Department of Microbiology, Institute of Plant Biology, University of Zurich, Zurich, Switzerland.
Environ Microbiol Rep. 2012 Oct;4(5):562-8. doi: 10.1111/j.1758-2229.2012.00372.x. Epub 2012 Aug 19.
Burkholderia species are highly resistant to heavy metals (HMs), yet their resistance mechanisms are largely unknown. In this study we screened 5000 mini-Tn5 transposon insertion mutants of Burkholderia cenocepacia H111 for loss of cadmium tolerance. Of the four genes identified three affected outer membrane biogenesis and integrity or DNA repair. The fourth gene, BCAE0587, encoded a P1-type ATPase belonging to the CadA family of HM exporters. CadA-deficient strains lost the ability to grow in the presence of cadmium, zinc and lead, whereas resistance to nickel, copper and cobalt was not affected. Expression studies using a transcriptional fusion of the cadA promoter to gfp confirmed this specificity, as induction was only observed in presence of cadmium, zinc and lead. The promoter activity was found to be highest at neutral pH with an activation threshold of 30 nM cadmium. Inoculation of the HM-hyperaccumulating plant Arabidopsis halleri with a RFP-marked derivative of B. cenocepacia H111 containing the PcadA -gfp fusion demonstrated the applicability of this biosensor for monitoring cadmium at the single cell level in a natural environment.
伯克霍尔德氏菌属物种对重金属（HMs）具有很强的抗性，但它们的抗性机制在很大程度上尚不清楚。在这项研究中，我们筛选了 5000 个伯克霍尔德氏菌属中肠杆菌 H111 的 mini-Tn5 转座子插入突变体，以寻找对镉耐受性丧失的突变体。在鉴定的四个基因中，有三个基因影响外膜生物发生和完整性或 DNA 修复。第四个基因 BCAE0587 编码属于 HM 外排 CadA 家族的 P1 型 ATP 酶。CadA 缺陷型菌株丧失了在镉、锌和铅存在下生长的能力，而对镍、铜和钴的抗性不受影响。使用 cadA 启动子与 gfp 的转录融合进行表达研究证实了这种特异性，因为只有在存在镉、锌和铅的情况下才观察到诱导。发现启动子活性在中性 pH 下最高，镉的激活阈值为 30 nM。用含有 PcadA-gfp 融合的 RFP 标记的 B. cenocepacia H111 的衍生物接种重金属超积累植物拟南芥，证明了这种生物传感器可用于在自然环境中监测单个细胞水平的镉。
