DIFFERENTIAL PROTEOMICS AND METABOLOMICS OF SHOOTS OF VETIVER GRASS (CHRYSOPOGON ZIZANIOIDES) USED TO TREAT ACID MINE DRAINAGE

Mining operations can produce acid mine drainage (AMD) that presents dangers to the environment through soil and water contamination. The abandoned Tab-Simco coal mining site in Southern Illinois generates AMD that is highly acidic with elevated sulfate and metal content. Floating treatment wetlands (FTWs) with vetiver grass (Chrysopogon zizanioides) have demonstrated the effectiveness of this non-invasive and fast-growing perennial grass for the remediation of Tab-Simco AMD at mesocosm and microcosm levels over extended periods. Here we report a proteomic investigation of vetiver shoots in response to AMD over shorter- and longer-term growth in concurrence with differential metabolism from a previous experiment. Proteomics revealed more upregulation after 56-days of AMD-growth (65% upregulated) while 7-day growth revealed more differential regulation (50% upregulated). There was significant upregulation of photosynthetic light-dependent proteins at 56 days and differential regulation of C₄ carbon fixation enzymes at 7 days. Amino acid and nitrogen metabolism were also impacted, including response to nitrogen starvation with the upregulation of ethylene and flavonoid biosynthesis. Moreover, there were increases in glutathione reductase and methionine sulfoxide reductase to manage oxidative stress and longer-term adaptations to ER-associated protein degradation and intracellular trafficking. Physical observation and chlorophyll content demonstrated negative impacts on photosynthesis after 7 days, though vetiver was able to acclimate and regrow by 56 days. Meanwhile, metabolomics suggested that vetiver shoots were accumulating larger amounts of oxaloacetate and ornithine for carbon and nitrogen storage. These findings largely correlated with proteomic changes, indicating that AMD-grown vetiver was better acclimated to AMD after longer-term exposure, likely suffering more from nutrient deprivation rather than oxidative stress from metals and acidity. These findings offer critical insight into the improvement of our FTWs in the treatment of AMD.