Masanori Tamaoki

Ethylene Inhibits Abscisic Acid-Induced Stomatal Closure in Arabidopsis

To examine the cross talk between the abscisic acid (ABA) and ethylene signal transduction pathways, signaling events during ABA-induced stomatal closure were examined in Arabidopsis (Arabidopsis thaliana) wild-type plants, in an ethylene-overproducing mutant (eto1-1), and in two ethylene-insensitive mutants (etr1-1 and ein3-1). Using isolated epidermal peels, stomata of wild-type plants were found to close within a few minutes in response to ABA, whereas stomata of the eto1-1 mutant showed a similar but less sensitive ABA response. In addition, ABA-induced stomatal closure could be inhibited by application of ethylene or the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC). In contrast, stomata of the etr1-1 and ein3-1 mutants were able to close in response to concomitant ABA and ACC application, although to a lesser extent than in wild-type plants. Moreover, expression of the ABA-induced gene RAB18 was reduced following ACC application. These results indicate that ethylene delays stomatal closure by inhibiting the ABA signaling pathway. The same inhibitive effects of ethylene on stomatal closure were observed in ABA-irrigated plants and the plants in drought condition. Furthermore, upon drought stress, the rate of transpiration was greater in eto1-1 and wild-type plants exposed to ethylene than in untreated wild-type control plants, indicating that the inhibitive effects of ethylene on ABA-induced stomatal closure were also observed in planta.

Complete Genomic Structure of the Bloom-forming Toxic Cyanobacterium Microcystis aeruginosa NIES-843

Abstract The nucleotide sequence of the complete genome of a cyanobacterium, Microcystis aeruginosa NIES-843, was determined. The genome of M. aeruginosa is a single, circular chromosome of 5 842 795 base pairs (bp) in length, with an average GC content of 42.3%. The chromosome comprises 6312 putative protein-encoding genes, two sets of rRNA genes, 42 tRNA genes representing 41 tRNA species, and genes for tmRNA, the B subunit of RNase P, SRP RNA, and 6Sa RNA. Forty-five percent of the putative protein-encoding sequences showed sequence similarity to genes of known function, 32% were similar to hypothetical genes, and the remaining 23% had no apparent similarity to reported genes. A total of 688 kb of the genome, equivalent to 11.8% of the entire genome, were composed of both insertion sequences and miniature inverted-repeat transposable elements. This is indicative of a plasticity of the M. aeruginosa genome, through a mechanism that involves homologous recombination mediated by repetitive DNA elements. In addition to known gene clusters related to the synthesis of microcystin and cyanopeptolin, novel gene clusters that may be involved in the synthesis and modification of toxic small polypeptides were identified. Compared with other cyanobacteria, a relatively small number of genes for two component systems and a large number of genes for restriction-modification systems were notable characteristics of the M. aeruginosa genome.

Spermidine Synthase Genes Are Essential for Survival of Arabidopsis

The cellular polyamines putrescine, spermidine, and spermine are ubiquitous in nature and have been implicated in a wide range of growth and developmental processes. There is little information, however, on mutant plants or animals defective in the synthesis of polyamines. The Arabidopsis genome has two genes encoding spermidine synthase, SPDS1 and SPDS2. In this paper, we describe T-DNA insertion mutants of both of these genes. While each mutant allele shows normal growth, spds1-1 spds2-1 double-mutant seeds are abnormally shrunken and they have embryos that are arrested morphologically at the heart-torpedo transition stage. These seeds contain significantly reduced levels of spermidine and high levels of its precursor, putrescine. The embryo lethal phenotype of spds1-1 spds2-1 is complemented by the wild-type SPDS1 gene. In addition, we observed a nearly identical seed phenotype among an F2 seed population from the cross between the spds2-1 allele and SPDS1 RNA interference transgenic lines. These data provide the first genetic evidence indicating a critical role of the spermidine synthase in plant embryo development.

