Chromosomal Targeting by the Type III-A CRISPR-Cas System Can Reshape Genomes in Staphylococcus aureus.
Chromosomal Targeting by the Type III-A CRISPR-Cas System Can Reshape Genomes in Staphylococcus aureus.
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DOI：
10.1128/msphere.00403-17
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发表时间：
2017-11
期刊：
影响因子：
4.8
通讯作者：
Sun B
中科院分区：
文献类型：
--
作者：
Guan J;Wang W;Sun B
金黄色葡萄球菌是一种可引起人类广泛感染的病原体。研究表明，CRISPR-Cas 系统可以通过染色体靶向导致整合移动遗传元件 (MGE) 的丢失。在这里，我们证明 CRISPR 介导的切割有助于部分删除耐甲氧西林金黄色葡萄球菌 (MRSA) 中整合的 SCCmec，这为治疗 MRSA 感染提供了一种策略。人工 CRISPR 阵列中的间隔区应包含超过 25 个核苷酸以实现免疫，并且选定靶标与 crRNA 5' 标签之间的连续三核苷酸配对可以阻止靶向。这些发现增加了我们对 III-A 型 CRISPR-Cas 系统分子机制的理解，并为在临床和工业应用中开发针对细菌中整合 MGE 的工程 CRISPR 免疫提供了新的策略。 CRISPR-Cas（成簇规则间隔短回文重复序列 [CRISPR]-CRISPR 相关蛋白 [Cas]）系统可以通过使用 CRISPR RNA (crRNA) 作为定位和降解目标 DNA 的指导来提供针对入侵遗传元件的保护。根据cas基因的含量，CRISPR-Cas系统分为两类五类。此前的研究表明，CRISPR-Cas系统可以避免病毒感染并阻止质粒转移。在这里，我们证明金黄色葡萄球菌 III-A 型 CRISPR-Cas 系统的染色体靶向可以驱动集成葡萄球菌盒式染色体 mec (SCCmec) 内的大规模基因组删除和改变。 CRISPR-Cas系统的靶向活性与crRNA和原型间隔区之间的互补性有关，间隔区的10至13个核苷酸截断部分阻断CRISPR攻击，超过13个核苷酸截断可以完全废除靶向，这表明需要最小长度来许可切割。避免原型间隔子上游区域的碱基配对对于 CRISPR 靶向也是必要的。 crRNA 的 5' 标签和原型间隔子之间的连续三核苷酸互补性可能会破坏靶向。我们的研究结果表明，III-A 型 CRISPR-Cas 系统可以调节细菌基因组稳定性，并可能作为删除细菌中抗性或毒力基因的高效工具。重要性 金黄色葡萄球菌是一种可引起人类多种感染的病原体。研究表明，CRISPR-Cas 系统可以通过染色体靶向导致整合移动遗传元件 (MGE) 的丢失。在这里，我们证明 CRISPR 介导的切割有助于部分删除耐甲氧西林金黄色葡萄球菌 (MRSA) 中整合的 SCCmec，这为治疗 MRSA 感染提供了一种策略。人工 CRISPR 阵列中的间隔区应包含超过 25 个核苷酸以实现免疫，并且选定靶标与 crRNA 5' 标签之间的连续三核苷酸配对可以阻止靶向。这些发现增加了我们对 III-A 型 CRISPR-Cas 系统分子机制的理解，并为在临床和工业应用中开发针对细菌中整合 MGE 的工程 CRISPR 免疫提供了新的策略。
Staphylococcus aureus is a pathogen that can cause a wide range of infections in humans. Studies have suggested that CRISPR-Cas systems can drive the loss of integrated mobile genetic elements (MGEs) by chromosomal targeting. Here we demonstrate that CRISPR-mediated cleavage contributes to the partial deletion of integrated SCCmec in methicillin-resistant S. aureus (MRSA), which provides a strategy for the treatment of MRSA infections. The spacer within artificial CRISPR arrays should contain more than 25 nucleotides for immunity, and consecutive trinucleotide pairings between a selected target and the 5′ tag of crRNA can block targeting. These findings add to our understanding of the molecular mechanisms of the type III-A CRISPR-Cas system and provide a novel strategy for the exploitation of engineered CRISPR immunity against integrated MGEs in bacteria for clinical and industrial applications. CRISPR-Cas (clustered regularly interspaced short palindromic repeat [CRISPR]-CRISPR-associated protein [Cas]) systems can provide protection against invading genetic elements by using CRISPR RNAs (crRNAs) as a guide to locate and degrade the target DNA. CRISPR-Cas systems have been classified into two classes and five types according to the content of cas genes. Previous studies have indicated that CRISPR-Cas systems can avoid viral infection and block plasmid transfer. Here we show that chromosomal targeting by the Staphylococcus aureus type III-A CRISPR-Cas system can drive large-scale genome deletion and alteration within integrated staphylococcal cassette chromosome mec (SCCmec). The targeting activity of the CRISPR-Cas system is associated with the complementarity between crRNAs and protospacers, and 10- to 13-nucleotide truncations of spacers partially block CRISPR attack and more than 13-nucleotide truncation can fully abolish targeting, suggesting that a minimal length is required to license cleavage. Avoiding base pairings in the upstream region of protospacers is also necessary for CRISPR targeting. Successive trinucleotide complementarity between the 5′ tag of crRNAs and protospacers can disrupt targeting. Our findings reveal that type III-A CRISPR-Cas systems can modulate bacterial genome stability and may serve as a high-efficiency tool for deleting resistance or virulence genes in bacteria. IMPORTANCE Staphylococcus aureus is a pathogen that can cause a wide range of infections in humans. Studies have suggested that CRISPR-Cas systems can drive the loss of integrated mobile genetic elements (MGEs) by chromosomal targeting. Here we demonstrate that CRISPR-mediated cleavage contributes to the partial deletion of integrated SCCmec in methicillin-resistant S. aureus (MRSA), which provides a strategy for the treatment of MRSA infections. The spacer within artificial CRISPR arrays should contain more than 25 nucleotides for immunity, and consecutive trinucleotide pairings between a selected target and the 5′ tag of crRNA can block targeting. These findings add to our understanding of the molecular mechanisms of the type III-A CRISPR-Cas system and provide a novel strategy for the exploitation of engineered CRISPR immunity against integrated MGEs in bacteria for clinical and industrial applications.