No abstract

The accumulation of high amounts of specific metabolites in transgenic plants increases tolerance to salt and water stress . This effect has been observed in tobacco after the transfer of genes or cDNAs encoding enzymes that lead to the biosynthesis of mannitol, Gly betaine, Pro, trehalose, or fructan (Tarczynski et al., 1993;Kishor et al., 1995;Nomura et al., 1995;Pilon-Smits et al., 1995;Holmstrom et al., 1996). Because many naturally salt-or drought-tolerant plants accumulate such compounds when stressed, these metabolites are considered to be compatible solutes that act by providing osmotic adjustment and by lowering the osmotic potential, i.e. increasing the capacity to retain water (LeRudelier and Boullard, 1983). Other mechanisms by which these metabolites might enhance resistance to stress are, however, still not well understood. Some of these metabolites could act, for example, as scavengers of hydroxyl radicals-extremely toxic, short-lived active oxygen species that have no known enzymatic systems for breakdown (Smirnoff and Cumbes, 1989;Smirnoff, 1993;Asada, 1994;Shen et al., 1997)--or their protective function might be in maintaining the hydration sphere of proteins under water-stress conditions (Galinski, 1993;Papageorgiou and Murata, 1995).We have previously shown that tobacco, transformed to accumulate mannitol, survives high salinity better when stressed at the beginning of the exponential growth phase (Tarczynski et al., 1993). However, the amount of mannitol accumulated was usually not greater than approximately 6 to 8 pmol g-' fresh weight, possibly because of the fact that the Escherickia coli mtlD gene product catalyzed an equilibrium reaction that could also result in a conversion of mannitol-6-P to Fru-6-P, owing to the equal affinity of MtlD for either mannitol-6-P or Fru-6-P (Teschner et al., 1990). The direction of the reaction is determined by a pH-dependent change in affinity for NAD/NADH. W e decided to generate tobacco plants with a capacity for higher polyol production by exploiting a pathway that is induced in the halophyte ice plant (Mesembryantkemum crystallinum) following salt stress (Vernon and Bohnert, 1992). This pathway shunts carbon from Glu-6-P to myoinositol and then to methylated inositols. Methylated inositols are formed by a specific O-methyltransferase, IMT1, which seems to be absent in tobacco. IMTl methylates myo-inositol to form D( +)-ononitol (1D-1-O-methyl-myoinositol). Our expectation was that D-ononitol, which is dependent on the concentration of myo-inositol, might accumulate and provide stress tolerance, but it also seemed possible that the activity of the methyltransferase would generate myo-inositol deficiency.Tobacco (Nicotiana tabacum L., cv SRl) plants, transformed with the IMTl cDNA from M. crystallinum producing the enzyme D-myo-inositol methyltransferase (Vernon et al., 1993), were phenotypically normal. We compare here the physiology of the transformed line, I5A, with untransformed tobacco cv SR1. When the 15A plants were salt stressed or water...

Nf-E2 related factor-2 (Nrf2) is a basic leucine zipper transcription factor that binds and activates the antioxidant response element (ARE) in the promoters of many antioxidant and detoxification genes. also induced phosphorylation of eukaryotic translation initiation factor (eIF) 4E and eIF2␣ within 30 and 10 min, respectively. Inhibiting eIF4E with small interfering siRNA or increasing eIF2␣ phosphorylation with salubrinal did not affect Nrf2 elevation by H 2 O 2 . Our data present a novel phenomenon of quick onset of the antioxidant/detoxification response via increased translation of Nrf2 by oxidants. The mechanism underlying such stress-induced de novo protein translation may involve multiple components of translational machinery.

We analyzed transgenic tobacco (Nicotiana tabacum L.) expressing Stpd1, a cDNA encoding sorbitol-6-phosphate dehydrogenase from apple, under the control of a cauliflower mosaic virus 35S promoter. In 125 independent transformants variable amounts of sorbitol ranging from 0.2 to 130 mol g ؊1 fresh weight were found. Plants that accumulated up to 2 to 3 mol g ؊1 fresh weight sorbitol were phenotypically normal, with successively slower growth as sorbitol amounts increased. Plants accumulating sorbitol at 3 to 5 mol g ؊1 fresh weight occasionally showed regions in which chlorophyll was partially lost, but at higher sorbitol amounts young leaves of all plants lost chlorophyll in irregular spots that developed into necrotic lesions. When sorbitol exceeded 15 to 20 mol g ؊1 fresh weight, plants were infertile, and at even higher sorbitol concentrations the primary regenerants were incapable of forming roots in culture or soil. In mature plants sorbitol amounts varied with age, leaf position, and growth conditions. The appearance of lesions was correlated with high sorbitol, glucose, fructose, and starch, and low myo-inositol. Supplementing myo-inositol in seedlings and young plants prevented lesion formation. Hyperaccumulation of sorbitol, which interferes with inositol biosynthesis, seems to lead to osmotic imbalance, possibly acting as a signal affecting carbohydrate allocation and transport.

Evidence is presented for the recent, horizontal transfer of a self-splicing, homing group II intron from a cyanobacteria to the chloroplast genome of Euglena myxocylindracea. The psbA gene of E.myxocylindracea was found to contain a single 2566 nt group II intron with a gene in domain 4 for a 575 amino acid maturase. The predicted secondary structure and tertiary interactions of the group II intron, as well as the derived maturase primary sequence, most closely resemble the homing intron of the cyanobacterium Calothrix and the rnl introns of Porphyra purpurea mitochondria, while being only distantly related to all other Euglena plastid introns and maturases. All main functional domains of the intron-encoded proteins of known homing introns are conserved, including reverse transcriptase domains 1-7, the zinc finger domain and domain X. The close relationship with cyanobacterial introns was confirmed by phylogenetic analysis. Both the full-length psbA intron and a Delta-maturase variant self-splice in vitro in two independent assays. The psbA intron is the first example of a self-splicing chloroplast group II intron from any organism. These results support the conclusion that the psbA intron is the result of a recent horizontal transfer into the E.myxocylindracea chloroplast genome from a cyanobacterial donor and should prompt a reconsideration of horizontal transfer mechanisms to account for the origin of other chloroplast genetic elements.