Lu Mingmei, Jiao Shuo, Gao Enting, Song Xiuyong, Li Zhefei, Hao Xiuli, Rensing Christopher, Wei Gehong
State Key Laboratory of Crop Stress Biology in Arid Areas, College of Life Sciences, Northwest A&F University, Yangling, Shaanxi, China.
Key Laboratory of Urban Environment and Health, Institute of Urban Environment, Chinese Academy of Sciences, Xiamen, China.
Appl Environ Microbiol. 2017 Sep 29;83(20). doi: 10.1128/AEM.01244-17. Print 2017 Oct 15.
The symbiosis of the highly metal-resistant CCNWSX0020 and has been considered an efficient tool for bioremediation of heavy metal-polluted soils. However, the metal resistance mechanisms of CCNWSX00200 have not been elucidated in detail. Here we employed a comparative transcriptome approach to analyze the defense mechanisms of CCNWSX00200 against Cu or Zn exposure. Six highly upregulated transcripts involved in Cu and Zn resistance were identified through deletion mutagenesis, including genes encoding a multicopper oxidase (CueO), an outer membrane protein (Omp), sulfite oxidoreductases (YedYZ), and three hypothetical proteins (a CusA-like protein, a FixH-like protein, and an unknown protein), and the corresponding mutant strains showed various degrees of sensitivity to multiple metals. The Cu-sensitive mutant (Δ) and three mutants that were both Cu and Zn sensitive (Δ, Δ-like, and Δ-like) were selected for further study of the effects of these metal resistance determinants on bioremediation. The results showed that inoculation with the Δ mutant severely inhibited infection establishment and nodulation of under Cu stress, while inoculation with the Δ and Δ-like mutants decreased just the early infection frequency and nodulation under Cu and Zn stresses. In contrast, inoculation with the Δ-like mutant almost led to loss of the symbiotic capacity of to even grow in uncontaminated soil. Moreover, the antioxidant enzyme activity and metal accumulation in roots of inoculated with all mutants were lower than those with the wild-type strain. These results suggest that heavy metal resistance determinants may promote bioremediation by directly or indirectly influencing formation of the rhizobium-legume symbiosis. Rhizobium-legume symbiosis has been promoted as an appropriate tool for bioremediation of heavy metal-contaminated soils. Considering the plant-growth-promoting traits and survival advantage of metal-resistant rhizobia in contaminated environments, more heavy metal-resistant rhizobia and genetically manipulated strains were investigated. In view of the genetic diversity of metal resistance determinants in rhizobia, their effects on phytoremediation by the rhizobium-legume symbiosis must be different and depend on their specific assigned functions. Our work provides a better understanding of the mechanism of heavy metal resistance determinants involved in the rhizobium-legume symbiosis, and in further studies, genetically modified rhizobia harboring effective heavy metal resistance determinants may be engineered for the practical application of rhizobium-legume symbiosis for bioremediation in metal-contaminated soils.
具有高度金属抗性的CCNWSX0020的共生关系被认为是重金属污染土壤生物修复的有效工具。然而，CCNWSX00200的金属抗性机制尚未得到详细阐明。在此，我们采用比较转录组方法来分析CCNWSX00200对铜或锌暴露的防御机制。通过缺失诱变鉴定出6个与铜和锌抗性相关的高度上调转录本，包括编码多铜氧化酶（CueO）、外膜蛋白（Omp）、亚硫酸盐氧化还原酶（YedYZ）的基因，以及3个假定蛋白（一种类CusA蛋白、一种类FixH蛋白和一种未知蛋白），相应的突变菌株对多种金属表现出不同程度的敏感性。选择对铜敏感的突变体（Δ）和对铜和锌均敏感的3个突变体（Δ、Δ-like和Δ-like），进一步研究这些金属抗性决定因素对生物修复的影响。结果表明，在铜胁迫下接种Δ突变体严重抑制了根瘤菌的感染建立和结瘤，而接种Δ和Δ-like突变体仅降低了铜和锌胁迫下的早期感染频率和结瘤。相反，接种Δ-like突变体几乎导致根瘤菌在未受污染土壤中生长的共生能力丧失。此外，接种所有突变体的根瘤菌根中的抗氧化酶活性和金属积累均低于野生型菌株。这些结果表明，重金属抗性决定因素可能通过直接或间接影响根瘤菌 - 豆科植物共生体的形成来促进生物修复。根瘤菌 - 豆科植物共生体已被推广为重金属污染土壤生物修复的合适工具。考虑到金属抗性根瘤菌在污染环境中的植物生长促进特性和生存优势，对更多的金属抗性根瘤菌和基因工程菌株进行了研究。鉴于根瘤菌中金属抗性决定因素的遗传多样性，它们对根瘤菌 - 豆科植物共生体植物修复的影响必然不同，且取决于其特定的指定功能。我们的工作有助于更好地理解参与根瘤菌 - 豆科植物共生体的重金属抗性决定因素的机制，并且在进一步的研究中，可以构建携带有效重金属抗性决定因素的基因工程根瘤菌，用于根瘤菌 - 豆科植物共生体在金属污染土壤生物修复中的实际应用。
