Patterns of evolutionary rate variation among genes of the anthocyanin biosynthetic pathway.
The anthocyanin biosynthetic pathway is responsible for the production of anthocyanin pigments in plant tissues and shares a number of enzymes with other biochemical pathways. The six core structural genes of this pathway have been cloned and characterized in two taxonomically diverse plant species (maize and snapdragon). We have recently cloned these genes for a third species, the common morning glory, Ipomoea purpurea. This additional information provides an opportunity to examine patterns of evolution among genes within a single biochemical pathway. We report here that upstream genes in the anthocyanin pathway have evolved substantially more slowly than downstream genes and suggest that this difference in evolutionary rates may be explained by upstream genes being more constrained because they participate in several different biochemical pathways. In addition, regulatory genes associated with the anthocyanin pathway tend to evolve more rapidly than the structural genes they regulate, suggesting that adaptive evolution of flower color may be mediated more by regulatory than by structural genes. Finally, for individual anthocyanin genes, we found an absence of rate heterogeneity among three major angiosperm lineages. This rate constancy contrasts with an accelerated rate of evolution of three CHS-like genes in the Ipomoea lineage, indicating that these three genes have diverged without coordinated adjustment by other pathway genes. (+info)
Molecular cloning and biochemical characterization of a novel anthocyanin 5-O-glucosyltransferase by mRNA differential display for plant forms regarding anthocyanin.
UDP-glucose: anthocyanin 5-O-glucosyltransferase (5-GT) is responsible for the modification of anthocyanins to more stable molecules in complexes for co-pigmentation, supposedly resulting in a purple hue. The cDNA encoding 5-GT was isolated by a differential display applied to two different forms of anthocyanin production in Perilla frutescens var. crispa. Differential display was carried out for mRNA from the leaves of reddish-purple and green forms of P. frutescens, resulting in the isolation of five cDNA clones predominantly expressed in the red form. The cDNA encoded a polypeptide of 460 amino acids, exhibiting a low homology with the sequences of several glucosyltransferases including UDP-glucose: anthocyanidin 3-O-glucosyltransferase. By using this cDNA as the probe, we also isolated a homologous cDNA clone from a petal cDNA library of Verbena hybrida. To identify the biochemical function of the encoded proteins, these cDNAs were expressed in Saccharomyces cerevisiae cells. The recombinant proteins in the yeast extracts catalyzed the conversion of anthocyanidin 3-O-glucosides into the corresponding anthocyanidin 3,5-di-O-glucosides using UDP-glucose as a cofactor, indicating the identity of the cDNAs encoding 5-GT. Several biochemical properties (optimum pH, Km values, and sensitivity to inhibitors) were similar to those reported previously for 5-GTs. Southern blot analysis indicated the presence of two copies of 5-GT genes in the genome of both red and green forms of P. frutescens. The mRNA accumulation of the 5-GT gene was detected in the leaves of the red form but not in those of the green form and was induced by illumination of light, as observed for other structural genes for anthocyanin biosynthesis in P. frutescens. (+info)
Regulation of mitochondrial KATP channel by redox agents.
The ATP-dependent K+ channel (KATP) was purified from the inner mitochondrial membrane and reconstituted into lipid bilayer membranes. KATP activity was inhibited by high concentrations of ATP and ADP, but activated by low concentrations (up to 200 microM) of ADP. p-Diethylaminoethylbenzoate (DEB) acted as a KATP opener: at micromolar concentrations, it reversed inhibition by ATP and ADP and it also prevented KATP rundown. Pelargonidine, extracted from flowers of Pelargonium, reduced spontaneous activity of KATP channels and diminished their potentiation by DEB. Their opposite action on KATP corresponded with their opposite redox properties in reactions with free radicals: DEB behaved as an electron donor, whereas pelargonidine acted as an electron acceptor. We hypothesize that thiol groups on mitoKATP are targets for redox-active ligans. (+info)
Antimutagenicity of sweetpotato (Ipomoea batatas) roots.
