Inositol Hexakis Phosphate is the Seasonal Phosphorus Reservoir in the Deciduous Woody Plant Populus alba L.

Seasonal recycling of nutrients is an important strategy for deciduous perennials. Deciduous perennials maintain and expand their nutrient pools by the autumn nutrient remobilization and the subsequent winter storage throughout their long life. Phosphorus (P), one of the most important elements in living organisms, is remobilized from senescing leaves during autumn in deciduous trees. However, it remains unknown how phosphate is stored over winter. Here we show that in poplar trees (Populus alba L.), organic phosphates are accumulated in twigs from late summer to winter, and that IP6 (myo-inositol-1,2,3,4,5,6-hexakis phosphate: phytic acid) is the primary storage form. IP6 was found in high concentrations in twigs during winter and quickly decreased in early spring. In parenchyma cells of winter twigs, P was associated with electron-dense structures, similar to globoids found in seeds of higher plants. Various other deciduous trees were also found to accumulate IP6 in twigs during winter. We conclude that IP6 is the primary storage form of P in poplar trees during winter, and that it may be a common strategy for seasonal P storage in deciduous woody plants.

Loss of heterophylly in aquatic plants: not ABA-mediated stress but exogenous ABA treatment induces stomatal leaves in Potamogeton perfoliatus.

Heterophyllous aquatic plants produce aerial (i.e., floating and terrestrial) and submerged leaves-the latter lack stomata-while homophyllous plants contain only submerged leaves, and cannot survive on land. To identify whether differences in morphogenetic potential and/or physiological stress responses are responsible for variation in phenotypic plasticity between two plants types, responses to abscisic acid (ABA) and salinity stress were compared between the closely related, but ecologically diverse pondweeds, Potamogeton wrightii (heterophyllous) and P. perfoliatus (homophyllous). The ABA-treated (1 or 10 µM) P. wrightii plants exhibited heterophylly and produced leaves with stomata. The obligate submerged P. perfoliatus plants were able to produce stomata on their leaves, but there were no changes to leaf shape, and stomatal production occurred only at a high ABA concentration (10 µM). Under salinity stress conditions, only P. wrightii leaves formed stomata. Additionally, the expression of stress-responsive NCED genes, which encode a key enzyme in ABA biosynthesis, was consistently up-regulated in P. wrightii, but only temporarily in P. perfoliatus. The observed species-specific gene expression patterns may be responsible for the induction or suppression of stomatal production during exposure to salinity stress. These results suggest that the two Potamogeton species have an innate morphogenetic ability to form stomata, but the actual production of stomata depends on ABA-mediated stress responses specific to each species and habitat.

Establishment of a shortened annual cycle system; a tool for the analysis of annual re-translocation of phosphorus in the deciduous woody plant (Populus alba L.).

The supply of phosphorus, the essential element for plant growth and development, is often limited in natural environments. Plants employ multiple physiological strategies to minimize the impact of phosphate deficiency. In deciduous trees, phosphorus is remobilized from senescing leaves in autumn and stored in other tissues for reuse in the following spring. We previously monitored the annual changes in leaf phosphate content of white poplar (Populus alba) growing under natural conditions and found that about 75 % of inorganic and 60 % of organic leaf phosphates observed in May were remobilized by November. In order to analyze this process (such annual events), we have established a model system, in which an annual cycle of phosphate re-translocation in trees can be simulated under laboratory conditions by controlling temperature and photoperiod (='shortened annual cycle'). This system evidently allowed us to monitor the annual changes in leaf color, phosphate remobilization from senescent leaves, and bud break in the next spring within five months. This will greatly facilitate the analysis of cellular and molecular mechanisms of annual phosphate re-translocation in deciduous trees.

Circumnutation on the water surface: female flowers of Vallisneria.

