In bacteria, copper (Cu) can support metabolic processes as an enzymatic cofactor but can also cause cell damage if present in excess, leading to intoxication. In group B Streptococcus (GBS) a system for control of Cu efflux based on the canonical cop operon supports survival during Cu stress. In some other bacteria, genetic systems additional to the cop operon are engaged during Cu stress and also contribute to Cu management. Here, we examined genetic systems beyond the cop operon in GBS for regions that contribute to survival of GBS in Cu stress using a forward genetic screen and probe of the entire bacterial genome. A high-density mutant library, generated using pGh9-ISS1, was used to expose GBS to Cu stress and compared to non-exposed controls en masse. Nine genes were identified as essential for GBS survival in Cu stress, whereas five genes constrained GBS growth in Cu stress. The genes encode varied factors including enzymes for metabolism, cell wall synthesis, transporters and global transcriptional regulators. Targeted mutation of the genes validated their roles in GBS resistance to Cu stress. Notably, several genes, including stp1, yceG, plyB and rfaB were also essential for resistance to Zn stress. Excepting copA, the genes identified are new to the area of bacterial metal ion intoxication. We conclude that a discrete and limited suite of genes beyond the cop operon in GBS contribute to a repertoire of mechanisms used to survive Cu stress in vitro and achieve cellular homeostasis.

中文翻译：

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B组链球菌的铜抵抗组揭示了对金属离子应激期间细胞存活的遗传基础的洞察

在细菌中，铜 (Cu) 作为酶辅助因子可以支持代谢过程，但如果过量存在，也会导致细胞损伤，导致中毒。在 B 组链球菌(GBS) 中，基于规范cop操纵子的用于控制 Cu 流出的系统支持在 Cu 胁迫期间的存活。在其他一些细菌中，除了cop操纵子之外的遗传系统在铜胁迫期间参与，也有助于铜管理。在这里，我们使用正向遗传筛选和整个细菌基因组的探针检查了 GBS 中除了cop操纵子之外的遗传系统，以确定有助于 GBS 在 Cu 胁迫中存活的区域。使用 pGh9-IS S1生成的高密度突变文库中，使用以暴露GBS与Cu应力和相对于未暴露对照集体。九个基因被确定为在铜胁迫下 GBS 存活所必需的，而五个基因限制了铜胁迫下 GBS 的生长。这些基因编码多种因子，包括代谢酶、细胞壁合成酶、转运蛋白和全局转录调节因子。基因的靶向突变验证了它们在 GBS 对铜胁迫的抗性中的作用。值得注意的是，包括stp1、yceG、plyB和rfaB在内的几个基因对于锌胁迫的抗性也是必不可少的。除了copA之外，所鉴定的基因在细菌金属离子中毒领域是新的。我们得出结论，除了cop之外的一组离散且有限的基因GBS 中的操纵子有助于在体外存活铜胁迫和实现细胞稳态的机制。