Plant‐based FRET biosensor discriminates environmental zinc levels

Abstract

SummaryHeavy metal accumulation in the environment poses great risks to flora and fauna. However, monitoring sites prone to accumulation poses scale and economic challenges. In this study, we present and test a method for monitoring these sites using fluorescent resonance energy transfer (FRET) change in response to zinc (Zn) accumulation in plants as a proxy for environmental health. We modified a plant Zn transport protein by adding flanking fluorescent proteins (FPs) and deploying the construct into two different species. In Arabidopsis thaliana, FRET was monitored by a confocal microscope and had a 1.4-fold increase in intensity as the metal concentration increased. This led to a 16.7% overall error-rate when discriminating between a control (1 lM Zn) and high (10 mM Zn) treatment after 96 h. The second host plant (Populus tremula · Populu salba) also had greater FRET values (1.3-fold increase) when exposed to the higher concentration of Zn, while overall error-rates were greater at 22.4%. These results indicate that as plants accumulate Zn, protein conformational changes occur in response to Zn causing differing interaction between FPs. This results in greater FRET values when exposed to greater amounts of Zn and monitored with appropriate light sources and filters. We also demonstrate how this construct can be moved into different host plants effectively including one tree species. This chimeric protein potentially offers a method for monitoring large areas of land for Zn accumulation, is transferable among species, and could be modified to monitor other specific heavy metals that pose environmental risks.

Live Imaging of Inorganic Phosphate in Plants with Cellular and Subcellular Resolution

“…Sensors with a single fluorescent protein report ligand-dependent changes in conformation as changes in fluorescence intensity, whereas sensors with two fluorescent proteins can yield changes in fluorescence resonance energy transfer (FRET), which can be quantified through ratiometric imaging. FRET-based sensors have been used in live plants to assess a variety of analytes, including Glc, maltose, Suc, Gln, calcium, zinc, and pH (Deuschle et al, 2006;Chaudhuri et al, 2008Chaudhuri et al, , 2011Kaper et al, 2008;Rincón-Zachary et al, 2010;Adams et al, 2012;Gjetting et al, 2012Gjetting et al, , 2013Krebs et al, 2012). Gu et al (2006) engineered a FRET-based Pi sensor named fluorescence indicator protein for inorganic phosphate (FLIPPi) that consists of a cyanobacterial inorganic phosphate binding protein (PiBP) fused to enhanced cyan fluorescent protein (eCFP) and enhanced yellow fluorescent protein (eYFP) and showed the use of one of these sensors for monitoring cytosolic Pi in cultured animal cells.…”

Biological and Molecular Components for Genetically Engineering Biosensors in Plants

“…While GEPBs allow scientists to monitor molecular events, cell activities, and metabolic pathways in real-time, they also offer multiple desirable features for environmental monitoring. First, the integrated aboveground leaves and belowground root system allow plants to not only accept signals in the air, but to also sense the dynamic signals from a broad region of the soil [ 9 ]. Second, engineering biosensor in perennial plants species allows for cost-effective continuous monitoring of environmental conditions [ 9 ].…”

“…First, the integrated aboveground leaves and belowground root system allow plants to not only accept signals in the air, but to also sense the dynamic signals from a broad region of the soil [ 9 ]. Second, engineering biosensor in perennial plants species allows for cost-effective continuous monitoring of environmental conditions [ 9 ]. Third, with visible plant-based biosensors responsive to environmental factors, cumbersome sampling and laboratory analysis can be avoided [ 10 ].…”