The CRISPR system: a new frontier in prokaryotic molecular biology

病毒与它们试图感染和破坏的细胞之间的斗争是分子进化的主要驱动力之一。原核生物利用“限制性内切酶”来定位和降解入侵的DNA，这种酶的发现彻底改变了分子生物学。真核生物，包括人类，使用RNA干扰(RNAi)途径来靶向和降解病毒RNA。在过去的几年里，RNAi途径的一个原核版本，即所谓的CAS-CRISPR系统，已经浮出水面。病毒序列以“间隔区”的形式存储在宿主基因组中，两侧是直接重复序列(CRISPR)。众所周知，宿主细胞将新的病毒DNA整合到CRISPR中，这赋予了对未来病毒感染的免疫力。CRISPR系统的作用方式被认为与RNAi途径类似。CRISPR序列与一些广泛保守的‘CAS’蛋白质(每个基因组多达50个)有关，这些蛋白质被认为负责CRISPR的处理、病毒防御和新病毒序列的整合。这些蛋白质的功能还不清楚。我们拥有研究模式生物Sulfolobus solfararicus中所有CaS蛋白的经验和技术诀窍。将解决CAS蛋白质的结构，并描绘CAS蛋白质之间的相互作用。我们的实验室将使用常规使用的各种技术来测定单个CaS蛋白质和复合体的活性。最终目的是从分子上了解CRISPR系统。除了为原核生物生物学提供基本的新见解外，这项工作还可能带来操纵有用生物体基因组(合成生物学)或治疗重要微生物病原体的新方法。

The battle between viruses and the cells they attempt to infect and subvert is one of the main driving forces in molecular evolution. Prokaryotes target and degrade invading DNA using 'restriction enzymes', the discovery of which revolutionised molecular biology. Eukaryotes, including humans, use the RNA interference (RNAi) pathway to target and degrade viral RNA. In the last few years a prokaryotic version of the RNAi pathway, the so-called CAS-CRISPR system, has come to light. Viral sequences are stored in the host genome as 'spacers' flanked by direct repeat sequences (CRISPRs). It is known that host cells incorporate new viral DNA into the CRISPRs, and that this confers immunity against future viral infection. The CRISPR system is envisaged to act in an analogous manner to the RNAi pathway A number of widely conserved 'Cas' proteins (up to 50 per genome) are associated with the CRISPR sequences, and these are thought to be responsible for CRISPR processing, viral defence and the incorporation of new viral sequences. The functions of these proteins are not clearly understood. We have the experience and technological know-how to study all the Cas proteins from the model organism Sulfolobus solfataricus. Cas protein structures will be solved, and Cas protein interactions delineated. The activities of individual Cas proteins and complexes will be determined using a variety of techniques in routine use in our laboratories. The ultimate aim is to acheive a molecular understanding of the CRISPR system. As well as providing fundamental new insights into prokaryotic biology, the work may lead to new methods to manipulate the genomes of useful organisms (synthetic biology) or to treatments for important microbial pathogens.