Vitamin C (L-ascorbic acid) has important antioxidant and metabolic functions in both plants and animals, but humans, and a few other animal species, have lost the capacity to synthesize it. Plant-derived ascorbate is thus the major source of vitamin C in the human diet. Although the biosynthetic pathway of L-ascorbic acid in animals is well understood, the plant pathway has remained unknown-one of the few primary plant metabolic pathways for which this is the case. L-ascorbate is abundant in plants (found at concentrations of 1-5 mM in leaves and 25 mM in chloroplasts) and may have roles in photosynthesis and transmembrane electron transport. We found that D-mannose and L-galactose are efficient precursors for ascorbate synthesis and are interconverted by GDP-D-mannose-3,5-epimerase. We have identified an enzyme in pea and Arabidopsis thaliana, L-galactose dehydrogenase, that catalyses oxidation of L-galactose to L-galactono-1,4-lactone. We propose an ascorbate biosynthesis pathway involving GDP-D-mannose, GDP-L-galactose, L-galactose and L-galactono-1,4-lactone, and have synthesized ascorbate from GDP-D-mannose by way of these intermediates in vitro. The definition of this biosynthetic pathway should allow engineering of plants for increased ascorbate production, thus increasing their nutritional value and stress tolerance.

Centre d'études et d'expertise sur les risques, l'environnement, la mobilité et l'aménagement

Root hairs provide a model system for the study of cell polarity. We examined the possibility that one or more members of the distinct plant subfamily of RHO monomeric GTPases, termed Rop, may function as molecular switches regulating root hair growth. Specific Rops are known to control polar growth in pollen tubes. Overexpressing Rop2 (Rop2 OX) resulted in a strong root hair phenotype, whereas overexpressing Rop7 appeared to inhibit root hair tip growth. Overexpressing Rops from other phylogenetic subgroups of Rop did not give a root hair phenotype. We confirmed that Rop2 was expressed throughout hair development. Rop2 OX and constitutively active GTP-bound rop2 (CA-rop2) led to additional and misplaced hairs on the cell surface as well as longer hairs. Furthermore, CA-rop2 depolarized root hair tip growth, whereas Rop2 OX resulted in hairs with multiple tips. Dominant negative GDP-bound Rop2 reduced the number of hair-forming sites and led to shorter and wavy hairs. Green fluorescent protein-Rop2 localized to the future site of hair formation well before swelling formation and to the tip throughout hair development. We conclude that the Arabidopsis Rop2 GTPase acts as a positive regulatory switch in the earliest visible stage in hair development, swelling formation, and in tip growth.

No abstract

One of the first responses of plants to microbial attack is the production of extracellular superoxide surrounding infection sites. Here, we report that Magnaporthe grisea, the causal agent of rice blast disease, undergoes an oxidative burst of its own during plant infection, which is associated with its development of specialized infection structures called appressoria. Scavenging of these oxygen radicals significantly delayed the development of appressoria and altered their morphology. We targeted two superoxide-generating NADPH oxidaseencoding genes, Nox1 and Nox2, and demonstrated genetically, that each is independently required for pathogenicity of M. grisea. ⌬nox1 and ⌬nox2 mutants are incapable of causing plant disease because of an inability to bring about appressorium-mediated cuticle penetration. The initiation of rice blast disease therefore requires production of superoxide by the invading pathogen.O ne of the earliest manifestations of the plant defense response is the production of reactive oxygen species (ROS), including superoxide and its dismutation product, hydrogen peroxide (1) . These ROS can kill pathogens directly (2) but also strengthen plant cell walls through the oxidative cross-linking of cell wall structural proteins (3) and may function in the regulation of programmed cell death (4). Although numerous studies have documented the detection of plant-derived ROS (2, 3, 5-7), very little is known about the role of ROS generation in invading plant pathogenic microorganisms. However, the recent discovery of functional members of the superoxide-generating NADPH oxidase (Nox) family within filamentous fungi (8) has led to increased speculation regarding the possible role of ROS in pathogenic species.The most well characterized Nox remains that of the human phagocytic leukocyte, a multisubunit oxidase formed by the cytosolic regulatory components Rac, p67 phox , p47 phox , and p40 phox and the integral membrane protein flavocytochrome b 558 , composed of the catalytic subunit gp91 phox and p22 phox (9). In activated macrophages, Nox enzymes induce K ϩ influx, causing pH changes in the phagocytic vacuole, leading to the killing of pathogens through activation of neutral proteases (10). Mutations in the catalytic gp91 phox subunit result in chronic granulomatous disease, an immunological disorder in which macrophages are unable to prevent the spread of infection (11). Plants contain enzymes that are homologous to gp91 phox , designated respiratory burst oxidase homologues (Rboh) (12). Arabidopsis thaliana, for example, possesses 10 Rboh isoforms involved in a diverse range of plant processes. ROS generated by the A. thaliana RHD2/RBOHC regulate root hair growth through the activation of Ca 2ϩ channels (13), whereas RBOHD and RBOHF regulate stomatal closure, seed germination, and root elongation through abscisic acid signaling (14). Recent studies have shown that it is the activation of RBOHD and RBOHF that is responsible for ROS accumulation in several plant-microbe interactions (15). However, r...