图谱（S。使用AP/MS策略（Krogan等人，Nature，2006）。此外，最近，Yu也在芽殖酵母中生成了迄今为止最全面的双杂交数据集（Yu et al.，Science，2008）。因此，我们认为，在这两个小组之间，我们有实验和生物信息学的专业知识来对裂殖酵母中的蛋白质相互作用组进行全面分析。从纯进化的角度来看，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，因为这两种酵母在大约3.3亿至4.2亿年前分化。此外，还对S. pombe将提供不能从S.啤酒。例如，在其大的复杂的着丝粒结构中，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，通过组蛋白甲基化和染色质结构域异染色质蛋白的基因调控，通过RNAi途径的基因和转座子调控，端粒结合蛋白，以及大多数基因中内含子的存在，S. pombe比S更类似于后生动物。酿酒。因此，该项目不仅将对裂变酵母领域产生巨大影响，而且还将推动进化研究，并将允许对 存在于高等生物中但不存在于芽殖酵母中的生物系统。 公共卫生相关性： 虽然我们对特定疾病引起途径的了解一直在迅速增加，但我们利用这些知识用于治疗策略的能力受到不同蛋白质和途径的复杂相互连接性质的阻碍。在这项研究中，我们的目标是在模式生物S中生成蛋白质-蛋白质相互作用数据集。粟酒裂殖酵母，这将允许不同物种之间的物理相互作用组的进化比较，并将促进也存在于哺乳动物细胞中的许多途径的功能询问，其中许多在疾病状态下受到干扰。

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. PUBLIC HEALTH RELEVANCE: While our knowledge of specific disease causing pathways has been increasing rapidly, our ability to harness this knowledge for therapeutic strategies has been hampered by the complex inter-connected nature of the different proteins and pathways. In this study, we aim to generate a protein-protein interaction dataset in the model organism S. pombe which will allow for evolutionary comparison of physical interactomes across different species and will facilitate the functional interrogation of many pathways that also exist in mammalian cells, many of which are perturbed in disease states.