Plant growth and crop yields are very sensitive to toxic chemicals in their environment, including not only heavy metals and xenobiotics in the soil, but also oxygen in the air, which can react to produce destructive oxygen radicals. Glutathione (GSH) is a small molecule made from common amino acids that is key in protecting plants and animals from many toxic chemicals, and the GSH-ascorbate cycle detoxifies dangerous oxygen radicals. Creating plants that are more resistant to environmental stresses and therefore crops that maintain their yields under unfavorable conditions, requires detailed knowledge of how GSH levels are controlled, both through how it is made and how it is broken down. While a lot is known about GSH synthesis and regulation, very little is known about how GSH is broken down. This Arabidopsis 2010 research project investigates a newly discovered enzyme activity, termed gamma-glutamyl cyclotransferase, which experiments suggest catalyzes a majority of GSH break down in the model plant Arabidopsis thaliana. Experiments in this project will define the gene and protein that are responsible for this enzyme activity in plants. A combination of protein chemistry and genomics techniques will be employed to identify the gene in Arabidopsis. Subsequent genetic experiments will be used to verify that the isolated gene is responsible for GSH turnover and how GSH turnover is regulated relative to synthesis. Information gained will enable experiments designed to engineer or select plants with elevated potential for defending themselves from environmental threats. In addition, because GSH is found in all organisms, discovery of a new branch in the pathway of GSH metabolism is transformative with regard to understanding how all organisms survive in an oxygen containing environment.
Broader Impacts: To benefit the research community, data and information generated through this project will be made available through the Arabidopsis Information Resource (TAIR: www.arabidopsis.org). Seed stocks and any unique DNA materials will be made available through ABRC (www.biosci.ohio-state.edu/~plantbio/Facilities/abrc/abrchome.htm). To enhance education, the project will engage undergraduate students in research and professional development activities that are designed to expose them to advanced biotechnology, and enable them to continue careers in STEM disciplines. A postdoctoral research associate will be trained in research and mentoring, in preparation for a faculty career. In addition, undergraduates will work with the PI and the postdoctoral associate to develop a module on mitochondria to be included in the "Meta!Blast" virtual 3D cell, a web-based video environment designed to teach the basics of plant cell biology to precollege and beginning college students.
The chemical glutathione is produced by most organisms and plays an essential role in protecting those individuals from dangerous chemicals in their environment. Plants cannot survive without glutathione. It protects them from poisonous heavy metals in the soil, a broad range of naturally occurring and human-produced chemicals, and even the toxic byproducts of oxygen metabolism that accumulate when plants grow in less than optimal conditions. Because of its many essential roles, plants have evolved complex mechanisms for regulating the rate that glutathione is made. Many different stressful conditions cause plants to increase the rate of glutathione synthesis in order to protect the plant from those conditions. Despite all of the work done in understanding how glutathione is made, very little work has been done to determine how it is broken down. So while we understand its synthesis we know little about its degradation. Earlier we had discovered a family of genes that encode proteins called γ-glutamyltransferases. While these enzymes are responsible for glutathione breakdown in animals we were surprised to discover that they were not the major enzymes that broke glutathione down in plants. During this project we discovered that a second enzyme called γ-glutamyl cyclotransferase was responsible for glutathione breakdown in plants. We identified and characterized the enzyme and even identified the gene(s) that encode it. We have proven its role in glutathione synthesis by creating mutants that lack this activity. These plants will now allow us to completely understand glutathione synthesis and breakdown in plants. This knowledge should allow us to engineer plants with altered glutathione levels that may be more stress-resistance. Such crop plants should maintain higher yields under less than favorable growth conditions.
