Department of Biotechnology, Thapar Institute of Engineering and Technology, Patiala, Punjab, 147004, India.
Univ Lyon, Université Claude Bernard Lyon 1, CNRS, INRAE, VetAgro Sup, UMR Ecologie Microbienne, F-69622, Villeurbanne, France.
Environ Sci Pollut Res Int. 2021 Apr;28(14):17132-17145. doi: 10.1007/s11356-020-12152-6. Epub 2021 Jan 4.
Pollution in the environment due to accumulation of potentially toxic metals results in deterioration of soil and water quality, thus impacting health of all living organisms including microbes. In the present investigation, a functional metagenomics approach was adopted to mine functional genes involved in metal tolerance from potentially toxic metal contaminated site. Eukaryotic cDNA library (1.0-4.0 kb) was screened for the genes providing tolerance to cadmium (Cd) toxicity through a functional complementation assay using Cd-sensitive Saccharomyces cerevisiae mutant ycf1. Out of the 98 clones able to recover growth on Cd-supplemented selective medium, one clone designated as PLCc43 showed more tolerance to Cd along with some other clones. Sequence analysis revealed that cDNA PLCc43 encodes a 284 amino acid protein harbouring four characteristic zinc finger motif repeats (CXXCXGXG) and showing partial homology with heat shock protein (Hsp40) of Acanthamoeba castellanii. qPCR analysis revealed the induction of PLCc43 in the presence of Cd, which was further supported by accumulation of Cd in ycf1/PLCc43 mutant. Cu-sensitive (cup1), Zn-sensitive (zrc1) and Co-sensitive (cot1) yeast mutant strains were rescued from sensitivity when transformed with cDNA PLCc43 indicating its ability to confer tolerance to various potentially toxic metals. Oxidative stress tolerance potential of PLCc43 was also confirmed in the presence of HO. Present study results suggest that PLCc43 originating from a functional eukaryotic gene of soil community play an important role in detoxification of potentially toxic metals and may be used as biomarker in various contaminated sites.
由于潜在有毒金属的积累导致环境中的污染，从而导致土壤和水质恶化，从而影响包括微生物在内的所有生物的健康。在本研究中，采用功能宏基因组学方法从潜在有毒金属污染的地点挖掘参与金属耐受性的功能基因。通过使用 Cd 敏感的酿酒酵母突变体 ycf1 的功能互补测定筛选真核 cDNA 文库（1.0-4.0 kb），以寻找耐受 Cd 毒性的基因。在能够在补充有 Cd 的选择培养基上恢复生长的 98 个克隆中，有一个克隆被指定为 PLCc43，该克隆对 Cd 具有更高的耐受性，还有一些其他克隆也具有耐受性。序列分析表明，cDNA PLCc43 编码一个 284 个氨基酸的蛋白质，该蛋白质含有四个特征锌指模体重复（CXXCXGXG），并与嗜热变形菌的热休克蛋白（Hsp40）显示出部分同源性。qPCR 分析显示 PLCc43 在 Cd 存在下的诱导，这在 ycf1/PLCc43 突变体中 Cd 的积累进一步得到支持。当用 cDNA PLCc43 转化时，Cu 敏感（cup1），Zn 敏感（zrc1）和 Co 敏感（cot1）酵母突变体从敏感性中得到拯救，表明其能够赋予对各种潜在有毒金属的耐受性。在 HO 存在下，还证实了 PLCc43 的氧化应激耐受性潜力。本研究结果表明，源自土壤群落的功能真核基因的 PLCc43 在潜在有毒金属解毒中发挥重要作用，并且可以作为各种污染地点的生物标志物。
