Function and structure of the RNA-processing exosome in the hyperthermophilic archaeon Sulfolobus solfataricus

虽然在真核生物和真细菌中对RNA加工和降解的不同方面进行了很好的研究，但在古细菌（生命的第三领域）中对这些基本过程知之甚少。我们从嗜热古菌Sulfolobus solfataricus中分离出一种蛋白质复合物，它是真核外泌体的简化版本。真核外泌体参与RNA的成熟和降解，其亚基对细胞活力至关重要。本项目旨在揭示S. solfataricus外泌体的生理功能，并鉴定其天然RNA底物。外泌体亚基的体内功能将通过s.s solfataricus基因沉默和条件酵母突变体的互补来研究。此外，亚基的生化活性将在体外测定。真核外泌体的确切结构至今还不能确定。我们喜欢利用古细菌外泌体的简单组织进行结构分析。我们将研究单个亚基之间的蛋白质-蛋白质相互作用，并重建不同组成的复合物进行电子显微镜和晶体学分析。我们的研究结果将为未来古细菌转录后水平基因调控的研究奠定基础，将有助于我们对高温下蛋白质-蛋白质和RNA-蛋白质相互作用的理解，并将为RNA加工蛋白复合物的进化提供有趣的方面。

While different aspects of RNA processing and degradation are well studied in Eubacteria and Eukarya, little is known about these essential processes in Archaea, the third domain of life. We have isolated a protein complex from the hyperthermophilic archaeon Sulfolobus solfataricus which is a simplified version of the eukaryotic exosome. The eukaryotic exosome is involved in maturation and degradation of RNA, its subunits are essential for cell viability. This project aims to unravel the physiological function of the S. solfataricus exosome and to identify its natural RNA substrates. The in vivo function of the exosome subunits will be studied by gene silencing in S. solfataricus and by complementation of conditional yeast mutants. In addition the biochemical activity of the subunits will be determined in vitro. The exact structure of the eukaryotic exosome could not be solved to date. We like to take advantage of the simpler organization of the archaeal exosome for structural analyses. We will study protein-protein interaction between the individual subunits and reconstitute complexes of different composition for electron microscopic and crystallographic analyses. Our results will build the fundament for future investigations of gene regulation at post-transcriptional level in Archaea, will contribute to our understanding of protein-protein and RNA-protein interactions at high temperatures and will provide interesting aspects on the evolution of RNA processing protein complexes.