Differential Effects of Carbohydrates on Arabidopsis Pollen Germination.

Pollen germination as a crucial process in plant development strongly depends on the accessibility of carbon as energy source. Carbohydrates, however, function not only as a primary energy source, but also as important signaling components. In a comprehensive study, we analyzed various aspects of the impact of 32 different sugars on in vitro germination of Arabidopsis pollen comprising about 150 variations of individual sugars and combinations. Twenty-six structurally different mono-, di- and oligosaccharides, and sugar analogs were initially tested for their ability to support pollen germination. Whereas several di- and oligosaccharides supported pollen germination, hexoses such as glucose, fructose and mannose did not support and even considerably inhibited pollen germination when added to germination-supporting medium. Complementary experiments using glucose analogs with varying functional features, the hexokinase inhibitor mannoheptulose and the glucose-insensitive hexokinase-deficient Arabidopsis mutant gin2-1 suggested that mannose- and glucose-mediated inhibition of sucrose-supported pollen germination depends partially on hexokinase signaling. The results suggest that, in addition to their role as energy source, sugars act as signaling molecules differentially regulating the complex process of pollen germination depending on their structural properties. Thus, a sugar-dependent multilayer regulation of Arabidopsis pollen germination is supported, which makes this approach a valuable experimental system for future studies addressing sugar sensing and signaling.

Homo- and heterodimers of tobacco bZIP proteins counteract as positive or negative regulators of transcription during pollen development.

Expression of BZI-1 Delta N, a dominant-negative form of the tobacco (Nicotiana tabacum) basic leucine zipper (bZIP) transcription factor BZI-1 leads to severe defects in pollen development which coincides with reduced transcript abundance of the stamen specific invertase gene NIN88 and decreased extracellular invertase enzymatic activity. This finding suggests a function of BZI-1 in regulating carbohydrate supply of the developing pollen. BZI-1 heterodimerises with the bZIP factors BZI-2, BZI-3 and BZI-4 in vitro and in planta. Whereas BZI-1 exhibits only weak activation properties, BZI-1/BZI-2 heterodimers strongly activate transcription. Consistently, approaches leading to reduced levels of functional BZI-1 or BZI-2 both significantly interfere with pollen development, auxin responsiveness and carbohydrate partitioning. In situ hybridisation studies for BZI-1 and BZI-2 confirmed temporal and spatial overlapping expression patterns in tapetum and pollen supporting functional cooperation of these factors during pollen development. Plants over-expressing BZI-4 produce significantly reduced amounts of intact pollen and are also impaired in NIN88 transcription and enzymatic activity. BZI-4 homodimer efficiently binds to a G-box located in the NIN88 promoter but exhibits almost no transcriptional activation capacity. As BZI-4 does not actively repress transcription, we propose that its homodimer blocks G-box mediated transcription. In summary, these data support a regulatory model in which BZI-4 homodimers and BZI-1/BZI-2 heterodimers perform opposing functions as negative or positive transcriptional regulators during pollen development.

Hovenia dulcis--an Asian traditional herb.

Hovenia dulcis Thunb., known as Japanese raisin tree, is commonly found in East Asia. It has a long history as a food supplement and traditional medicine in Japan, China and Korea, but is little known and used in Western countries so far. This minireview summarizes traditional uses and current knowledge on the pharmacology and phytochemistry of H. duclcis and covers, in particular, literature from specialized Asian journals that are not readily accessible. Extracts from H. dulcis accelerate detoxification of ethanol, and possess hepatoprotective, antioxidative, antimicrobial and antidiabetic properties. Although the underlying molecular mechanisms are not fully understood, free radical scavenging and enhancement of ethanol catabolism have been reported.

Cloning of a CACTA transposon-like insertion in intron I of tomato invertase Lin5 gene and identification of transposase-like sequences of Solanaceae species.

Very few CACTA transposon-like sequences have been described in Solanaceae species. Sequence information has been restricted to partial transposase (TPase)-like fragments, and no target gene of CACTA-like transposon insertion has been described in tomato to date. In this manuscript, we report on a CACTA transposon-like insertion in intron I of tomato (Lycopersicon esculentum) invertase gene Lin5 and TPase-like sequences of several Solanaceae species. Consensus primers deduced from the TPase region of the tomato CACTA transposon-like element allowed the amplification of similar sequences from various Solanaceae species of different subfamilies including Solaneae (Solanum tuberosum), Cestreae (Nicotiana tabacum) and Datureae (Datura stramonium). This demonstrates the ubiquitous presence of CACTA-like elements in Solanaceae genomes. The obtained partial sequences are highly conserved, and allow further detection and detailed analysis of CACTA-like transposons throughout Solanaceae species. CACTA-like transposon sequences make possible the evaluation of their use for genome analysis, functional studies of genes and the evolutionary relationships between plant species.

