How does a chimeric partition machine mediate chromosome segregation in Archaea?

古生菌在数十亿年前演化为生命的第三个领域，但它们是相对较新的生物有机体普遍树地图上的新成员。他们37年前的发现代表着一个重要的生物学里程碑。古生菌是单细胞生物，与细菌和真核生物一起存在于我们的星球上。细菌和古菌都是原核生物，即它们的遗传物质不被膜包裹到一个单独的隔室，称为细胞核，这是真核生物(面包酵母、真菌、植物、动物和人类等)的标志。古生菌最初是从极端生态系统中分离出来的，现在人们知道它无处不在，构成了生物圈的相当大一部分。例如，据报道，仅世界海洋就有大约1.3x10到28个古细菌细胞：这是一个巨大的数字。作为比较，地球上所有海滩上的沙粒数量估计为7.5x10到18，与海洋古细菌相比，这个数量仍然要少得多。它们的无处不在和丰富使它们成为调节地球上全球生物地球化学循环的关键角色。从功能和机制的角度来看，古生菌是细菌和真核生物的镶嵌，但它们也具有独特的分子特征，如产生甲烷。嗜热古生菌是超级微生物，在80摄氏度和更高的温度下在温泉、火山、深海喷口生长，并显示出不同寻常的特性，这使得这些生物对于开发新的生物技术应用非常有价值，但对于被推向极端的生命的基础研究也非常有趣。在嗜热古菌中发现的耐热分子(例如蛋白质和脂链)向我们揭示了我们所知的生命的边界可以比之前预期的更远地推进。它们在其他陆地生物无法生存的极端环境中生长的能力，也重新燃起了在不适宜居住的星球上发现外星生命的希望。尽管在过去30年里，这些生物在破译分子机制方面取得了重大进展，但到目前为止，关于古生物染色体分离的基本过程的信息很少，这个课题仍然是一个等待研究的黑匣子。基因组分离是每个细胞生命周期的关键阶段：遗传物质首先复制，然后分离并平均分配到两个子细胞中。我们打算在嗜热古生物Sulfolobus solfararicus中剖析这一过程，它的基因组编码两种蛋白质，SegA和SegB，这两种蛋白质相互作用形成一个简单的染色体分离机器。该项目旨在发现SegAB复合体在细胞分裂时调节索尔法特链霉菌染色体分离和均匀分布的机制。我们将通过将它们融合到一个热稳定的荧光蛋白，并使用传统的显微镜和一种名为超分辨率显微镜的新成像技术来阐明这些蛋白质在细胞中的定位。这种方法将使我们能够获得由SegA和SegB在细胞中形成的结构的高分辨率图像。我们还希望研究每种蛋白质与DNA的相互作用，以绘制它们在染色体上的结合位置，并了解这些结合是如何导致染色体分离的。这项工作的另一个目标是识别与细胞内SegAB复合体相互作用的其他蛋白质：将探索两种不同的筛选策略，一种寻找基因，另一种寻找潜在伴侣的蛋白质。此外，我们还想确定SegA和SegB的三维结构。从各种研究中得到的多个拼图将被组合在一起，以产生一张关于索尔法里克氏链霉菌染色体分离的详细图片。

Archaea evolved as the third domain of life billions of years ago, but they are a relatively recent addition to the map of the universal tree of living organisms. Their discovery 37 years ago represented a major biological milestone. Archaea are unicellular organisms that populate our planet together with bacteria and eukaryotes. Both bacteria and archaea are prokaryotes, i.e. their genetic material is not wrapped by a membrane into a separate compartment, called nucleus, which is instead a hallmark of eukaryotes (baker yeast, fungi, plants, animals and humans to mention some). Initially isolated from extreme ecosystems, archaea are now known to be ubiquitous, constituting a considerable fraction of the biosphere. For example, it has been reported that the world ocean alone contains approximately 1.3 x 10 to the 28 archaeal cells: this is an enormous number. To provide a comparison, the estimated number of grains of sand on all the beaches on earth is 7.5 x 10 to the 18, a quantity still much smaller as compared with that of marine archaeal cells. Their ubiquity and abundance make them key players in regulating global biogeochemical cycles on Earth. From a functional and mechanistic standpoint, archaea are a mosaic of tesserae from bacteria and eukaryotes, but they are also characterized by unique molecular features like methane production.Thermophilic archaea are super microbes thriving at 80 degrees C and higher temperatures in hot springs, volcanoes, deep sea vents and exhibiting unusual properties, which make these organisms valuable for the development of novel biotechnological applications, but also extremely interesting for basic studies on life pushed to extremes. The heat resistant molecules found in thermophilic archaea (for example proteins and lipidic chains) have revealed to us that the boundaries of life as we know it can be pushed much further than previously anticipated. Their ability to grow in extreme environments where no other terrestrial organism can survive has also rejuvenated hopes of discovering extraterrestrial life on inhospitable planets.Despite the significant progress made in decoding molecular mechanisms in these organisms in the last three decades, to date little information is available on the fundamental process of chromosome segregation in archaea and the subject remains a black box awaiting investigation. Genome segregation is a crucial stage of the life cycle of every cell: the genetic material is first duplicated, then separated and equally distributed into the two daughter cells. We intend to dissect this process in the thermophilic archaeon Sulfolobus solfataricus, whose genome encodes for two proteins, SegA and SegB, which interact to form a simple chromosome segregation machine. The proposed project intends to discover the mechanisms adopted by the SegAB complex to mediate the separation and equi-distribution of chromosomes in S. solfataricus at cell division. We will shed light on the localization of these proteins in the cell by fusing them to a heat stable fluorescent protein and using conventional microscopy and a novel imaging technique, called super resolution microscopy. This approach will allow us to acquire a high-resolution picture of the structures formed by SegA and SegB in the cell. We also wish to investigate the interaction of each of the proteins with DNA to map their binding sites on the chromosome and to understand how these associations result in chromosome segregation. Another aim of the work is to identify other proteins that interact with the SegAB complex inside the cell: two different screening strategies will be expolited, one looking for genes and the other looking for proteins of potential partners. In addition, we want to determine the three-dimensional structure of SegA and SegB. The multiple pieces of the jigsaw deriving from the various investigations will be combined to generate a detailed picture of chromosome segregation in S. solfataricus