IBBTEC, Instituto de Biomedicina y Biotecnologia de Cantabria (CSIC-UC-SODERCAN), Facultad de Medicina, Universidad de Cantabria, Santander, Spain.
PLoS Genet. 2011 May;7(5):e1002073. doi: 10.1371/journal.pgen.1002073. Epub 2011 May 19.
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.
R388 结合质粒和许多其他质粒携带一个操纵子 stbABC，位于结合转移原点的附近。我们研究了 stbA、stbB 和 stbC 基因的作用。stbA 的缺失既影响结合又影响稳定性。它导致 R388 转移频率增加了 50 倍，同时也导致质粒大量丢失。相比之下，stbB 的缺失会阻断结合，但不会改变质粒的稳定性。stbC 的缺失没有影响，无论是在结合还是稳定性方面。stb 操纵子的整个缺失对结合没有影响，仍与野生型质粒相同，但导致类似于 R388ΔstbA 突变体的质粒丢失表型。我们得出结论，StbA 是质粒稳定性所必需的，StbA 和 StbB 控制结合。我们接下来观察了 R388 DNA 分子的细胞内定位，并表明它们作为离散焦点均匀分布在活大肠杆菌细胞中。R388ΔΔstbA 突变体的质粒不稳定性与质粒 DNA 分子作为簇的异常定位相关，要么在一个细胞极，要么在两个细胞极，要么在细胞中心。相比之下，R388ΔΔstbB 突变体中的质粒分子主要被排除在细胞极之外。因此，结果表明，质粒维持和转移的缺陷是质粒 DNA 细胞内定位变化的结果。我们提出 StbA 和 StbB 构成了一种非典型的质粒稳定化系统，它协调了质粒 R388 的两种生理模式：一种是维持模式（复制和分离），另一种是繁殖模式（结合）。我们将讨论这个质粒生理学新概念的后果。
