Accumulation of Mercury in Transgenic Tobacco Expressing Bacterial Polyphosphate

Abstract

Mercury pollution is a worldwide problem in environments, resulting primarily from its industrial use in bleaching operations, as a catalyst, as a pigment in paints, and in the mining of gold. The effects of this heavy metal on the ecosystem and human health are growing concerns. Decontamination of mercury in the sediment and soil around industrial plants has been a challenge for a long time. Unfortunately, numerous sites contaminated with mercurials have not been remediated because of the high costs of dredging and landfilling.2) Phytoremediation, using plants to extract and/or sequester mercurials, is often regarded as a relatively inexpensive and efficient way of cleaning up the mercury-contaminated sites.Recently, with advances in biotechnology, phytoremediation utilizing specific plants to remove mercurials from contaminated sites has become one of the most rapidly developing fields for environmental restoration. Several studies have successfully integrated bacterial mer-genes into plant genome to create superior plants for phytoremediation of mercurial contaminated sites, based on the merA-mediated mercury reduction and volatilization mechanism.3-8) However, phytovolatilization of mercurials mediated by bacterial merA gene from the polluted sites would still cause public anxiety because of the release of mercury vapor into the ambient air that will redistribute in the ecosphere and be recycled to environment. To help solve this environmental problem, a new mercury-scavenging mechanism that can be expressed in plant cells and accumulate mercury from contaminated sites without releasing mercury vapor into the ambient air is required in place of the merA-mediated mercury reduction mechanism.In the previous papers, we reported that expression of ppkspecified polyphosphate (polyP) in a genetically engineered Escherichia coli strain resulted in significantly increased the bioaccumulation of Hg 2ϩ from simulated and actual wastewaters, and protected the bacterial cells from the toxic accumulated Hg 2ϩ . [9][10][11] In theory, plants engineering expression of polyP would also reduce mercury toxicity and accumulate more mercury from the polluted sites. As expected, we found that tobacco plant genetically engineered to express bacterial polyP was more resistant to Hg 2ϩ than its wild-type progenitors.12) The transported Hg 2ϩ seemed to be converted to a less toxic molecule, probably via chelation formation with ppk-specific polyP in the tobacco tissues just as the fact previously described in a bacterial system. 9,10) From these results we communicated that expression of ppk-specific polyP in transgenic tobacco may provide an ecologically compatible approach for environmental phytoremediation of mercury pollution in soils. However, the feasibility of ppk-specific polyP for phytoremediation of mercury had not uncovered. In this study, the ability of ppk-transgenic tobacco to accumulate mercury from the contaminated sites was investigated and the feasibility of this engineered tobacco for environmental remediation of ...

Genetic Engineering of Transgenic Tobacco for Enhanced Uptake and Bioaccumulation of Mercury

“…In agreement with our previous results, 9,10) the ppk-transgenic tobacco showed more resistance to Hg 2ϩ than its wild-type progenitors (Fig. 3).…”

“…20) Previously, we reported that tobacco plants expressing polyP in the absence of MerT, accumulated 2-6-fold more mercury than its wild-type progenitors from soils containing Hg 2ϩ concentrations over a range of 2.5-10 mM, but did not accumulate appreciable amounts of Hg 2ϩ compared with the wild-type control from soils containing Hg 2ϩ concentrations below 1.25 mM, despite its possession of polyP. 9,10) These previous observations together with the fact that mercury accumulation was accelerated by expression of MerT in tobacco plants inspired us to ascertain the efficiency and potential of ppk/merT-transgenic tobacco for use in the phytoremediation of low levels of mercury contamination, which is generally believed to be difficult to cleanup using natural or genetically-engineered plants without transport systems for the pollutant. Tobacco engineered to express mercury transporter and polyP accumulates effectively Hg 2ϩ at a range of low concentrations of Hg 2ϩ (0.1-1.25 mM) (Fig.…”

“…These results suggest that the incorporated mercury from medium was converted to a less toxic molecule, probably via chelation with ppk-specified polyP in the transgenic tobacco tissue as previously described. 21,22) These characteristics in addition to the ability to overexpress mercury chelator, polyP 9,10) shown by the ppk/ merT-transgenic tobacco may fulfill the proposed prerequisites for plants used for phytoremediation of mercury in soils.…”

“…[6][7][8] In previous papers, we reported that expression of polyphosphate (polyP) encoded by bacterial polyphosphate kinase gene (ppk) in a genetically engineered tobacco resulted in significantly increased accumulation of Hg 2ϩ from simulated soils without releasing mercury vapor into the ambient, and protected the tobacco from the toxicity of the accumulated Hg 2ϩ . 9,10) These phenotypes in addition to their high biomass production, the engineered tobacco should satisfy the proposed prerequisites for plants used in the phytoremediation of mercury in soils.…”

“…After 2 weeks incubation at 25°C, mercury tolerance was evaluated by monitoring the fresh weight according to the method previously described. 9,10) Mercury Uptake and Accumulation Assay The 4 weeks-old seedlings were hydroponically grown in fresh MS liquid medium containing 0.75 mM Hg 2ϩ . After incubation for appropriate times at 25°C, mercury taken up into the tobacco leaves was determined by flameless cold-vapor atomic adsorption spectrometry using an atomic mercury analyzer (RA-2A, Nippon Instruments, Tokyo, Japan) after the samples were digested with concentrated nitric acid.…”