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An arsenate reductase (Car1) from the Bacteroidetes species Rufibacter tibetensis 1351T was isolated from the Tibetan Plateau. The strain exhibits resistance to arsenite [As(III)] and arsenate [As(V)] and reduces As(V) to As(III). Here we shed light on the mechanism of enzymatic reduction by Car1. AlphaFold2 structure prediction, active site energy minimization, and steady‐state kinetics of wild‐type and mutant enzymes give insight into the catalytic mechanism. Car1 is structurally related to calcineurin‐like metallophosphoesterases (MPPs). It functions as a binuclear metal hydrolase with limited phosphatase activity, particularly relying on the divalent metal Ni2+. As an As(V) reductase, it displays metal promiscuity and is coupled to the thioredoxin redox cycle, requiring the participation of two cysteine residues, Cys74 and Cys76. These findings suggest that Car1 evolved from a common ancestor of extant phosphatases by incorporating a redox function into an existing MPP catalytic site. Its proposed mechanism of arsenate reduction involves Cys74 initiating a nucleophilic attack on arsenate, leading to the formation of a covalent intermediate. Next, a nucleophilic attack of Cys76 leads to the release of As(III) and the formation of a surface‐exposed Cys74‐Cys76 disulfide, ready for reduction by thioredoxin. [ABSTRACT FROM AUTHOR]

Cadmium (Cd) is highly toxic to plants, but the targets and modes of toxicity remain unclear. We isolated a Cd-hypersensitive mutant of Arabidopsis thaliana , Cd-induced short root 2 (cdsr2), in the background of the phytochelatin synthase-defective mutant cad1-3. Both cdsr2 and cdsr2 cad1-3 displayed shorter roots and were more sensitive to Cd than their respective wild type. Using genomic resequencing and complementation, IAR4 was identified as the causal gene, which encodes a putative mitochondrial pyruvate dehydrogenase E1 α subunit. cdsr2 showed decreased pyruvate dehydrogenase activity and NADH content, but markedly increased concentrations of pyruvate and alanine in roots. Both Cd stress and IAR4 mutation decreased auxin level in the root tips, and the effect was additive. A higher growth temperature rescued the phenotypes in cdsr2. Exogenous alanine inhibited root growth and decreased auxin level in the wild type. Cadmium stress suppressed the expression of genes involved in auxin biosynthesis, hydrolysis of auxin-conjugates and auxin polar transport. Our results suggest that auxin homeostasis is a key target of Cd toxicity, which is aggravated by IAR4 mutation due to decreased pyruvate dehydrogenase activity. Decreased auxin level in cdsr2 is likely caused by increased auxin-alanine conjugation and decreased energy status in roots. [ABSTRACT FROM AUTHOR]

Summary Rice grains typically contain relatively high levels of toxic arsenic (As) but low levels of essential micronutrients. Biofortification of essential micronutrients while decreasing As accumulation in rice would benefit human nutrition and health. We generated transgenic rice expressing a gain‐of‐function mutant allele astol1 driven by the OsGPX1 promoter. astol1 encodes a plastid‐localized O‐acetylserine (thiol) lyase (OAS‐TL) with Ser189Asn substitution (OsASTOL1S189N), which enhances cysteine biosynthesis by forming an indissociable cysteine synthase complex with its partner serine acetyltransferase (SAT). The effects on growth, As tolerance, and nutrient and As accumulation in rice grain were evaluated in hydroponic, pot and field experiments. The expression of OsASTOL1S189N in pOsGPX1::astol1 transgenic lines enhanced SAT activity, sulphate uptake, biosynthesis of cysteine, glutathione, phytochelatins and nicotianamine, and enhanced tolerance to As. The expression of OsASTOL1S189N decreased As accumulation while increased the accumulation of multiple macronutrients (especially sulphur, nitrogen and potassium) and micronutrients (especially zinc and selenium) in rice grain in a pot experiment and two field experiments, and had little effect on plant growth and grain yield. Our study provides a new strategy to genetically engineer rice to biofortify multiple essential nutrients, reducing As accumulation in rice grain and enhancing As tolerance simultaneously. [ABSTRACT FROM AUTHOR]