Circumnutation, the helical movement of growing organ tips, is ubiquitous in land plants. The mechanisms underlying circumnutation have been debated since Darwin's time. Experiments in space and mutant analyses have revealed that internal oscillatory (tropism-independent) movement and gravitropic response are involved in circumnutation. Female flower buds of tape grass (Vallisneria asiatica var. biwaensis) circumnutate on the water surface. Our observations and experiments with an artificial model indicated that gravitropism is barely involved in circumnutation. Instead, we show that helical intercalary growth at the base of peduncle plays the primary role in all movements in Vallisneria. This growth pattern produces torsional bud rotation, and gravity and buoyancy forces have a physical effect on the direction of peduncle elongation, resulting in bud circumnutation on the water surface. In contrast to other water-pollinated hydrophilous plants, circumnutation in Vallisneria enables female flowers to actively collect male flowers from a larger surface area of water.

Comparative studies of thermotolerance: different modes of heat acclimation between tolerant and intolerant aquatic plants of the genus Potamogeton.

BACKGROUND AND AIMS: Molecular-based studies of thermotolerance have rarely been performed on wild plants, although this trait is critical for summer survival. Here, we focused on thermotolerance and expression of heat shock transcription factor A2 (HSFA2) and its putative target gene (chloroplast-localized small heat shock protein, CP-sHSP) in two allied aquatic species of the genus Potamogeton (pondweeds) that differ in survival on land. METHODS: The degree of thermotolerance was examined using a chlorophyll bioassay to assess heat injury in plants cultivated under non- and heat-acclimation conditions. Potamogeton HSFA2 and CP-sHSP genes were identified and their heat-induction was quantified by real-time PCR. KEY RESULTS: The inhibition of chlorophyll accumulation after heat stress showed that Potamogeton malaianus had a higher basal thermotolerance and developed acquired thermotolerance, whereas Potamogeton perfoliatus was heat sensitive and unable to acquire thermotolerance. We found two duplicated HSFA2 and CP-sHSP genes in each species. These genes were induced by heat shock in P. malaianus, while one HSFA2a gene was not induced in P. perfoliatus. In non-heat-acclimated plants, transcript levels of HSFA2 and CP-sHSP were transiently elevated after heat shock. In heat-acclimated plants, transcripts were continuously induced during sublethal heat shock in P. malaianus, but not in P. perfoliatus. Instead, the minimum threshold temperature for heat induction of the CP-sHSP genes was elevated in P. perfoliatus. CONCLUSIONS: Our comparative study of thermotolerance showed that heat acclimation leads to species-specific changes in heat response. The development of acquired thermotolerance is beneficial for survival at extreme temperatures. However, the loss of acquired thermotolerance and plasticity in the minimum threshold temperature of heat response may be favourable for plants growing in moderate habitats with limited daily and seasonal temperature fluctuations.

Proton-induced tuning of metal-metal communication in rack-type dinuclear Ru complexes containing benzimidazolyl moieties.

Ru complexes bearing a bis-tridentate benzimidazolyl ligand have been synthesized. The dinuclear ones act as a bibasic acid with pK(a1)=4.36 and pK(a2)=5.90. The protonated form of the dinuclear complex exhibited two one-electron oxidations at +0.91 and +1.02 V versus the ferrocenium/ferrocene (Fc/Fc(+)) couple (the potential difference (ΔE)=0.11 V), but the di-deprotonated form showed two waves at +0.50 and +0.58 V versus Fc/Fc(+) (ΔE=0.08 V). Since the potential difference between two waves reflects the strength of the metal-metal interaction, the deprotonation of the benzimidazole moieties in the complexes weakened the Ru-Ru communication. The degree of electronic coupling between two metal centers, estimated from the intervalence charge transfer (IVCT) band, was greater for the protonated form. DFT calculations for the protonated and deprotonated forms of the dinuclear complex suggest that the Ru(II)-L(H(2)) π* interaction plays a key role in the Ru-Ru interaction.

AQUATIC PLANT SPECIATION AFFECTED BY DIVERSIFYING SELECTION OF ORGANELLE DNA REGIONS(1).

