Improving the nutritive value of tubers: elevation of cysteine and glutathione contents in the potato cultivar White Lady by marker-free transformation.

The amino acids that limit the nutritive value of potato are the sulfur containing amino acids methionine and cysteine. Manipulation of the targeted amino acid biosynthesis is a way to circumvent this problem. Cysteine is synthesised from O-acetyl-l-serine formed by serine acetyltransferase (SAT). To increase the cysteine content of the commercial potato cultivar White Lady the chimeric SAT-coding cysE gene from Escherichia coli under the control of the constitutive CaMV 35S promoter and fused to the chloroplast targeting rbcS 5'-transit peptide sequence was introduced into the White Lady genome. Novelty of the approach was the application of marker-free transformation. Two transgenic lines were obtained that accumulated the cysE mRNA in high amounts. Crude leaf extracts of these plants exhibited up to 80- and 20-fold higher SAT activity in leaves and tubers, respectively, than those prepared from non-transformed plants. Levels of cysteine and glutathione both in leaves and tubers were 1.5-fold higher in average than in control plants. The alterations observed had no effect on tuber yield and sprouting behaviour. Gas chromatography coupled to mass spectrometry showed that all other amino acids than cysteine were unaffected. Here we demonstrate for the first time that the cysteine content of tubers can be enhanced by metabolic engineering.

The Level of ALR is Regulated by the Quantity of Mitochondrial DNA.

Augmenter of liver regeneration (ALR) contributes to mitochondrial biogenesis, maintenance and to the physiological operation of mitochondria. The depletion of ALR has been widely studied and had serious consequences on the mitochondrial functions. However the inverse direction, the effect of the depletion of mitochondrial electron transfer chain and mtDNA on ALR expression has not been investigated yet. Thus mtDNA depleted, ρ(0) cell line was prepared to investigate the role of mitochondrial electron transfer chain and mtDNA on ALR expression. The depletion of mtDNA has not caused any difference at mRNA level, but at protein level the expression of ALR has been markedly increased. The regulatory role of ATP and ROS levels could be ruled out because the treatment of the parental cell line with different respiratory inhibitors and uncoupling agent could not provoke any changes in the protein level of ALR. The effect of mtDNA depletion on the protein level of ALR has been proved not to be liver specific, since the phenomenon could be observed in the case of two other, non-hepatic cell lines. It seems the level of mtDNA and/or its products may have regulatory role on the protein level of ALR. The up-regulation of ALR can be a part of the adaptive response in ρ(0) cells that preserves the structural integrity and the transmembrane potential despite the absence of protein components encoded by the mtDNA.

Physiological functions of presenilins; beyond γ-secretase.

Presenilin (PS) was identified in screens for mutations causing the early onset forms of familial Alzheimer's disease (FAD) in 1995. As catalytic units of the γ-secretase complex, presenilins participate in the processing of amyloid beta protein (Aß), the main component of deposits in brain of patients with AD. The more than 90 substrates of γ-secretase isolated so far demonstrate its contribution to wide range of cellular processes and signaling events. However, recent findings have revealed numerous γ-secretase-independent presenilin functions, including involvement in calcium homeostasis, endoplasmic reticulum (ER) stress and autophagy. This mini-review attempts to summarize the multiple physiological and pathological functions of presenilin.

Production of H2O2 in the endoplasmic reticulum promotes in vivo disulfide bond formation.

AIMS: Oxidative protein folding in the luminal compartment of endoplasmic reticulum (ER) is thought to be accompanied by the generation of H2O2, as side-product of disulfide bond formation. We aimed to examine the role of H2O2 produced in the lumen, which on one hand can lead to redox imbalance and hence can contribute to ER stress caused by overproduction of secretory proteins; on the other hand, as an excellent electron acceptor, H2O2 might serve as an additional pro-oxidant in physiological oxidative folding. RESULTS: Stimulation of H2O2 production in the hepatic ER resulted in a decrease in microsomal GSH and protein-thiol contents and in a redox shift of certain luminal oxidoreductases in mice. The oxidative effect, accompanied by moderate signs of ER stress and reversible dilation of ER cisternae, was prevented by concomitant reducing treatment. The imbalance also affected the redox state of pyridine nucleotides in the ER. Antibody producing cells artificially engineered with powerful luminal H2O2 eliminating system showed diminished secretion of mature antibody polymers, while incomplete antibody monomers/dimers were accumulated and/or secreted. INNOVATION: Evidence are provided by using in vivo models that hydrogen peroxide can promote disulfide bond formation in the ER. CONCLUSION: The results indicate that local H2O2 production promotes, while quenching of H2O2 impairs disulfide formation. The contribution of H2O2 to disulfide bond formation previously observed in vitro can be also shown in cellular and in vivo systems.

Effects of drought on water content and photosynthetic parameters in potato plants expressing the trehalose-6-phosphate synthase gene of Saccharomyces cerevisiae.

Two transgenic potato lines, T1 and T2, expressing the trehalose-6-phosphate synthase (TPS1) gene of yeast were isolated. In our experimental approach, we applied two novelties, namely the fusion of the drought-inducible promoter StDS2 to TPS1 and a marker-free transformation method. In contrast to the expected drought-induced expression, only a very low constitutive TPS1 expression was detected in the transgenic lines, probably due to chromosomal position effects. The observed expression pattern, however, was sufficient to alter the drought response of plants. Detached leaves of T1 and T2 showed an 8 h delay in wilting compared to the non-transformed control. Potted plants of T1 and T2 kept water 6 days longer than control plants and maintained high stomatal conductance and a satisfactory rate of net photosynthesis. During drought treatment, CO2 assimilation rate measured at saturating CO2 level was maintained at maximum level for 6-9 days in transgenic plants while it decreased rapidly after 3 days in the wild type plants. Under optimal growth conditions, lower CO2 fixation was detected in the transgenic than in the control plants. Stomatal densities of T1 and T2 leaves were reduced by 30-40%. This may have contributed to the lower CO2 fixation rate and altered drought response.

Prevalence and avidity of human herpesvirus-6 specific IgG antibodies in pregnant women in Hungary.

The prevalence, the level and the avidity of human herpesvirus-6 (HHV-6) specific IgG were examined in pregnant women and age-matched female blood donors. The study group consisted of 180 women (age 14-45); 60 women with normal pregnancy, 60 pregnant women with fetuses suspected of having any viral infection and 60 healthy blood donors with no history of pregnancy. Plasma or serum samples were tested for HHV-6 IgG antibodies by an immunofluorescence assay. Ninety-eight percent of blood donors and 97% of 120 pregnant women had IgG antibodies to HHV-6. The rate of seropositivity in women with normal pregnancies and women with fetuses suspected to have viral infection was the same. Pregnant women (n = 120) had significantly lower antibody titer than blood donors. No significant differences were found in the same respect between the two groups of pregnant women. Low avidity of IgG antibodies to HHV-6 was detected in 5% of pregnant women.