Circadian and developmental regulation of vacuolar invertase expression in petioles of sugar beet plants.

The expression pattern of the genes coding for vacuolar and extracellular invertase activity was analyzed in sugar beet (Beta vulgaris) and compared with the expression of sucrose synthase in this important sucrose-storing crop. Northern blot analysis revealed that sucrose synthase is the predominant sucrose-cleaving enzyme in tap roots, whereas vacuolar invertase was specifically expressed in petioles. Extracellular invertase transcripts showed low abundance in all the sugar beet organs and were not detected in northern blots. Relative RT-PCR analysis revealed differential expression of the two extracellular invertase genes: BVInv-CW1 was almost exclusively expressed in tap roots and BVInv-CW2 was widely expressed in all the organs analyzed. A remarkable result of this analysis was the high expression of vacuolar invertase (BVInv-V3) in petioles. Two factors had a clear influence on vacuolar invertase gene expression in petioles: light and the developmental stage, so that expression was higher in petioles from juvenile plants. BVInv-V3 transcripts showed circadian oscillation in petioles, with maximal accumulation during the light period. A similar pattern of diurnal oscillation was also observed for the vacuolar invertase activity, showing a delay with respect to the level of transcripts. The analysis of sugars in petioles revealed oscillation of the hexoses, with a remarkably higher content of glucose than fructose. In contrast, the level of sucrose in petioles was very low. This pattern of expression suggests an important role of petiole vacuolar invertase in plant development and photoassimilate partitioning.

Cyclopentenone isoprostanes induced by reactive oxygen species trigger defense gene activation and phytoalexin accumulation in plants.

Lipid peroxidation may be initiated either by lipoxygenases or by reactive oxygen species (ROS). Enzymatic oxidation of alpha-linolenate can result in the biosynthesis of cyclic oxylipins of the jasmonate type while free-radical-catalyzed oxidation of alpha-linolenate may yield several classes of cyclic oxylipins termed phytoprostanes in vivo. Previously, we have shown that one of these classes, the E1-phytoprostanes (PPE1), occurs ubiquitously in plants. In this work, it is shown that PPE1 are converted to novel cyclopentenone A1- and B1-phytoprostanes (PPA1 and PPB1) in planta. Enhanced formation of PPE1, PPA1, and PPB1 is observed after peroxide stress in tobacco cell cultures as well as after infection of tomato plants with a necrotrophic fungus, Botrytis cinerea. PPA1 and PPB1 display powerful biologic activities including activation of mitogen-activated protein kinase (MAPK) and induction of glutathione-S-transferase (GST), defense genes, and phytoalexins. Data collected so far infer that enhanced phytoprostane formation is a general consequence of oxidative stress in plants. We propose that phytoprostanes are components of an oxidant-injury-sensing, archaic signaling system that serves to induce several plant defense mechanisms.

Biochemical evidence for the activation of distinct subsets of mitogen-activated protein kinases by voltage and defense-related stimuli.

Activation of mitogen-activated protein (MAP) kinases is a common reaction of plant cells in defense-related signal transduction pathways. To gain insight into the mechanisms that determine specificity in response to a particular stimulus, a biochemical approach has been employed. Photoautotrophic suspension culture cells of tomato (Lycopersicon peruvianum) were used as experimental system to characterize MAP kinase activation by different stress-related stimuli. An elicitor preparation of the tomato-specific pathogen Fusarium oxysporum lycopersici was shown to result in the simultaneous induction of four kinase activities that could be separated by ion-exchange chromatography. The simultaneous activation of multiple MAP kinases was further substantiated by distinct pharmacological and immunological properties: a differential sensitivity toward various protein kinase inhibitors and a differential cross-reaction with isoform-specific MAP kinase antibodies. In contrast to the two fungal elicitors chitosan and the F. oxysporum lycopersici preparation, the plant-derived stimuli polygalacturonic acid and salicylic acid were shown to activate distinctly different subsets of MAP kinases. Application of a voltage pulse was introduced as a transient stress-related stimulus that does not persist in the culture. Voltage application activates a distinct set of MAP kinases, resembling those activated by salicylic acid treatment, and generates a refractory state for the salicylic acid response. The inhibitory effect of nifedipine indicates that current application may directly affect voltage-gated calcium channels, thus, providing a tool to study various calcium-dependent pathways.