Detoxification of Multiple Heavy Metals by a Half-Molecule ABC Transporter,HMT-1, and Coelomocytes of Caenorhabditis elegans

Background

Developing methods for protecting organisms in metal-polluted environments is contingent upon our understanding of cellular detoxification mechanisms. In this regard, half-molecule ATP-binding cassette (ABC) transporters of the HMT-1 subfamily are required for cadmium (Cd) detoxification. HMTs have conserved structural architecture that distinguishes them from other ABC transporters and allows the identification of homologs in genomes of different species including humans. We recently discovered that HMT-1 from the simple, unicellular organism, Schizosaccharomyces pombe, SpHMT1, acts independently of phytochelatin synthase (PCS) and detoxifies Cd, but not other heavy metals. Whether HMTs from multicellular organisms confer tolerance only to Cd or also to other heavy metals is not known.

Methodology/Principal Findings

Using molecular genetics approaches and functional in vivo assays we showed that HMT-1 from a multicellular organism, Caenorhabditis elegans, functions distinctly from its S. pombe counterpart in that in addition to Cd it confers tolerance to arsenic (As) and copper (Cu) while acting independently of pcs-1. Further investigation of hmt-1 and pcs-1 revealed that these genes are expressed in different cell types, supporting the notion that hmt-1 and pcs-1 operate in distinct detoxification pathways. Interestingly, pcs-1 and hmt-1 are co-expressed in highly endocytic C. elegans cells with unknown function, the coelomocytes. By analyzing heavy metal and oxidative stress sensitivities of the coelomocyte-deficient C. elegans strain we discovered that coelomocytes are essential mainly for detoxification of heavy metals, but not of oxidative stress, a by-product of heavy metal toxicity.

Conclusions/Significance

We established that HMT-1 from the multicellular organism confers tolerance to multiple heavy metals and is expressed in liver-like cells, the coelomocytes, as well as head neurons and intestinal cells, which are cell types that are affected by heavy metal poisoning in humans. We also showed that coelomocytes are involved in detoxification of heavy metals. Therefore, the HMT-1-dependent detoxification pathway and coelomocytes of C. elegans emerge as novel models for studies of heavy metal-promoted diseases.     

Induction of oxidative stress and antioxidative mechanisms in Phaseolus vulgaris after Cd application.

Oxidative stress has been shown to be of great importance in the toxicity of several metals (copper, zinc, ...). In this study, the relationship of cadmium phytotoxicity and antioxidative reactions in bean (Phaseolus vulgaris L.) plants was investigated. Eleven-day-old seedlings were exposed to an environmentally realistic concentration of cadmium (2 microM CdSO(4)). Several biochemical and physiological parameters were influenced even by these low concentrations. At the biochemical level, the antioxidative defence mechanism was significantly activated after 24 h of cadmium exposure. Some enzymes able of quenching reactive oxygen species (syringaldazine peroxidase, EC 1.11.1.7; guaiacol peroxidase, EC 1.11.1.7) as well as enzymes important in the reduction of NAD(P)(+) (isocitrate dehydrogenase, EC 1.1.1.42; malic enzyme, EC 1.1.1.40) were significantly elevated by cadmium exposure. Furthermore, the ascorbate-glutathione cycle appeared to be a very important mechanism against cadmium-induced oxidative stress. In leaves, significant increases of ascorbate peroxidase (EC 1.11.1.11) and glutathione reductase (EC 1.6.4.2) and significant changes in the ascorbate and glutathione pool were observed. Morphological and other biochemical parameters (lipid peroxidation) were significantly enhanced 48 h after the start of the cadmium exposure. At the end of the experiment (72 h after the start of the metal treatment), even visual effects, such as chlorosis, were observed. The present data indicate that cadmium, like other metals, induces cellular redox disequilibrium suggesting that an environmentally realistic concentration of cadmium can cause oxidative stress.     

富营养化与污染物间的相互作用及其生态效应

1　引　言进入 90年代 ,为了改善水环境条件 ,越来越多的国家认识到必须减少养分、稳定的有机物和金属元素的输入。富营养化和持续污染物 (Ｃｈｒｏｎｉｃｐｏｌｌｕｔａｎｔｓ)是当前全球水生态系统的两个主要问题[1 ,2 ]。尽管对富营养化和污染物已有很多研究 ,但很少调查二者之间的相互关系。有证据表明两者之间可能存在相互作用过程[5～ 7],但在自然的水生态系统中其作用的大小和关系尚不清楚。滇池是我国典型的遭受富营养化和重金属污染双重胁迫的高原湖泊 ,本研究以滇池富营养化和重金属污染为背景 ,通过室内模拟 ,初步探索富营养化和…     

Effect of methamidophos on the growth rate and esterase activity of the common carp Cyprinus carpio L.

