Minisymposium 13: Heavy metals

Abs # 26004: Arsenic hyperaccumulation in transgenic Arabidopsis by suppressing an endogenous arsenate reductase (AtACR2)

Presenter:	Dhankher, Om Parkash , parkash@uga.edu
Authors	Dhankher, Om Parkash  (A)   Meagher, Richard B. (A)
Affiliations:	(A): Department of Genetics, The University of Georgia

Arsenic is an extremely toxic metalloid pollutant that adversely affects the health of millions of people worldwide. Recently, we engineered Arabidopsis plants co-expressing the E. coli arsenate reductase (ArsC) gene in leaves and the γ-ECS (γ-glutamylcysteine synthetase) gene constitutively. These plants showed significantly greater arsenic tolerance and accumulation than wild type plants or transgenic plants expressing either ArsC or γ-ECS alone (Dhankher et al., 2002, Nature Biotech. 20:1140-1145). XANES analysis of arsenic speciation in leaves and roots from wild type plants grown on arsenate showed that 95% of arsenic was in the reduced form arsenite. This suggests that plants have strong endogenous arsenate reductase activity that favors accumulation in As(III)-thiol-complexes. Arsenate is a phosphate analogue that appears to be recognized by phosphate uptake and transport systems. Arsenite is a thiol-reactive reagent that binds tightly to root tissues and is not translocated, which is a major limitation to our phytoremediation strategy. In order to mobilize and hyperaccumulate arsenic in the aboveground parts, we need to block the activity of endogenous plant arsenate reductase in roots and enhance its activity in leaves. We have identified a single gene AtACR2, in the Arabidopsis genome, with weak homology to the yeast arsenic reductase gene Acr2, and phosphatase such as superfamily of cdc25. The AtACR2 cDNA complements the E. coli ArsC gene function in E. coli, suggesting that AtACR2 functions as an arsenate reductase. Knockingdown AtACR2 expression, using RNA interference, results in Arabidopsis plants that are significantly sensitive to arsenate. Consistent with our model, the sensitivity of AtACR2-RNAi knockdown lines to arsenate suggests that blocking the reduction of arsenate to arsenite in roots causes the translocation and accumulation of more arsenate in shoots. These transgenic lines hyperaccumulated 10- to 14-fold more arsenic in aboveground tissues and retained 2- to 3-fold less arsenic in roots than wild type plants. The arsenic sensitive phenotypes and arsenic accumulation in RNAi lines are consistent with the reduction in expression of AtACR2 protein levels. AtACR2 encodes the first known plant arsenate reductase and has conserved homologs in many plant species. Altering the expression of the plant arsenate reductase gene should play a vital role in the phytoremediation of environmental arsenic contamination.