Light-controlled expression of a gene encoding l-galactono-γ-lactone dehydrogenase which affects ascorbate pool size in Arabidopsis thaliana☆

Abstract The cDNA clones for l -galactono-γ-lactone dehydrogenase (EC 1.3.2.3; GLDHase) that catalyzes the final step in the ascorbic acid (AsA) synthetic pathway, were isolated from Arabidopsis thaliana. The gene (AtGLDH) encodes a polypeptide of 610 amino acids and the predicted amino acid sequence was highly homologous to GLDHases from other plant species. The AtGLDH mRNA level of seedlings exhibited a diurnal change; the level was low in the morning and increased during the day. Its level in the evening (at 18:00) was about 2-fold that in the morning (at 6:00). Then the level decreased during the night until dawn of the next day. Similar diurnal changes were also observed in GLDHase activity and AsA content, suggesting that the AsA pool size is, at least partly, determined by the transcription rate of AtGLDH. Moreover, the diurnal changes of mRNA level, enzyme activity, and AsA content were not observed if plants were kept in continuous darkness. These results suggest that the diurnal change in expression of AtGLDH, which may play an important role in the regulation of AsA pool size, is regulated not by a circadian rhythm but by light.

The role of phytohormone signaling in ozone‑induced cell death in plants

Ozone is the main photochemical oxidant that causes leaf damage in many plant species, and can thereby significantly decrease the productivity of crops and forests. When ozone is incorporated into plants, it produces reactive oxygen species (ROS), such as superoxide radicals and hydrogen peroxide. These ROS induce the synthesis of several plant hormones, such as ethylene, salicylic acid, and jasmonic acid. These phytohormones are required for plant growth, development, and defense responses, and regulate the extent of leaf injury in ozone-fumigated plants. Recently, responses to ozone have been studied using genetically modified plants and mutants with altered hormone levels or signaling pathways. These researches have clarified the roles of phytohormones and the complexity of their signaling pathways. The present paper reviews the biosynthesis of the phytohormones ethylene, salicylic acid, and jasmonic acid, their roles in plant responses to ozone, and multiple interactions between these phytohormones in ozone-exposed plants.

The Arabidopsis sweetie mutant is affected in carbohydrate metabolism and defective in the control of growth, development and senescence

SUMMARY Sugars modulate many vital metabolic and developmental processes in plants, from seed germination to flowering, senescence and protection against diverse abiotic and biotic stresses. However, the exact mechanisms involved in morphogenesis, developmental signalling and stress tolerance remain largely unknown. Here we report the characterization of a novel Arabidopsis thaliana mutant, sweetie, with drastically altered morphogenesis, and a strongly modified carbohydrate metabolism leading to elevated levels of trehalose, trehalose-6-phosphate and starch. We additionally show that the disruption of SWEETIE causes significant growth and developmental alterations, such as severe dwarfism, lancet-shaped leaves, early senescence and flower sterility. Genes implicated in sugar metabolism, senescence, ethylene biosynthesis and abiotic stress were found to be upregulated in sweetie. Our physiological, biochemical, genetic and molecular data indicate that the mutation in sweetie was nuclear, single and recessive. The effects of metabolizable sugars and osmolytes on sweetie morphogenesis were distinct; in light, sweetie was hypersensitive to sucrose and glucose during vegetative growth and a partial phenotypic reversion took place in the presence of high sorbitol concentrations. However, SWEETIE encodes a protein that is unrelated to any known enzyme involved in sugar metabolism. We suggest that SWEETIE plays an important regulatory function that influences multiple metabolic, hormonal and stress-related pathways, leading to altered gene expression and pronounced changes in the accumulation of sugar, starch and ethylene.

