The stb operon balances the requirements for vegetative stability and conjugative transfer of plasmid R388.
The stb operon balances the requirements for vegetative stability and conjugative transfer of plasmid R388.
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DOI：
10.1371/journal.pgen.1002073
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发表时间：
2011-05
期刊：
影响因子：
4.5
通讯作者：
de la Cruz F
中科院分区：
文献类型：
--
作者：
Guynet C;Cuevas A;Moncalián G;de la Cruz F
接合质粒R388和许多其他质粒携带操纵子stbABC，邻近接合转移的起点。我们研究了stbA、stbB和stbC基因的作用。stbA的缺失影响缀合和稳定性。它导致R388转移频率增加50倍，以及高质粒损失。与此相反，删除stbB废除接合，但没有引起质粒稳定性的变化。stbC的缺失没有显示出任何效果，无论是在缀合还是在稳定性上。整个stb操纵子的缺失对接合没有影响，其保持与野生型质粒相同，但导致与R388ΔstbA突变体相似的质粒丢失表型。我们得出结论，StbA是质粒稳定性所需的，StbA和StbB控制接合。我们接下来观察了R388 DNA分子的细胞内定位，并表明它们定位为均匀分布在活大肠杆菌细胞中的离散病灶。R388ΔΔstbA突变体的质粒不稳定性与质粒DNA分子簇的异常定位相关，无论是在一个细胞极，在两极，还是在细胞中心。相反，R388ΔΔstbB突变体中的质粒分子大多被排除在细胞极之外。因此，结果表明，质粒维持和转移中的缺陷是质粒DNA的细胞内定位变化的结果。我们建议，StbA和StbB构成一个非典型的质粒稳定系统，调和两种模式的质粒R388生理：一个维护模式（复制和分离）和传播模式（接合）。这一新概念在质粒生理学中的后果将被讨论。细菌进化和适应新环境的能力通常是通过水平转移获得新基因的结果。质粒酶通过其通过接合转移DNA的能力在基因交换中具有优势作用，接合是一种在细菌之间运输DNA的过程。此外，质粒是自主的DNA分子，在营养细胞增殖过程中忠实地传递给细胞后代。在这项研究中，我们报告了一个系统，由两种蛋白质，StbA和StbB，这两种模式之间的平衡质粒R388的生理作用：一个维护模式（垂直传输）和传播模式（水平传输）。我们证明，StbA是必不可少的，以确保忠实的质粒拷贝到子细胞的分类。反过来，StbB是质粒R388充分定位以进行接合所必需的。这是第一次报告的系统，调和质粒分离和接合。此外，R388属于接合质粒的IncW家族，由于其特别广泛的宿主范围而特别令人感兴趣。我们表明，StbAB系统是保守的各种各样的接合质粒，主要是广泛的主机范围的质粒。因此，Stb系统可以构成一个有趣的治疗靶点，以防止适应性基因的传播。
The conjugative plasmid R388 and a number of other plasmids carry an operon, stbABC, adjacent to the origin of conjugative transfer. We investigated the role of the stbA, stbB, and stbC genes. Deletion of stbA affected both conjugation and stability. It led to a 50-fold increase in R388 transfer frequency, as well as to high plasmid loss. In contrast, deletion of stbB abolished conjugation but provoked no change in plasmid stability. Deletion of stbC showed no effect, neither in conjugation nor in stability. Deletion of the entire stb operon had no effect on conjugation, which remained as in the wild-type plasmid, but led to a plasmid loss phenotype similar to that of the R388ΔstbA mutant. We concluded that StbA is required for plasmid stability and that StbA and StbB control conjugation. We next observed the intracellular positioning of R388 DNA molecules and showed that they localize as discrete foci evenly distributed in live Escherichia coli cells. Plasmid instability of the R388ΔΔstbA mutant correlated with aberrant localization of the plasmid DNA molecules as clusters, either at one cell pole, at both poles, or at the cell center. In contrast, plasmid molecules in the R388ΔΔstbB mutant were mostly excluded from the cell poles. Thus, results indicate that defects in both plasmid maintenance and transfer are a consequence of variations in the intracellular positioning of plasmid DNA. We propose that StbA and StbB constitute an atypical plasmid stabilization system that reconciles two modes of plasmid R388 physiology: a maintenance mode (replication and segregation) and a propagation mode (conjugation). The consequences of this novel concept in plasmid physiology will be discussed. The ability of bacteria to evolve and adapt to new environments most often results from the acquisition of new genes by horizontal transfer. Plasmids have a preponderant role in gene exchanges through their ability to transfer DNA by conjugation, a process that transports DNA between bacteria. Besides, plasmids are autonomous DNA molecules that are faithfully transmitted to cell progeny during vegetative cell multiplication. In this study, we report a system composed of two proteins, StbA and StbB, which act to balance plasmid R388 physiology between two modes: a maintenance mode (vertical transmission) and a propagation mode (horizontal transmission). We demonstrate that StbA is essential to ensure faithful assortment of plasmid copies to daughter cells. In turn, StbB is required for plasmid R388 adequate localization for conjugation. This is the first report of a system which reconciles plasmid segregation and conjugation. Furthermore, R388 belongs to the IncW family of conjugative plasmids, which are of particular interest due to their exceptionally broad host range. We show that the StbAB system is conserved among a wide variety of conjugative plasmids, mainly broad host range plasmids. Thus, the Stb system could constitute an interesting therapeutic target to prevent the spread of adaptive genes.
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