Introduction
With the growing world population and changing climate conditions, feeding the world in the future will pose a severe challenge (Dhankher and Foyer 2018; Ray et al . 2013). Global food, feed, and fuel requirements are putting tremendous stress on our limited land resources. To achieve food safety and security goals, utilizing degraded and contaminated land for agricultural production safely and sustainably will assist in feeding the future population. Heavy metals and metalloids are major contributors to soil contamination (Li et al . 2019). Metal(loid)s such as arsenic (As) can be present naturally in agricultural soil or due to anthropogenic activities like fertilizer and pesticide application, sewage waste, coal burning, and industrial pollution (Li et al . 2019). The presence of metal(loid)s in soil impacts the growth and development of food crops as these toxic elements compete with nutrient uptake, negatively impact photosynthesis, create reactive oxygen species (ROS), and damage crucial biochemical processes in plant cells (Farooq et al . 2016; Li et al. 2019). Besides severely impacting food production, toxic metals also affect food quality and safety. Food contaminated with toxic metals poses serious threats to the health of humans and livestock. Chronic exposure to As can cause skin, liver, and kidney cancer (Farooq et al . 2016; Jomova et al . 2011), mercury (Hg) exposure leads to nervous system damage (Yang et al . 2020), cadmium (Cd) can cause kidney diseases and cancers (Genchi et al . 2020), and lead (Pb) causes reduced cognitive skills and mental development issues (Mason, Harp, and Han 2014). In order to utilize degraded land resources, we need to develop strategies for the safe use of contaminated sites without compromising the quality and quantity of the produce grown on them. One such approach is utilizing genetic engineering to create plants that can withstand phytotoxicity while limiting the uptake and accumulation of toxic metals.
One of the significant toxic metalloids of global concern is As. Arsenic is a highly toxic, group I carcinogen, and unlike some heavy metals like iron, nickel, and cobalt, it is not required by plants (Farooq et al . 2016). Globally, As contamination is a major concern in ~108 countries, impacting more than 230 million people (Shaji et al . 2021). Arsenic toxicity also reduces plant growth and development by damaging the plant’s cellular and molecular functions (Meselhy et al . 2021; Verma et al . 2021). Plants can take up As in two primary forms – inorganic forms such as arsenite (AsIII) and arsenate (AsV), and organic forms such as methylated arsenic –monomethylarsenic acid (MMA) and dimethylarsenic acid (DMA) (Bali and Sidhu 2021; Farooq et al . 2016). Plants take up inorganic AsV through phosphate transporters and AsIII, MMA, and DMA via membrane-bound aquaporin transporters, like nodulin 26-like proteins (NIPs) and plasma membrane intrinsic proteins (PIPs) (Ali 2022; Farooqet al . 2016; Mosa et al . 2012). After entering plants, As impacts the plants’ physiology and biochemistry by hindering photosynthetic, respiratory, and other growth and development processes. To protect themselves from the damaging effects of As and other toxic metals, plants utilize their innate capability to detoxify toxic metal(loid)s efficiently via binding with glutathione (GSH) and GSH derivatives, phytochelatins (PCs). Plant cells maintain GSH homeostasis via the γ-glutamyl cycle (Paulose et al. , 2013; Noctor et al. , 2012; Emamverdian et al. , 2015; Hasanuzzaman et al. , 2017). Glutathione and its oligomer PCs can help sequester heavy metals in the vacuole and extracellular spaces and limit their uptake and transport, thereby preventing the plant’s cellular machinery from damaging toxic effects (Hasanuzzaman et al. 2017). Exogenous GSH application has improved plant tolerance to Hg, cesium (Cs), silver nanoparticles (AgNPs), and salinity in Arabidopsis, soybean, and poplar (Adams et al . 2020; Akram et al . 2017; Kim et al . 2017; Ma et al . 2020). Moreover, improving GSH synthesis via overexpression of the GSH1 gene showed increased tolerance to Cd and As in Arabidopsis (Guo et al . 2008) and enhanced tolerance to Pb in poplar (Samuilov et al . 2016) and better overall growth of poplar grown in soil contaminated with heavy metals (Ivanova et al . 2011). Overexpression of bacterial γ-ECS, a GSH1homolog, also leads to increased tolerance to As in Arabidopsis (Dhankher et al. 2002; Li et al. 2005). Similarly, overexpressing GSH recycling genes, GGCT2;1 (gamma-glutamyl cyclotransferases 2;1 ), improved tolerance to As in Arabidopsis which led to higher biomass, lower As accumulation, and better nutrient utilization as compared to wild-type plants (Paulose et al.2013). Here, we translate those Arabidopsis research findings of Pauloseet al. (2013) into Camelina sativa , a biofuel crop, to potentially develop plants that can provide economic returns while cultivated on soils contaminated with toxic metals.
The selection of Camelina as the crop to work with was based on its unique properties. Camillina sativa , commonly known as the gold of pleasure or false flax, is an oilseed crop of the Brassicaceae family and is considered a dedicated biofuel crop (Chhikara et al . 2018). Camelina has various agronomic advantages for production, including early maturity, low requirement for water and nutrients, adaptability to adverse environmental conditions, and resistance to common cruciferous pests (Abdullah et al . 2018; Kagale et al . 2014; Stamenković et al . 2021). Camelina seeds contain more than 30% oil which can be utilized to produce biodiesel. Based on its shorter life cycle (85-100 days), it can be included in double and relay cropping systems. Life cycle assessment studies of Camelina have shown its economic viability and sustainability for use as a biofuel (Bertiet al . 2017; Keske et al . 2013). However, Camelina is still an underutilized resource, and its potential as a biofuel crop could be increased by translating and applying beneficial research findings from other model plants (Sainger et al . 2017; Stamenković et al . 2021).
In congruence with this aim to improve Camelina, as a translational approach, we overexpressed the CsGGCT2;1 gene in Camelina and analyzed its effect on the plant’s ability to grow on arsenite-contaminated media. Our results showed that overexpression ofCsGGCT2;1 enabled Camelina plants to grow better than wild-type (WT) plants on media supplemented with AsIII. The CsGGCT2;1transgenic lines also had significantly higher shoot and root biomass, improved chlorophyll content, lower lipid peroxidation, and lower arsenic accumulation than WT plants.