In this article we will discuss about:- 1. Definition of Phytoremediation 2. Transgenic Plants for Phytoremediation 3. Transgenic Approach.
Definition of Phytoremediation:
Extensive activity of mankind has been introducing hazardous compounds into the environment at an alarming rate. The soil has been contaminated with toxic materials which is jeopardising its fertility. Heavy metals accumulation is primary concern because they cannot be destroyed by degradation. Heavy metals such as cadmium, copper, lead, mercury, arsenic zinc etc. contaminate urban soil at many sites throughout the world.
Accumulation of toxic heavy metals in soil and water is mainly due to mining, electroplating operations, industrial manufacturing practices, plastic industry, leaded gasoline and solid disposal site. In addition, military ammunitions are also a major worldwide source of soil heavy metal contamination.
Remediation of contaminated soil and ground water requires the removal of toxic metals from contaminated areas. Traditional methods of removing heavy metals from the contaminated soils and water are expensive and laborious. These systems are often threatened environment, e.g., contaminated sites can be clean up from the sand and placed in a sanitary landfill requires cost of approximately $ 2,000,000 per acre.
These methods are not only an expensive, but it may pose risk to spread contaminated soil. Therefore, several possible alternative strategies have been exploited to clean up contaminated soil. Among those, plants have been considered as favourable candidates in a novel process known as phytoremediation.
The potential use of plants and genetically modified plants to clean up contaminated soil and ground water has received great deal of attraction. Plants of one kind or another can be instrumental in the biological treatment of large number of contaminated soils. Accordingly, they may be exploited to remediate industrial pollution, waste water effluents etc.
Thus, utility of green plants to clean up contaminated hazardous waste site is known as phytoremediation. The use of aquatic plants in water quality assessment has been a common practice as biomonitor and is considered to be the development of water quality to protect aquatic life, the toxicity evaluation of municipal and industrial waste.
Phytoremediation is evolving as a cost-effective alternative to high cost conventional methods. It is being part of green revolution in the field of innovative clean up technology. The idea of using metal accumulating plants to remove heavy metal and other hazardous compounds was first introduced in 1983. Phytoremediation can be used as an insitu or exsitu application.
Insitu application is often encouraged because it minimizes distribution of soil and reduces spread of contaminants. Phytoremediation does not require any expensive instruments or skilled person. It is often evolved with selection of appropriate plant that can able to accumulate hazardous material in a wide range of environment. Green plants are able to survive in the hazardous contaminated soils.
Economy of phytoremediation has been assessed by number of agencies for example, the cost of cleaning up one acre of sand loam soil with a contaminate depth of 50 cm with plants was established at $ 60,000 to $ 100,000 compared to $ 400,000 for the conventional excavation and disposal method. Other common approaches used to treat metal- polluted soils are fixation, and teaching.
Transgenic Plants for Phytoremediation:
The overview on conventional method of phytoremediation particularly heavy metals discussed earlier concluded that the process could significantly reduce contamination over other traditional methods, and permanently accumulated or contaminate material disposed in landfills. However, the method has limitation because of plant’s feasibility for remediation due to toxicity of the metal to the plants.
In addition, method is restricted to shallow contamination sites and cannot guarantee of leaching of contamination into ground water. Besides, severe risk of bioaccumulation occurs of the plants enter the food chain in the ecosystem. Therefore, several laboratories devised transgenic plant a strategy for reducing some of the constraints of metal phytoremediation.
Several research teams have developed transgenic plants capable of tolerating high level of accumulated cadmium and lead.
These transgenic plants exhibits enhanced capacity for:
(i) Uptake and sequestration of trace elements, particularly heavy metal like cadmium.
(ii) Uptake, assimilate and volatilization of selenium.
(iii) Uptake at detoxification of mercury.
Many of the naturally occurring plant species used in phytoremediation process that can be genetically engineered, including Brassica juncea, Helianthus annus are more important tree species poplar. Genetic engineering strategy is to enhance the ability of metal ions to enter plant cells.
Many plant genes are involved in metal uptake, translocation and sequestration and transfer of any of these genes into candidate plants is a possible strategy for genetic engineering of plants for novel phytoremediation process.
Transgenic plants can be developed which will be engineered to accumulate high concentrations of metals in harvested plants (Fig. 23.2). Over expression of desirable genes will lead to enhanced metal uptake, translocation, sequestration or intracellular targeting.
For the production of transgenic plants for phytoremediation, genes can be transferred from hyper-accumulators or from other sources. Following are some of the possible areas of genetic engineering process for phytoremediation. In addition, bioinformatics analysis of metal transporter gene sequence data has greatly advanced rational engineering of strategy.
Metallothioneins, Phytochelatins and Metal Chelators:
Transgenic plants have been produced by successful expression of metallothionein genes. Transfer of human MT-2 genes in tobacco resulted in plants with increased Cd tolerance and pea MT genes in Arabidopsis thaliana enhanced Cu accumulation.
Selection of promoter was found to of great significance for metallothionein genes. The ribulose biphosphate carboxylase (rbcs) promoter was repressed by high Cd concentration while mannose synthase promoter was induced by Cd.