Many of the genes that control photosynthesis are carried in the chloroplast. These genes differ among species. However, evidence has yet to be reported revealing the involvement of organelle genes in the initial stages of plant speciation. To elucidate the molecular basis of aquatic plant speciation, we focused on the unique plant species Chara braunii C. C. Gmel. that inhabits both shallow and deep freshwater habitats and exhibits habitat-based dimorphism of chloroplast DNA (cpDNA). Here, we examined the "shallow" and "deep" subpopulations of C. braunii using two nuclear DNA (nDNA) markers and cpDNA. Genetic differentiation between the two subpopulations was measured in both nDNA and cpDNA regions, although phylogenetic analyses suggested nuclear gene flow between subpopulations. Neutrality tests based on Tajima's D demonstrated diversifying selection acting on organelle DNA regions. Furthermore, both "shallow" and "deep" haplotypes of cpDNA detected in cultures originating from bottom soils of three deep environments suggested that migration of oospores (dormant zygotes) between the two habitats occurs irrespective of the complete habitat-based dimorphism of cpDNA from field-collected vegetative thalli. Therefore, the two subpopulations are highly selected by their different aquatic habitats and show prezygotic isolation, which represents an initial process of speciation affected by ecologically based divergent selection of organelle genes.

Physiology and gene expression of the rice landrace Horkuch under salt stress.

Good donors in breeding for salt tolerance are a prerequisite for food security under changing climatic conditions. Horkuch, a farmer-popular salt tolerant rice (Oryza sativa L.) variety from the south-west coast of Bangladesh was characterised up to maturity under NaCl stress, together with a modern variety (BRRI dhan41), a sensitive control (BRRI dhan29) and Pokkali, the salt-tolerant benchmark for rice. Horkuch had low reduction in shoot biomass, a low Na:K ratio in flag leaves, a low percent reduction in yield and good partitioning of Na in the older leaves, and maintained high levels of Ca and Mg in the flag leaves. In order to understand the physiology at the molecular level, the expression of salt-responsive genes was investigated using microarray analysis. Salt-stressed cDNA of Horkuch seedlings were hybridised with cDNA probes synthesised mainly from database sequences of Arabidopsis thaliana (L.) Heynh. The upregulated genes included transcription factors, signal transducers, metabolic enzymes, reactive oxygen species (ROS) scavengers, osmoprotectants and some specific salt-induced transcripts. An increase in expression of photosynthesis-related genes as well ROS scavengers suggested that this could be the reason for the better yield performance of Horkuch. The data therefore indicate Horkuch as a potential donor alternative to Pokkali in breeding programs for salt tolerance.

Molecular adaptation of rbcL in the heterophyllous aquatic plant Potamogeton.

BACKGROUND: Heterophyllous aquatic plants show marked phenotypic plasticity. They adapt to environmental changes by producing different leaf types: submerged, floating and terrestrial leaves. By contrast, homophyllous plants produce only submerged leaves and grow entirely underwater. Heterophylly and submerged homophylly evolved under selective pressure modifying the species-specific optima for photosynthesis, but little is known about the evolutionary outcome of habit. Recent evolutionary analyses suggested that rbcL, a chloroplast gene that encodes a catalytic subunit of RuBisCO, evolves under positive selection in most land plant lineages. To examine the adaptive evolutionary process linked to heterophylly or homophylly, we analyzed positive selection in the rbcL sequences of ecologically diverse aquatic plants, Japanese Potamogeton. PRINCIPAL FINDINGS: Phylogenetic and maximum likelihood analyses of codon substitution models indicated that Potamogeton rbcL has evolved under positive Darwinian selection. The positive selection has operated specifically in heterophyllous lineages but not in homophyllous ones in the branch-site models. This suggests that the selective pressure on this chloroplast gene was higher for heterophyllous lineages than for homophyllous lineages. The replacement of 12 amino acids occurred at structurally important sites in the quaternary structure of RbcL, two of which (residue 225 and 281) were identified as potentially under positive selection. CONCLUSIONS/SIGNIFICANCE: Our analysis did not show an exact relationship between the amino acid replacements and heterophylly or homophylly but revealed that lineage-specific positive selection acted on the Potamogeton rbcL. The contrasting ecological conditions between heterophyllous and homophyllous plants have imposed different selective pressures on the photosynthetic system. The increased amino acid replacement in RbcL may reflect the continuous fine-tuning of RuBisCO under varying ecological conditions.

