Mutant Plants to the Rescue!

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Bioremediation is a management tool that addresses environmental pollution. In bioremediation organisms remove or neutralize pollutants from a contaminated site. Often these organisms are already present at the site.  For example a group of scientists from Rutgers University discovered a strain of bacteria that can breathe either oxygen or uranium in the soil of an old ore mill in Rifle Colorado. Other times organisms adept at containing or converting the specific pollutant are introduced to the site. In such cases the use of native species is encouraged in order to maintain the integrity of the local ecosystem.  

One area of biotechnology research involves finding organisms that are optimally suited for the clean up of a certain chemical and understanding what adaptations enable this service. An example of such work can be found in a Science article that received lots of attention because of its explosive subject matter – the bioremediation of TNT by the plant species Arabidopsis. 

NEWS_10.13.15_Arabidopsis_Fig1Arabidopsis, or more specifically Arabidopsis thaliana, is one of the most common plant model organisms. One advantage of Arabidopsis’ popularity is that its genetic makeup has been well studied and documented. Additionally, there are many existing mutant Arabidopsis lines, allowing scientists to rapidly screen for useful adaptations.

The TNT study began with planting several different mutant lines of Arabidopsis thaliana in both uncontaminated and TNT treated soil and looking for individuals with enhanced TNT tolerance. This is exactly what the scientists found – a line of plants that had significantly more root growth in the TNT soil than the other lines but the same amount of growth in the uncontaminated soil.

The next step was to discover the genetic underpinning of this trait. The mutation turned out to be a single nucleotide deletion in a gene called MDHAR6. This was a surprise to the scientists who were expecting to find a new or over-expressed detoxifying enzyme. MDHAR genes code for the antioxidant enzyme monodehydroascorbate reductase which reduces oxidative stress in plants by helping to recycle vitamin C. How could a loss of function mutation in this gene allow plants to tolerate TNT? 

To investigate this question scientist expressed MDHAR6 in E. coli and then purified the resulting protein by affinity chromatography. Experimenting with this protein product they found that the enzyme caused a one electron reduction in TNT. This reaction used NADH and created reactive oxygen species (ROS) that at high levels can damage cell structures. 

Returning to the plants, the scientists measured oxidative stress in both mutated and wild-type plants planted in TNT and found higher levels of stress in the normal plants. Finally, they transformed the MDHA6 gene back into the mutated line. This resulted in restored TNT toxicity and confirmed that plants deficient in MDHAR6 are more tolerant to TNT.

Arabidopsis is too small to efficiently remove TNT from polluted sites. However, the MDHAR6 gene can be disrupted or “knocked out” in other plants using traditional breeding methods. This is good news for thousands of sites that have been contaminated with residual TNT following explosions. It’s also possible that this strategy might lead to novel bioremediation strategies for other contaminants. 

To explore bioremediation in your classroom, we recommend starting with the “Bioremediation by Oil Eating Bacteria” Kit.  You can also test the effects of environmental pollution on plants with our “The Dose Makes the Poison” kit