Pathological phenotypes and in vivo DNA cleavage by unrestrained activity of a phosphorothioate-based restriction system in Salmonella.
Pathological phenotypes and in vivo DNA cleavage by unrestrained activity of a phosphorothioate-based restriction system in Salmonella.
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DOI：
10.1111/mmi.12692
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发表时间：
2014-08
影响因子：
3.6
通讯作者：
You D
中科院分区：
文献类型：
--
作者：
Cao B;Cheng Q;Gu C;Yao F;DeMott MS;Zheng X;Deng Z;Dedon PC;You D
原核生物通过多种防御机制保护其基因组免受外来DNA的侵害，包括广泛的限制修饰（R-M）系统，涉及DNA骨架的硫代磷酸酯（PT）修饰。与经典的R-M系统不同，细菌基因组中共有基序的高度部分PT修饰表明PT依赖性限制的不寻常机制。在肠道沙门氏菌中，PT修饰由四个基因dptB-E介导，而限制涉及另外三个基因dptF-H。在这里，我们进行了一系列的研究来表征PT依赖的限制，并发现它呈现出一些不同于传统R-M系统的特征。PT缺陷突变体中限制性基因的存在不是致命的，而是导致几种病理表型。随后的转录谱分析显示，在缺乏PT的细胞中，超过600个基因的表达受到限制性酶的影响，包括噬菌体的诱导、SOS反应和DNA修复相关基因。这些转录反应与观察结果一致，即在体内没有PT修饰的情况下，限制性内切酶引起广泛的DNA切割。然而，尽管存在PT修饰，限制性基因的过表达对宿主是致命的。这些结果指出了一个不寻常的机制PT依赖性DNA切割的限制性内切酶在面对部分PT修改。
Prokaryotes protect their genomes from foreign DNA with a diversity of defense mechanisms, including a widespread restriction-modification (R-M) system involving phosphorothioate (PT) modification of the DNA backbone. Unlike classical R-M systems, highly partial PT-modification of consensus motifs in bacterial genomes suggests an unusual mechanism of PT-dependent restriction. In Salmonella enterica, PT modification is mediated by four genes dptB-E, while restriction involves additional three genes dptF-H. Here, we performed a series of studies to characterize the PT-dependent restriction, and found that it presented several features distinct with traditional R-M systems. The presence of restriction genes in a PT-deficient mutant was not lethal, but instead resulted in several pathological phenotypes. Subsequent transcriptional profiling revealed the expression of >600 genes was affected by restriction enzymes in cells lacking PT, including induction of bacteriophage, SOS response and DNA repair-related genes. These transcriptional responses are consistent with the observation that restriction enzymes caused extensive DNA cleavage in the absence of PT modifications in vivo. However, over-expression of restriction genes was lethal to the host in spite of the presence PT modifications. These results point to an unusual mechanism of PT-dependent DNA cleavage by restriction enzymes in the face of partial PT modification.
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