Reconstruction of mreB Expression in Staphylococcus aureus via a Collection of New Integrative Plasmids
Reconstruction of mreB Expression in Staphylococcus aureus via a Collection of New Integrative Plasmids
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DOI：
10.1128/aem.00759-14
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发表时间：
2014-04
影响因子：
4.4
通讯作者：
Ana Yepes;G. Koch;A. Waldvogel;Juan-Carlos García-Betancur;Daniel López
中科院分区：
文献类型：
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作者：
Ana Yepes;G. Koch;A. Waldvogel;Juan-Carlos García-Betancur;Daniel López
摘要蛋白质定位一直是单细胞生物的传统研究，其易于遗传操作，有利于分子表征。两种杆状细菌模型大肠杆菌和枯草芽孢杆菌已被突出地用于这一目的，并取代了其他细菌，其遗传操作的挑战使任何细胞生物学研究复杂化。在这些细菌中有球形致病菌金黄色葡萄球菌。在这份报告中，我们提出了一个新的分子工具箱，促进基因缺失的葡萄球菌在一个1步重组过程和额外的载体，促进插入不同的报告融合到新确定的中性位点的S。金黄色染色体报告基因的插入不会将任何抗生素抗性基因添加到所得菌株的染色体上，从而使它们适合于进一步的遗传操作。我们使用这个工具箱在S中重建mreB的表达。金黄色葡萄球菌，一种编码肌动蛋白样细胞骨架蛋白的基因，这种蛋白在球菌细胞中不存在，可能在物种形成过程中丢失。我们观察到在S.在金黄色葡萄球菌中，MreB以与膜相关的离散结构组织，导致细胞壁材料的不寻常的重新分布。MreB的产生也会引起细胞增大，但不会使葡萄球菌的形状恢复。我们提出了MreB与关键葡萄球菌细胞壁相关蛋白的相互作用。这一工作为S.金黄色葡萄球菌作为探索细胞微生物学各个方面的模型系统。
ABSTRACT Protein localization has been traditionally explored in unicellular organisms, whose ease of genetic manipulation facilitates molecular characterization. The two rod-shaped bacterial models Escherichia coli and Bacillus subtilis have been prominently used for this purpose and have displaced other bacteria whose challenges for genetic manipulation have complicated any study of cell biology. Among these bacteria is the spherical pathogenic bacterium Staphylococcus aureus. In this report, we present a new molecular toolbox that facilitates gene deletion in staphylococci in a 1-step recombination process and additional vectors that facilitate the insertion of diverse reporter fusions into newly identified neutral loci of the S. aureus chromosome. Insertion of the reporters does not add any antibiotic resistance genes to the chromosomes of the resultant strains, thereby making them amenable for further genetic manipulations. We used this toolbox to reconstitute the expression of mreB in S. aureus, a gene that encodes an actin-like cytoskeletal protein which is absent in coccal cells and is presumably lost during the course of speciation. We observed that in S. aureus, MreB is organized in discrete structures in association with the membrane, leading to an unusual redistribution of the cell wall material. The production of MreB also caused cell enlargement, but it did not revert staphylococcal shape. We present interactions of MreB with key staphylococcal cell wall-related proteins. This work facilitates the use S. aureus as a model system in exploring diverse aspects of cellular microbiology.