Higher levels of ectopic expression of Arabidopsis phytochelatin synthase do not lead to increased cadmium tolerance and accumulation

Abstract

Phytochelatins (PCs) are enzymatically synthesized peptides that play an important role in heavy metal homeostasis and detoxification. Recently, genes encoding PC synthase have been cloned from Arabidopsis and other species. Herein, we report on manipulating the expression of Arabidopsis thaliana PC synthase ( AtPCS1) in order to increase tolerance and accumulation of cadmium in plants. Approximately 50 transgenic Arabidopsis lines have been generated following transformation with a construct containing the AtPCS1 cDNA under the control of the cauliflower mosaic virus (CaMV) 35S promoter ( 35::AtPCS1). Following screening, two lines with the highest level of expression of the 35S::AtPCS1 transgene have been selected. When seedlings of these two lines, exhibiting ~14-fold increase in level of the AtPCS1 transcript, are subjected to 85 μM CdCl 2 for a 3-d period, ~30% increase in the level of PC production is observed compared to that of wild-type. However, these two transgenic lines are no more cadmium-tolerant, nor do they accumulate more cadmium than wild-type seedlings. In fact, these transgenic lines show higher sensitivity to cadmium than wild-type seedlings when the agar medium is supplemented with various concentrations of essential metals (EMs) in the presence of 75 μM CdCl 2. When all remaining transgenic lines are evaluated for cadmium tolerance, regardless of their levels of AtPCS1 transgene expression, six transgenic lines exhibit approximately twofold increase in cadmium tolerance compared to wild-type plants. Surprisingly, these six cadmium-tolerant lines show only slight increases (30–40%) in expression of AtPCS1 and ~15% increase in PC content when compared to wild-type. They also show increased accumulation of cadmium in their shoots. Therefore, critical levels of AtPCS1 are important in heavy metal detoxification.