Antimutagenicity of the water extracts prepared from the storage roots of four varieties of sweetpotato with different flesh colors was investigated using Salmonella typhimurium TA 98. The extract from the whole roots of the purple-colored Ayamurasaki variety effectively decreased the reverse mutation induced not only by Trp-P-1, Trp-P-2, IQ, B[a]P, and 4-NQO but also by dimethyl sulfoxide extracts of grilled beef. Comparison of the inhibitory activity of the extracts from the normal Ayamurasaki and its anthocyanin-deficient mutant one suggested that the anthocyanin pigment in the flesh decreases the mutagenic activity of the mutagens as heterocyclic amines. Two anthocyanin pigments purified from purple-colored sweet-potato, 3-(6,6'-caffeylferulylsophoroside)-5-glucoside of cyanidin (YGM-3) and peonidin (YGM-6) effectively inhibited the reverse mutation induced by heterocyclic amines, Trp-P-1, Trp-P-2, and IQ in the presence of rat liver microsomal activation systems. (+info)
Absorption and metabolism of cyanidin 3-O-beta-D-glucoside in rats.
We have clarified for the first time how cyanidin 3-O-beta-D-glucoside (C3G), which is a potent antioxidant anthocyanin, is absorbed and metabolized in vivo. Rats were orally administered C3G (0.9 mmol/kg body weight), and C3G rapidly appeared in the plasma. However, the aglycon of C3G (cyanidin; Cy) was not detected, although it was present in the jejunum. Protocatechuic acid (PC), which may be produced by degradation of Cy, was present in the plasma and the concentration was 8-fold higher than that of C3G. These results suggest that plasma PC and C3G may contribute to the antioxidant activity of the plasma. In the liver and kidney, C3G was metabolized to methylated C3G (methyl-C3G), suggesting that C3G and/or methyl-C3G act as antioxidants in the tissues. (+info)
Hyperoxia-induced changes in antioxidant capacity and the effect of dietary antioxidants.
We investigated, by measuring oxygen radical absorbance capacity (ORAC), whether hyperoxia causes alterations in antioxidant status and whether these alterations could be modulated by dietary antioxidants. Rats were fed for 8 wk a control diet or a control diet supplemented with vitamin E (500 IU/kg) or with aqueous extracts (ORAC: 1.36 mmol Trolox equivalents/kg) from blueberries or spinach and then were exposed to air or >99% O2 for 48 h. Although the constituents of the extracts were not extensively characterized, HPLC indicated that blueberry extract was particularly rich in anthocyanins, and the spinach extract did not contain any anthocyanins. The ORAC was determined in samples without proteins [serum treated with perchloric acid (PCA); ORACPCA] and with proteins (ORACtot). Hyperoxia induced a decrease in serum protein concentration, an increase in serum ORACPCA, decreases in lung ORACPCA and ORACtot, and an equilibration of proteins and ORACPCA between serum and pleural effusion. These alterations suggested a redistribution of antioxidants between tissues and an increase in capillary permeability during hyperoxia. Only the blueberry extract was effective in alleviating the hyperoxia-induced redistribution of antioxidants between tissues. (+info)
Protective effects of dietary nasunin on paraquat-induced oxidative stress in rats.
The preventive effects of nasunin (delphinidin-3-[4-p-coumaroyl-rhamnosyl(1-->6)glucosid e]-5-glucoside) on paraquat-induced oxidative stress were determined in rats. Decreased food intake and body weight gain and increased lung weight by feeding the rats a diet containing paraquat were clearly suppressed by supplementing nasunin to the paraquat diet. Paraquat feeding increased the concentration of thiobarbituric acid-reactive substances (TBARS) in liver lipids and the atherogenic index, and decreased the liver triacylglycerol level. These effects were also suppressed by supplementing nasunin to the paraquat diet. In addition, catalase activity in the liver mitochondrial fraction was markedly decreased by feeding the paraquat diet, this decrease being partially suppressed by supplementing the paraquat diet with nasunin. These results suggest that nasunin acted preventively against the oxidative stress in vivo that may have been due to active oxygen species formed through the action of paraquat. (+info)
ANTHOCYANINLESS2, a homeobox gene affecting anthocyanin distribution and root development in Arabidopsis.
The ANTHOCYANINLESS2 (ANL2) gene was isolated from Arabidopsis by using the maize Enhancer-Inhibitor transposon tagging system. Sequencing of the ANL2 gene showed that it encodes a homeodomain protein belonging to the HD-GLABRA2 group. As we report here, this homeobox gene is involved in the accumulation of anthocyanin and in root development. Histological observations of the anl2 mutant revealed that the accumulation of anthocyanin was greatly suppressed in subepidermal cells but only slightly reduced in epidermal cells. Furthermore, the primary roots of the anl2 mutant showed an aberrant cellular organization. We discuss a possible role of ANL2 in the accumulation of anthocyanin and cellular organization of the primary root. (+info)