Respiratory Burst Oxidase Homologues (RBOHs) are involved in plant growth, development, and stress adaptation. How OsRBOHs affect root hair formation and consequently nutrient acquisition and drought resistance in rice is not well understood. We knocked out six OsRBOH genes in rice that were expressed in roots and identified OsRBOHE as the only one affecting root hair formation. OsRBOHE was strongly expressed in the root epidermis, root hairs and tiller buds. OsRBOHE is localised at the plasma membrane. Knockout of OsRBOHE decreased reactive oxygen species generation in the root hairs and tiller buds, downregulated genes involved in cell wall biogenesis, and decreased root hair length and tillering by 90% and 30%, respectively. Knockout of OsRBOHE decreased phosphorus acquisition only in low available P soil under aerobic conditions, but not in high P soil or under flooded conditions when P was likely not limited by diffusion. Knockout of OsRBOHE markedly decreased drought resistance of rice plants through the effect on root hair formation and the associated rhizosheath. Taken together, OsRBOHE is crucial for root hair formation and tillering and consequently on drought resistance in rice. The contribution of root hairs to P acquisition in rice is limited to aerobic soil. [ABSTRACT FROM AUTHOR]

Cadmium (Cd) is a toxic metal that poses serious threats to human health. Rice is a major source of dietary Cd but how rice plants transport Cd to the grain is not fully understood. Here, we characterize the function of the ZIP (ZRT, IRT‐like protein) family protein, OsZIP2, in the root‐to‐shoot translocation of Cd and intervascular transfer of Cd in nodes. OsZIP2 is localized at the plasma membrane and exhibited Cd2+ transport activity when heterologously expressed in yeast. OsZIP2 is strongly expressed in xylem parenchyma cells in roots and in enlarged vascular bundles in nodes. Knockout of OsZIP2 significantly enhanced root‐to‐shoot translocation of Cd and alleviated the inhibition of root elongation by excess Cd stress; whereas overexpression of OsZIP2 decreased Cd translocation to shoots and resulted in Cd sensitivity. Knockout of OsZIP2 increased Cd allocation to the flag leaf but decreased Cd allocation to the panicle and grain. We further reveal that the variation of OsZIP2 expression level contributes to grain Cd concentration among rice germplasms. Our results demonstrate that OsZIP2 functions in root‐to‐shoot translocation of Cd in roots and intervascular transfer of Cd in nodes, which can be used for breeding low Cd rice varieties. [ABSTRACT FROM AUTHOR]

The biogeochemical cycling of arsenic (As) is often intertwined with iron (Fe) and sulfur (S) cycles, wherein Fe(III)- and sulfate-reducing bacteria (SRB) play a crucial role. Here, we isolated strain DS-1, a strictly anaerobic Fe(III)- and sulfate-reducing bacterium, from As-contaminated paddy soil. Using 16S rRNA gene sequence analysis, strain DS-1 was identified as a member of the genus Desulfovibrio. Strain DS-1 utilized energy derived from ferrihydrite reduction to support its cellular growth. Under anoxic sulfate-reducing conditions, the presence of strain DS-1 significantly increased As mobilization compared to sulfate-free conditions. Mechanistically, SRB-produced sulfide reacts with Fe(III) to form FeS, which disrupts Fe(III) minerals, thereby enhancing As release. These findings highlight the critical role of redox disequilibrium in As mobilization and suggest that SRB-produced sulfide may permeate to the rice rhizosphere, increasing As mobilization through Fe(III) reduction., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Methylated arsenicals, including highly toxic species, such as methylarsenite [MAs(III)], are pervasive in the environment. Certain microorganisms possess the ability to detoxify MAs(III) by ArsI-catalyzed demethylation. Here, we characterize a bifunctional enzyme encoded by the arsI gene from Acidovorax sp. ST3, which can detoxify MAs(III) through both the demethylation and oxidation pathways. Deletion of the 22 C-terminal amino acids of ArsI increased its demethylation activity while reducing the oxidation activity. Further deletion of 44 C-terminal residues enhanced the MAs(III) demethylation activity. ArsI has four vicinal cysteine pairs, with the first pair being necessary for MAs(III) demethylation, while at least one of the other three pairs contributes to MAs(III) oxidation. Molecular modeling and site-directed mutagenesis indicated that one of the C-terminal vicinal cysteine pairs is involved in modulating the switch between oxidase and demethylase activity. These findings underscore the critical role of the C-terminal region in modulating the enzymatic activities of ArsI, particularly in MAs(III) demethylation. This research reveals the structure-function relationship of the ArsI enzyme and advances our understanding of the MAs(III) metabolism in bacteria.