The acute toxicity of methamidophos and three other compounds to fingerlings of the common carp Cyprinus carpio L. was determined. The 96-h LC₅₀ values were 68 mg/litre for methamidophos, 1.7 mg/litre for carbaryl, 0.21 mg/litre for lindane and 50 mg/litre for diquat.The acetylcholinesterase (AChE) and carboxylesterase (CarE) activities of the brain and liver were monitored over six weeks in fish poisoned with sublethal doses of methamidophos. Brain CarE was more sensitive than AChE but for the liver the reverse was true. In either case the degree of enzyme inhibition increased with increasing insecticide concentrations in the water.After exposure to methamidophos at 20 mg/litre for 48h liver AChE and CarE recovered faster than those of the brain. For both organs CarE recovered faster than AChE.At sublethal doses methamidophos affected the growth rate of the fish but no direct relationship between growth and insecticide concentrations could be established.     

4种重金属离子对多刺裸腹溞的联合毒性效应

在研究了铜、铅、镉和锌4种单一因子对多刺裸腹溞（Moina macrocopa）急性毒性作用基础上,采用Marking 相加指数法研究了联合毒性效应.实验结果表明：4种重金属离子的毒性大小依次为Cd2＋＞Cu2＋＞Pb2＋＞Zn2＋,对多刺裸腹溞48h的LC50分别为0.0076mg/L,0.0306mg/L,0.0532mg/L,0.3568mg/L;Pb2＋-Cu2＋-Cd2＋、Pb2＋-Zn2＋-Cd2＋3种离子和Cu2＋-Pb2＋-cd2＋-Zn2＋4种离子共存时的毒性效应表现为拮抗作用;Cu2＋-Zn2＋-Cd2＋和Pb2＋-cu2＋-Zn2＋共存时的联合毒性较为复杂,三者浓度比1：1：1时表现为协同作用,而毒性比1∶1：1时在24h内表现为协同作用,至48h时为拮抗作用.     

Heavy metal-mediated activation of the rat Cu/Zn superoxide dismutase gene via a metal-responsive element

The Cu/Zn superoxide dismutase (SOD1) catalyzes the dismutation of superoxide radicals produced in the course of biological oxidations. When placed under the control of the rat SOD1 gene promoter and transfected into human HepG2 hepatoma cells, the activity of a chloramphenicol acetyltransferase reporter gene was found to increase three- to four-fold in the presence of heavy metals (cadmium, zinc and copper). Functional analysis of mutant derivatives of the SOD1 gene promoter and the use of a heterologous promoter system confirmed that the induction of the SOD1 gene by metal ions requires a metal-responsive element (MRE) located between positions −273 and −267 (GCGCGCA). It was also shown by gel mobility shift assays that an MRE binding protein is induced by the exposure of the human liver cell line HepG2 to heavy metals. These results suggest that the MRE participates in the induction of the SOD1 gene by heavy metals. Received: 5 February 1999 / Accepted: 21 May 1999     

生物淋滤技术在去除污泥中重金属的应用

利用微生物方法去除污泥中重金属（生物淋滤法）具有成本低,去除效率高,脱毒后污泥脱水性能好等优点,近年来在国际上备受关注,生物淋滤法采用的主要细菌为氧化亚铁硫杆菌（Thiobacillus ferrooxidans)和氧化硫硫杆菌（Thiobacillus thiooxidans),在其作用下,污泥中以难溶性金属硫化物被氧化成金属硫酸盐而溶出,通过固液分离即可达到去除重金属的目的,污泥的生物淋滤效果受温度、O2和CO2浓度,起始pH、污泥种类与浓度、底物种类与浓度、抑制因子、Fe^3 浓度等的影响,较为详细地介绍了生物淋滤法的作用机理及高效去除污泥中重金属的操作程序,并对其在环境污染治理方面的应用前景作了分析。     

Cumulative Title Index

Abstract

Metal fractionation is a powerful tool for studying the mobility, bioavailability and toxicity of metals in sediments and soils. A seven-step sequential extraction technique was used to determine the potential mobility of selected heavy metals (Fe, Mn, Zn, Cu, Pb, Cd and Ni) in the sediments of Lake Naivasha. Results indicate that residual fraction was the most important phase for the elements Fe, Mn, Cu and Zn. However, Pb and Cd are highly enriched in the non-residual phases. Nickel on the other hand was distributed evenly between the non-residual and the residual fractions.