The isochorismate pathway is negatively regulated by salicylic acid signaling in O3-exposed Arabidopsis

Ozone (O3), a major photochemical oxidant, causes leaf injury in plants. Plants synthesize salicylic acid (SA), which is reported to greatly affect O3 sensitivity. However, the mechanism of SA biosynthesis under O3 exposure remains unclear. Plants synthesize SA either by a pathway involving phenylalanine as a substrate or another involving isochorismate. To clarify how SA is produced in O3-exposed Arabidopsis, we examined the activities of phenylalanine ammonia lyase (PAL) and isochorismate synthase (ICS), which are components of the phenylalanine and isochorismate pathways, respectively. Exposure of Arabidopsis to O3 enhanced the accumulation of SA and the increase of ICS activity but did not affect PAL activity. In sid2 mutants, which have a defect in ICS1, the level of SA and the activity of ICS did not increase in response to O3 exposure. These results suggest that SA is mainly synthesized from isochorismate in Arabidopsis. Furthermore, the level of ICS1 expression and the activity of ICS during O3 exposure elevated in plants deficient for SA signaling (npr1 and eds5 mutants and NahG transgenics). Treatment of plants with SA also suppressed the enhancement of ICS1 expression by O3. These results suggest that SA synthesis is negatively regulated by SA signaling.

cDNA microarray assessment for ozone-stressed Arabidopsis thaliana

Various detrimental factors in the environment damage plants, resulting in growth inhibition or withering. However, it is not easy to identify causal factors by visually inspecting the damaged plants. Therefore, we have developed a sensitive and reliable method for plant diagnosis, based on measuring changes in expression of a set of genes in a DNA microarray. With this method, we have been able to detect and discriminate between plants stressed by ozone, drought, or wounding.

SAZ, a new SUPERMAN-like protein, negatively regulates a subset of ABA-responsive genes in Arabidopsis

Arabidopsis SUPERMAN (SUP) and members of its family are plant-unique C2H2-type zinc finger genes that have been implicated in plant growth and development. In this paper, we report that a new SUP-family gene, designated as SA- and ABA-downregulated zinc finger gene (SAZ), is involved in the negative regulation of ABA-mediated signaling. SAZ-GUS fusion proteins were predominantly localized in the nuclei when they were transiently expressed in onion epidermal cells. SAZ transcripts were expressed in the leaves and pistils of very young flower buds. In young seedlings, SAZ expression was downregulated in response to environmental stresses such as drought, salt, ozone and ultraviolet-B irradiation. This downregulation was also observed in response to the phytohormones salicylic acid (SA) and abscisic acid (ABA). SA-responsive downregulation of SAZ was not observed in the npr1-1 mutant, indicating that this regulation is NPR1 dependent. RNAi-mediated knockdown of SAZ (SAZ-kd) resulted in elevated expression of the drought- and ABA-responsive genes rd29B and rab18 under unstressed conditions, and it enhanced the response of these genes to drought and ABA treatment. The expression of several other drought- and/or ABA-responsive genes was not affected by SAZ-kd. Based on these results, we propose that SAZ plays a role in repressing a subset of the ABA-mediated stress-responsive genes in unstressed conditions.

Expression of genes encoding late nodulins characterized by a putative signal peptide and conserved cysteine residues is reduced in ineffective pea nodules

Five nodulin genes, PsN1, PsN6, PsN314, PsN335, and PsN466, with reduced expression in ineffective nodules on the pea (Pisum sativum) mutant E135 (sym13) were characterized. They encode small polypeptides containing a putative signal peptide and conserved cysteine residues and show homology to the nodulins PsENOD3/14 and PsNOD6. For each gene, multiple bands were detected by genomic Southern analysis. Northern analysis showed that all five genes were expressed exclusively in nodules and that their temporal expression patterns were similar to that of the leghemoglobin (Lb) gene during nodule development. Their transcripts were localized predominantly from the interzone II-III to the distal part of nitrogen-fixing zone III in effective nodules, resembling the Lb gene. However, transcripts in ineffective E135 nodules were localized in narrower regions than those in the effective nodules. These results indicate that these nodulins are abundant in pea nodules and that their successive expression during nodule development is associated with nitrogen-fixing activity.