Disruption ofptLPD1orptLPD2, Genes That Encode Isoforms of the Plastidial Lipoamide Dehydrogenase, Confers Arsenate Hypersensitivity in Arabidopsis    

Abstract

Arsenic is a ubiquitous environmental poison that inhibits root elongation and seed germination to a variable extent depending on the plant species. To understand the molecular mechanisms of arsenic resistance, a genetic screen was developed to isolate arsenate overly sensitive (aos) mutants from an activation-tagged Arabidopsis (Arabidopsis thaliana) population. Three aos mutants were isolated, and the phenotype of each was demonstrated to be due to an identical disruption of plastidial LIPOAMIDE DEHYDROGENASE1 (ptLPD1), a gene that encodes one of the two E3 isoforms found in the plastidial pyruvate dehydrogenase complex. In the presence of arsenate, ptlpd1-1 plants exhibited reduced root and shoot growth and enhanced anthocyanin accumulation compared with wild-type plants. The ptlpd1-1 plants accumulated the same amount of arsenic as wild-type plants, indicating that the aos phenotype was not due to increased arsenate in the tissues but to an increase in the innate sensitivity to the poison. Interestingly, a ptlpd1-4 knockdown allele produced a partial aos phenotype. Two loss-offunction alleles of ptLPD2 in Arabidopsis also caused elevated arsenate sensitivity, but the sensitivity was less pronounced than for the ptlpd1 mutants. Moreover, both the ptlpd1 and ptlpd2 mutants were more sensitive to arsenite than wild-type plants, and the LPD activity in isolated chloroplasts from wild-type plants was sensitive to arsenite but not arsenate. These findings show that the ptLPD isoforms are critical in vivo determinants of arsenite-mediated arsenic sensitivity in Arabidopsis and possible strategic targets for increasing arsenic tolerance.Arsenic (As) is a naturally occurring metalloid found in soil, water, and air, but anthropogenic activities, including smelting and fossil fuel combustion, have led to increased environmental exposure (Mandal and Suzuki, 2002). In the environment, As exists in both organic and inorganic forms. Arsenate [As(V)] is the principal inorganic form of As in aerobic soils, while arsenite [As(III)] is the main form found under anaerobic conditions (Marin et al., 1993; Hossner, 1995, 1996;Mandal and Suzuki, 2002;Masscheleyn et al., 2002).Both As(V) and As(III) are toxic to plants, inducing symptoms ranging from poor seed germination and inhibited root growth to death (Meharg and HartleyWhitaker, 2002;Lee et al., 2003;Ahsan et al., 2008;Smith et al., 2010). The modes of action of As(V) and As(III) differ, owing to their distinct chemical properties. As(V), with its structural similarity to phosphate, can compete with phosphate in oxidative phosphorylation, leading to the production of ADPAs(V) (Gresser, 1981). However, half-maximal stimulation of ADP-As(V) formation requires physiologically unlikely concentrations of approximately 0.8 mM As(V) (Moore et al., 1983). As(V) has been recently shown to enhance membrane fluidity, and thus membrane permeability, by binding and replacing phosphate or choline head groups (Tuan et al., 2008). The resulting damage to the membrane would disrupt the transp...

Reciprocal Regulation of Protein Synthesis and Carbon Metabolism for Thylakoid Membrane Biogenesis

“…Further indications for E2 subunits possessing an additional function besides being a cpPDC subunit are derived from A. thaliana T-DNA insertion lines. Assuming that all three cpPDC enzyme components are necessary for its functionality, it is surprising that T-DNA insertions in E2 genes lead to embryo lethality in the homozygous state, whereas the E3 subunit is dispensable, thereby indicating an essential function of E2 distinct from its role in the cpPDC [56]–[58]. Similarly, an extensive PCR-based search for a DLA2 insertion mutant in an indexed library of C. reinhardtii failed, suggesting that DLA2 also is essential in green algae (Bohne, Grossman, and Nickelsen, unpublished results).…”

The metabolic acclimation of Arabidopsis thaliana to arsenate is sensitized by the loss of mitochondrial LIPOAMIDE DEHYDROGENASE2, a key enzyme in oxidative metabolism

“…Seed sterilization and growth conditions were described previously (Chen et al . ). For liquid culture, surface‐sterilized seeds were stratified at 4 °C for 2 d before carefully floating them in jars on the surface of sterile Gamborg's B‐5 medium (Phytotechnology Laboratories, Shawnee Mission, KS, USA) supplemented with 2% (w/v) sucrose and 0.08% (w/v) bacteriological grade agar (Amresco, Solon, OH, USA), pH 5.8.…”

“…), caused increased sensitivity to both As(V) and As(III) compared with wild‐type (WT) plants (Chen et al . ). Plastidial LPD is the E3 subunit of the plastidial pyruvate decarboxylase complex (ptPDC) that is largely responsible for producing acetyl‐coenzyme A for fatty acid biosynthesis in the plastid (Dörmann ).…”

Arsenic Speciation in Phloem and Xylem Exudates of Castor Bean

“…As(III)-PC complexation in roots was found to result in reduced mobility for efflux and for long-distance transport, possibly because the complexes are stored in the vacuoles (Liu et al, 2010). Excess As(III) causes cellular toxicity by binding to the vicinal thiol groups of enzymes, such as the plastidial lipoamide dehydrogenase, which has been shown to be a sensitive target of As toxicity (Chen et al, 2010). The As hyperaccumulating Pteris species differ from nonhyperaccumulating plants because of enhanced As(V) uptake (Wang et al, 2002;Poynton et al, 2004), little As(III)-thiol complexation (Zhao et al, 2003;Raab et al, 2004), and efficient xylem loading of As(III) .…”