The total concentrations of the heavy metals suggested a decreasing order of iron ?> manganese ? zinc > nickel > copper ? lead > cadmium. However, the detailed sequential extraction data indicated an order of release or mobility of cadmium > lead ? nickel ? zinc > manganese > copper > iron. The high percentage of Cd and Pb in the mobile fractions suggests high bioavailability of these two elements in the study area and maybe a pointer to anthropogenic input of the two elements in the study area.     

Effect of heavy metals on isoperoxidases of Wheat

The influence of increasing concentrations of copper, zinc, lead, nickel, chromium and cadmium on 14-day-old seedlings of wheat (Triticum aestivum L. cv. Vergina) was studied. Plants were grown in 1/10 strength Rorison’s nutrient solution with increasing concentrations of each of the metals added separately. The toxicity of metals depressed shoot growth but the most evident symptoms were on roots. The concentration of each metal which caused inhibition of root growth was chosen to study the influence of metals on isoperoxidases of wheat shoots. The concentrations employed did not alter the number of peroxidase bands but almost in all cases enhanced the intensities of bands of pH 4.0-4.2 and 5.0-5.4, while they decreased the intensities of bands of pH 4.2-4.6 and 5.4-6.5. The similar effects of the different heavy metals employed may suggest similarity in metal action on wheat isoperoxidases. The increased intensities of peroxidase bands may be considered as an indication of enhanced senescence caused by the heavy metal treatments. Generally, our results suggest that the heavy metals employed have caused complex changes on the multiple forms of peroxidases.     

Blockade of heavy metals accumulation in Chlorella vulgaris cells by 24-epibrassinolide

This study was conducted on the influence of 24-epibrassinolide (24-epiBL) mixed with varying concentrations of heavy metals (copper, lead, cadmium, zinc) upon the growth and accumulation of these heavy metals in the cell of the alga Chlorella vulgaris Beijerinck (Chlorophyceae). Heavy metals at the concentration of 10^–3 M, alone or mixed with 24-epiBL, showed a lethal effect on C. vulgaris. At metal concentrations of 10^–6–10^–4 M, a combination with 24-epiBL appeared to have a stronger stimulatory effect on a number of cells than a single metal (a stronger inhibitory effect). 24-EpiBL at the concentration of 10^–8 M in combination with heavy metals (in the range 10^–6–10^–4 M) blocked metal accumulation in algal cells. 24-EpiBL has an anti-stress effect on C. vulgaris contaminated by heavy metals. The inhibitory effect on metal accumulation of 24-epiBL mixed with different heavy metals was arranged in the following order: zinc > cadmium > lead > copper. This process is correlated with the stimulation of growth of C. vulgaris. The stimulatory effect of 24-epiBL mixed with heavy metals leading to an increased pH in the medium (5.28–6.20) was significantly higher than the impact due to the increased acidity in the medium due to metals alone (pH 3.10–5.85). Lower pH increased the toxicity of heavy metals in C. vulgaris cells.     

Distribution of extractable heavy metals in different soil fractions

Abstract

Due to the difficulties of precisely characterizing environmentally contaminated soil, the effects of heavy metals on plants are studied using uncontaminated soil spiked with known quantities of heavy metals. One problem in using spiked soils is how accurately the distribution of metals mimics stabilized natural soils. We studied the distribution of cadmium, chromium, copper, lead, nickel, and zinc in soil fractions after application in soluble form. The soil samples included a control (an uncontaminated Typic Argiudoll) and two samples spiked with either a moderate or high heavy metal concentration). After application of the salts the soils were subjected to wet/dry cycles over the course of three months. The soils were fractionated using a sequential chemical extraction procedure employing: (1) CaCl₂,(2) NaOH, (3) Na₂EDTA and (4) HNO₃, HCl, and HF. Soil physical separation was carried out by ultrasonic dispersion. The heavy metal levels were determined using ICP-AES. Each heavy metal displayed a unique behavior when added to soil in the form of soluble salts. Cadmium and zinc remained in the soluble fraction, indicating that no equilibrium was attained, while nickel primarily appeared in the insoluble fraction. Chromium, copper and lead were distributed among various soil chemical fractions. The highest levels of all metals appeared in the clay fraction except lead which was mainly present in the silt fraction.     