Transgenic plants with increased phytochelatin levels through over expression of cystein synthase resulted in enhanced Cd tolerance. High tolerance towards cadmium was noticed when yeast CUPI gene transformed to cauliflower. Various MT genes like mouse MTIA, human MTIA, Chinese hamster MT II, have been cloned and introduced into tobacco and Brassica plants, resulting in constitutively enhanced Cd tolerance in these plants.
Genetically transformed Brassica juncea plant overexpressing different enzymes involved in phytochelatin synthesis were shown to extract more Cd. Cr, Cu, Pb and Zn than wild plants. Transgenic Indian mustard with higher levels of glutathione and phytochelatins were developed by overexpression of two enzymes glutanylcysteine synthase (r-ECS) or glutathione synthase (GS) and they showed enhanced Cd tolerance and accumulation (Table 23.3).
Transfer of nicotinamine amino-transferase genes resulted in overproduction of iron chelator-deoxymugineic acid in rice. Similarly, expression of citrate synthase gene resulted in plants with enhanced Al tolerance. These plants produced upto 10-fold citrate in their roots and released 4-fold more compared to control plants.
Metal Transporters:
Metal tolerance can be altered in plants by genetic manipulation of metal transporters. Transfer of Zn transporter. ZAT gene from Thlaspi, goisingense to A. thaliane resulted in 2 fold higher Zn accumulation in roots. Transgenic tobacco showed enhanced accumulation of Ca, Cd and Mn after transfer of calcium vacuolar transporter CAX-2 from A. thaliania engineering.
Engineering of Metabolic Pathways:
Transgenic plants exhibits hyper-accumulation property can be produced by the introduction of new metabolic pathways as in case of MerA and MerB genes which were introduced into plants, which resulted in increased tolerance to mercury (Hg) and volatilized elemental mercury. The transgenic Arabidopsis that transport oxy anion arsenate to above ground, reduce arsenite and sequester is to thiol peptide complexes by transfer of E. coli ars C and r-ECS genes.
Modification in Roots:
Plants modified with increased branched root systems with large surface area for efficient uptake of toxic metal are indispensable. It is possible to increase root biomass by inducing hairy roots in hyperaccumator plants through Agrobacterium rhizogenes infection. The hairy roots induced in some or the hyperaccumulators were shown to have high efficiency for rhizofiltration of radionuclides and heavy metals.
Enhanced Biomass Production:
Over production of hormones in hyper accumulator plants can alter their biomass yield. Biosynthetic pathways for phytohormone synthesis have been characterised and genes encoding enzymes have been cloned. Manipulation of hormone content and regulate their biosynthesis can play significant role in biomass production in hyperaccumulator plants. Enhanced production of gibberellin in transgenic trees was shown to promote growth and biomass production.
Phytoremediation by Transgenic Trees:
Tree biotechnology is becoming an interesting tool and can be effectively employed for the remediation of contaminated environment. Phytoremediation of trees have better advantage over using herbs or shrubs because trees have extensive root system to ensure an effective uptake of the pollutants from soil.
Tree species can take up additional responsibilities of several cycles of decontamination in soil. For example, engineering poplar tree can effectively detoxify heavy metals and pesticides. The detoxification process involves conjugation of contaminants with glutathione (GSA) by glutathionine S-transferase and subsequent excretion of these conjugate in the vacuoles.
Transgenic poplar trees over expressing bacterial gene for GSH led to an increased accumulation of cadmium. Transgenic trees have also been engineered by expressing phytochelatin synthase either alone or in combination with bacterial GSH for enhanced tolerance for metals.
Transgenic Approach to Improve Phytoremediation:
The greater research effort is directed at improving plants using genetic engineering for better phytoremediation. Improvement in this approach will have an important role to play in commercial phytoremediation within next 4 to 6 years.
Several conceivable approaches in the improvement process have been summarised and overviewed as follows:
i. Introducing genes for encoding transport protein such as IRTI. The genes encode for this protein that regulate uptake of iron.
ii. Efficient expression of MRP1 Mg-ATPase transporter.
iii. Over expression of phytochelating and metallothionins.
iv. Introduce genes to enhance metal transport into roots, from roots to other plant biomass.
v. Introduction of genes encoding efficient metal chelators.
vi. Introduce genes encoding key biodegradation enzymes like nitroreductases, dehydro- halogenases etc.
vii. Introduction of genes for the stimulation of rhizosphere microflora.
viii. Introduce genes for the enhancement of root depth and penetration, growth rates, biomass production rates.
Cellular targeting, especially in the vacuoles, is important since the heavy metal can be kept in a safe compartment without disturbing the cellular function. Hence, engineering vacuolar transporters, preferably in specific cell types, is a second generation approach for phytoremediation.
Greater studies have been made in the development of transgenic plants for phytoremediation but majority of genes have been transferred from other organisms to plants. Understanding hyper-accumulators will help in transfer of genes from hyper-accumulators to candidate plants.