Inherited maternal effects on the drought tolerance of a natural hybrid aquatic plant, Potamogeton anguillanus.

We tested whether maternal effects have led to the adaptive divergence of strains of the natural hybrid Potamogeton anguillanus, whose putative parents show contrastingly divergent ecologies. To examine the correlation between phenotypic characters and maternal types, we conducted drought experiments and DNA typing using nuclear and chloroplast genes. In the field, we investigated the distribution of the maternal type along the depth and the inshore-offshore gradient. Hybrids of P. malaianus mothers (M-hybrids) and those of P. perfoliatus mothers (P-hybrids) could not be distinguished morphologically under submerged conditions, but differed in drought tolerance. M-hybrids and P. malaianus formed more terrestrial shoots and exhibited higher survival than P-hybrids and P. perfoliatus in drought experiments. The distribution survey clarified that M-hybrids were dominant in shallow and inshore areas, whereas they were almost absent in deeper and offshore areas. These results indicate that the natural hybrid P. anguillanus differs in adaptive values depending on the maternal type. Bidirectional hybridization and heritable maternal effects may have played important roles in its phenotypic adaptation to local environmental conditions.

Phylogeny of vertebrate Src tyrosine kinases revealed by the epitope region of mAb327.

Mass fingerprinting and MS/MS analysis demonstrated that Xyk, a 57-kDa Src family tyrosine kinase that is activated within minutes of Xenopus egg fertilization, comprises a mixture of two Src proteins, Src1 and Src2. However, the Xenopus Src protein, denoted as xSrc, is hardly detectable with mAb327, a universal Src-specific antibody, whose target sequence has not yet been determined. We show that a point amino acid substitution in the Src homology 3 domain of xSrc is critical for improvement of the low efficiency of its recognition by mAb327. Namely, a point-mutated xSrc, in which Arg-121 was replaced by His that is conserved among mAb327-reactive Src in mammals and chicken, showed increased recognition by mAb327. On the other hand, a mutant chicken Src, in which the His-122 residue is replaced by Arg, showed decreased recognition by mAb327. Genomic sequencing analysis also demonstrated that reptile Src proteins are of either the R-type (snake) or H-type (caiman, turtle, and tortoise). These studies revealed, for the first time, a critical amino acid in the Src SH3 domain for mAb327 recognition, and suggest a novel scheme for the molecular evolution of Src, in which the H-type Src(s) are monophyletic and derived from the R-type Src.

Morphology and phylogenetic position of a mat-forming green plant from acidic rivers in Japan.

A green plant, which we have named "Misuzugoke", was found in acidic rivers in Nagano Prefecture, Japan, where it forms macroscopic mats or aggregates. The field-collected aggregates were nearly hemispherical and were composed of compact branched filaments radiating from a central base. The cells of the plant contained a single nucleus and numerous discoid chloroplasts lacking pyrenoids. Due to the plant's simple filamentous organization and absence of reproductive organs, its classification as an alga or a higher plant could not be determined from the field-collected material. When grown for more than 3 months on neutral agar medium (pH 7.0) containing hormones, leafy shoot buds characteristic of bryophytes appeared on the plant. A partial sequence of the plant's rbcL gene, which encodes the large subunit of ribulose-1, 5-bisphosphate carboxylase/oxygenase was 98% similar to that of Dicranella heteromalla (Hedw.) Schimp. (Haplolepideae, Bryopsida). Phylogenetic analyses based on rbcL gene sequences strongly indicate that Misuzugoke is positioned within the Haplolepideae. This mat-forming green plant is therefore considered to be a reduced form of moss in the Haplolepideae.