Concentrations of heavy metals in maternal and umbilical cord blood

Concentrations of lead, cadmium, methylmercury and total mercury were measured in maternal and umbilical cord blood using graphite atomic absorption spectrometry. Two essential metals, copper and zinc, were also determined using ion chromatography. Lead, copper and zinc were found to be lower in the cord blood, whereas methylmercury and total mercury were higher in cord blood than in maternal blood. Little differences were noted for cadmium in maternal and cord blood. Significant positive correlations were observed between the concentrations in maternal and cord blood with regard to lead (correlation coefficient, r = 0.44), copper (r = 0.34), zinc (r = 0.29), methylmercury (r = 0.44) and total mercury (r = 0.58). These results suggest that, like essential metals, most heavy metals can move rather freely across the human placenta. The potential health effects of heavy metal transfer from mothers to young infants cannot be discounted.     

Determination of zeta potential in Planctomycetes and its application in heavy metals toxicity assessment

Zeta potential of Planctomycetes was evaluated under different environmental conditions and correlated to cell viability. Phylogenetically distinct strains of the Planctomycetes presented different negative zeta potential values. More negative values were associated with Rhodopirellula spp. and related to the great amount of fimbriae in these species. Milli-Q water was chosen as the best dispersion media to perform the measurements. Zeta potential increased with ionic strength and varied with pH. In the physiological range of pH 5.0-9, zeta potential remained low and Rhodopirellula sp. strain LF2 cells were viable. Out of this range, zeta potential increased significantly and viability decreased. The effect on zeta potential of arsenic, cadmium, chromium, copper, lead, nickel, and zinc was assessed in Rhodopirellula sp. strain LF2. Zeta potential increased with increasing toxicity of the heavy metals in a dose-response way. This result was confirmed by the results observed for Rhodopirellula baltica strain SH1 under copper toxicity. Lead was the most toxic metal and zinc was the least toxic as observed by zeta potential and viability. The results support a correlation between zeta potential and cell viability which seem to indicate the possibility to use it as a viability predictor for the effects of heavy metals toxicity.     

Cleaning up of heavy metals-polluted water by a terrestrial hyperaccumulator Sedum alfredii Hance

Sedum alfredii Hance is a terrestrial zinc/cadmium （Zn/Cd）-hyperaccumulating and lead （Pb）-accumulating plant. Previous studies on S. alfredii were mostly focused on its physiological mechanism of heavy metal uptake and the application in phytoextraction of metals from contaminated soils. In this study, we evaluated the application potential of S. alfredii in the cleanup of heavy metals from contaminated lake water. Our research revealed that changing pH in lake water would not make particular difference on the final accumulation amount of heavy metals, because the acidic water environment negatively affected plant growth compared with the neutral and alkaline environments, but was more conducive for heavy metal absorption and accumulation. In addition, S. alfredii showed an increase of approximately 2.2-fold in dry weight （DW） when cultured with lake water for 25 d. At the same time, it accumulated approximately 5.0 mg/kg DW of Cd and 41.4 mg/kg DW of Pb. The absorption of heavy metals was highly effective during the first 10 d of culture. Also, the quality of lake water was greatly improved after only 2-d cleanup by S. aifredii. In general, this hyperaccumulator exhibits great potential for application in the cleanup of heavy metals-polluted waters.     

Survey of metal tolerance in moderately halophilic eubacteria.

The tolerance patterns, expressed as MICs, for 250 moderately halophilic eubacteria to 10 heavy metals were surveyed by using an agar dilution method. The moderate halophiles tested included 12 culture collection strains and fresh isolates representative of Deleya halophila (37 strains), Acinetobacter sp. (24 strains), Flavobacterium sp. (28 strains), and 149 moderately halophilic gram-positive cocci included in the genera Marinococcus, Sporosarcina, Micrococcus, and Staphylococcus. On the basis of the MICs, the collection strains showed, overall, similar responses to silver, cobalt, mercury, nickel, lead, and zinc. All were sensitive to silver, mercury, and zinc and tolerant of lead. The response to arsenate, cadmium, chromium, and copper was very heterogeneous. The metal susceptibility levels of the 238 freshly isolated strains were, in general, very heterogeneous among the four taxonomic groups as well as within the strains included in each group. The highest toxicities were found with mercury, silver, and zinc, while arsenate showed the lowest activity. All these strains were tolerant of nickel, lead, and chromium and sensitive to silver and mercury. Acinetobacter sp. strains were the most heavy-metal tolerant, with the majority of them showing tolerance of eight different metal ions. In contrast, Flavobacterium sp. strains were the most metal sensitive. The influence of salinity and yeast extract concentrations of the culture medium on the toxicity of the heavy metals tested for some representative strains was also studied. Lowering the salinity, in general, led to enhanced sensitivity to cadmium and, in some cases, to cobalt and copper. However, increasing the salinity resulted in only a slight decrease in the cadmium, copper, and nickel toxicities.(ABSTRACT TRUNCATED AT 250 WORDS)     

