Département de biochimie, de microbiologie et de bio-informatique, Faculté des sciences et de génie, Groupe de recherche en écologie buccale, Faculté de médecine dentaire, Félix d'Hérelle Reference Center for Bacterial Viruses, Université Laval, Quebec City, Quebec G1V 0A6, Canada.
Nature. 2010 Nov 4;468(7320):67-71. doi: 10.1038/nature09523.
Bacteria and Archaea have developed several defence strategies against foreign nucleic acids such as viral genomes and plasmids. Among them, clustered regularly interspaced short palindromic repeats (CRISPR) loci together with cas (CRISPR-associated) genes form the CRISPR/Cas immune system, which involves partially palindromic repeats separated by short stretches of DNA called spacers, acquired from extrachromosomal elements. It was recently demonstrated that these variable loci can incorporate spacers from infecting bacteriophages and then provide immunity against subsequent bacteriophage infections in a sequence-specific manner. Here we show that the Streptococcus thermophilus CRISPR1/Cas system can also naturally acquire spacers from a self-replicating plasmid containing an antibiotic-resistance gene, leading to plasmid loss. Acquired spacers that match antibiotic-resistance genes provide a novel means to naturally select bacteria that cannot uptake and disseminate such genes. We also provide in vivo evidence that the CRISPR1/Cas system specifically cleaves plasmid and bacteriophage double-stranded DNA within the proto-spacer, at specific sites. Our data show that the CRISPR/Cas immune system is remarkably adapted to cleave invading DNA rapidly and has the potential for exploitation to generate safer microbial strains.
细菌和古菌已经开发出几种防御策略来对抗外来核酸，如病毒基因组和质粒。其中，成簇规律间隔短回文重复序列 (CRISPR) 位点与 cas (CRISPR 相关) 基因一起构成了 CRISPR/Cas 免疫系统，该系统涉及部分回文重复序列，由称为间隔区的短 DNA 片段隔开，这些间隔区是从染色体外元件中获得的。最近的研究表明，这些可变的基因座可以从感染噬菌体的基因组中整合间隔区，然后以序列特异性的方式提供对后续噬菌体感染的免疫力。在这里，我们展示了嗜热链球菌的 CRISPR1/Cas 系统也可以从含有抗生素抗性基因的自我复制质粒中自然获得间隔区，从而导致质粒丢失。与抗生素抗性基因匹配的获得的间隔区为自然选择不能摄取和传播这些基因的细菌提供了一种新方法。我们还提供了体内证据表明，CRISPR1/Cas 系统可以在原间隔区的特定位置特异性切割质粒和噬菌体的双链 DNA。我们的数据表明，CRISPR/Cas 免疫系统非常适合快速切割入侵的 DNA，并且有可能被开发利用来产生更安全的微生物菌株。