Survey of metal tolerance in moderately halophilic eubacteria

The tolerance patterns, expressed as MICs, for 250 moderately halophilic eubacteria to 10 heavy metals were surveyed by using an agar dilution method. The moderate halophiles tested included 12 culture collection strains and fresh isolates representative of Deleya halophila (37 strains), Acinetobacter sp. (24 strains), Flavobacterium sp. (28 strains), and 149 moderately halophilic gram-positive cocci included in the genera Marinococcus, Sporosarcina, Micrococcus, and Staphylococcus. On the basis of the MICs, the collection strains showed, overall, similar responses to silver, cobalt, mercury, nickel, lead, and zinc. All were sensitive to silver, mercury, and zinc and tolerant of lead. The response to arsenate, cadmium, chromium, and copper was very heterogeneous. The metal susceptibility levels of the 238 freshly isolated strains were, in general, very heterogeneous among the four taxonomic groups as well as within the strains included in each group. The highest toxicities were found with mercury, silver, and zinc, while arsenate showed the lowest activity. All these strains were tolerant of nickel, lead, and chromium and sensitive to silver and mercury. Acinetobacter sp. strains were the most heavy-metal tolerant, with the majority of them showing tolerance of eight different metal ions. In contrast, Flavobacterium sp. strains were the most metal sensitive. The influence of salinity and yeast extract concentrations of the culture medium on the toxicity of the heavy metals tested for some representative strains was also studied. Lowering the salinity, in general, led to enhanced sensitivity to cadmium and, in some cases, to cobalt and copper. However, increasing the salinity resulted in only a slight decrease in the cadmium, copper, and nickel toxicities.(ABSTRACT TRUNCATED AT 250 WORDS)     

Multigenerational Effects of Heavy Metals on Feeding,Growth, Initial Reproduction and Antioxidants in Caenorhabditis elegans

Earlier studies showed that toxicities of excessive metals lasted over generations. Yet, these studies mainly employed one-generation exposure, and the effects of multigenerational challenges need further studies. Presently, Caenorhabditis elegans were exposed to cadmium, copper, lead and zinc for four consecutive generations (G1 to G4) at environmental concentrations. The feeding, growth, initial reproduction, superoxide dismutase (SOD) and catalase (CAT) were determined. All data were represented in the percentage of that in control (POC), and POC in the control was normalized to 100%. In G1 and G2, the POC values in feeding, growth and initial reproduction were generally within 10% of the control (100%), indicating non-significant effects. The POC values in SOD and CAT were significantly higher than 100%, showing stimulatory effects. In G3 and G4, the POC values in feeding, growth and initial reproduction were significantly lower than 100%, showing inhibitory effects which were more severe in G4 than in G3. Meanwhile, SOD and CAT continuously showed stimulatory effects, and the stimulatory effects on SOD increased from G1 to G4. The effects with multigenerational challenges were different from those in one-generation exposure. The effects in later generations demonstrated the importance of multigenerational challenges in judging long-term influences of metals.     

Resistance to and uptake of heavy metals in mushrooms

Resistance levels to heavy metals in 21 mushrooms obtained mainly from Japan, but also from Thailand, were tested. The mushrooms were 7 Pleurotus species, 2 Pycnoporus and Pholiota species and one each of Agrocybe, Cryptoporus, Coriolus, Inonotus, Lampteromyces, Grifola, Flammulina, Lyophyllum, Agaricus, and Polyporus species. The Pleurotus species strains showed higher resistance to the heavy metals, copper, cadmium, zinc, nickel, cobalt, mercury than the other species. Pleurotus ostreatus exhibited the highest resistance to all these heavy metals. Pholiota species, Flammulina veltipes, Lyophyllum ulmarium, Agaricus bisporus and Polyporus arcularius were rather sensitive to all the metals tested. The accumulation of copper, zinc and cadmium in Pleurotus ostreatus was studied. The uptake of heavy metals into the mycelia of P. ostreatus increased proportionally to an increasing concentration of these metals in the medium.