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Alteration in the leaf mesophyll anatomy by genetic modification is potentially a promising tool for improving the physiological functions of trees by improving leaf photosynthesis. Homeodomain leucine zipper (HD-Zip) transcription factors are candidates for anatomical alterations of leaves through modification of cell multiplication, differentiation, and expansion. Full-length cDNA encoding a Eucalyptus camaldulensis HD-Zip class II transcription factor (EcHB1) was over-expressed in vivo in the hybrid Eucalyptus GUT5 generated from Eucalyptus grandis and Eucalyptus urophylla. Overexpression of EcHB1 induced significant modification in the mesophyll anatomy of Eucalyptus with enhancements in the number of cells and chloroplasts on a leaf-area basis. The leaf-area-based photosynthesis of Eucalyptus was improved in the EcHB1-overexpression lines, which was due to both enhanced CO 2 diffusion into chloroplasts and increased photosynthetic biochemical functions through increased number of chloroplasts per unit leaf area. Additionally, overexpression of EcHB1 suppressed defoliation and thus improved the growth of Eucalyptus trees under drought stress, which was a result of reduced water loss from trees due to the reduction in leaf area with no changes in stomatal morphology. These results gave us new insights into the role of the HD-Zip II gene.

Direct measurements of ecophysiological processes such as leaf photosynthesis are often hampered due to the excessive time required for gas-exchange measurements and the limited availability of multiple gas analyzers. Although recent advancements in commercially available instruments have improved the ability to take measurements more conveniently, the amount of time required for each plant sample to acclimate to chamber conditions has not been sufficiently reduced. Here we describe a system of multiple gas-exchange chambers coupled with a laser spectrometer that employs tunable diode laser absorption spectroscopy (TDLAS) to measure leaf photosynthesis, stomatal conductance, and mesophyll conductance. Using four gas-exchange chambers minimizes the time loss associated with acclimation for each leaf sample. System operation is semiautomatic, and leaf temperature, humidity, and CO 2 concentration can be regulated and monitored remotely by a computer system. The preliminary results with rice leaf samples demonstrated that the system is capable of high-throughput measurements, which is necessary to obtain better representativeness of the ecophysiological characteristics of plant samples.

The physiological and anatomical responses of bryophytes to altered gravity conditions will provide crucial information for estimating how plant physiological traits have evolved to adapt to significant increases in the effects of gravity in land plant history. We quantified changes in plant growth and photosynthesis in the model plant of mosses, Physcomitrella patens, grown under a hypergravity environment for 25 days or 8 weeks using a custom-built centrifuge equipped with a lighting system. This is the first study to examine the response of bryophytes to hypergravity conditions. Canopy-based plant growth was significantly increased at 10×g, and was strongly affected by increases in plant numbers. Rhizoid lengths for individual gametophores were significantly increased at 10×g. Chloroplast diameters (major axis) and thicknesses (minor axis) in the leaves of P. patens were also increased at 10×g. The area-based photosynthesis rate of P. patens was also enhanced at 10×g. Increases in shoot numbers and chloroplast sizes may elevate the area-based photosynthesis rate under hypergravity conditions. We observed a decrease in leaf cell wall thickness under hypergravity conditions, which is in contrast to previous findings obtained using angiosperms. Since mosses including P. patens live in dense populations, an increase in canopy-based plant numbers may be effective to enhance the toughness of the population, and, thus, represents an effective adaptation strategy to a hypergravity environment for P. patens.

We investigated stomatal conductance (g(s)) and mesophyll conductance (g(m)) in response to atmospheric CO2 concentration [CO2] in two primitive land plants, the fern species Pteridium aquilinum and Thelypteris dentata, using the concurrent measurement of leaf gas exchange and carbon isotope discrimination. [CO2] was initially decreased from 400 to 200 μmol mol(-1), and then increased from 200 to 700 μmol mol(-1), and finally decreased from 700 to 400 μmol mol(-1). Analysis by tunable diode laser absorption spectroscopy (TDLAS) revealed a rapid and continuous response in g m within a few minutes. In most cases, both ferns showed rapid and significant responses of g m to changes in [CO2]. The largest changes (quote % decrease) were obtained when [CO2] was decreased from 400 to 200 μmol mol(-1). This is in contrast to angiosperms where an increase in g(m) is commonly observed at low [CO2]. Similarly, fern species observed little or no response of g(s) to changes in [CO2] whereas, a concomitant decline of g(m) and g(s) with [CO2] is often reported in angiosperms. Together, these results suggest that regulation of g(m) to [CO2] may differ between angiosperms and ferns.

Eucalyptus is a diverse genus of flowering trees with more than 700 genotypic species which are mostly native to Australia. We selected 19 wild provenances of Eucalyptus camaldulensis grown in Australia, compared their growth rate and drought tolerance and determined the protein levels of plasma membrane aquaporins (PIPs). There was a positive relationship between the drought tolerance and PIP content. PIPs are divided into two subgroups, PIP1 and PIP2. Most members of the PIP2 subgroup, but not PIP1 subgroup, exhibit water channel activity. We introduced two radish (Raphanus sativus L.) PIPs, RsPIP1;1 and RsPIP2;1, into a hybrid clone of Eucalyptus grandis and Eucalyptus urophylla to examine the effect of their overexpression. Expression of these genes was confirmed by real-time polymerase chain reaction (PCR) and the protein accumulation of RsPIP2;1 by immunoblotting. Drought tolerance was not enhanced in transgenic lines of either gene. However, one transgenic line expressing RsPIP2;1 showed high photosynthesis activity and growth rate under normal growth conditions. For RsPIP1;1-transformed lines, the RsPIP1;1 protein did not accumulate, and the abundance of endogenous PIP1 and PIP2 was decreased. The endogenous PIP1 and PIP2 genes were suppressed in these lines. Therefore, the decreased levels of PIP1 and PIP2 protein may be due to co-suppression of the PIP genes and/or high turnover of PIP proteins. RsPIP1;1-expressing lines gave low values of photosynthesis and growth compared with the control. These results suggest that down-regulation of PIP1 and PIP2 causes serious damage and that up-regulation of PIP2 improves the photosynthetic activity and growth of Eucalyptus trees.

The photosynthetic rate may be strongly limited by internal conductance from the intercellular airspace to the chloroplast stroma (g(i)). However, the effects of growth and leaf temperature on g(i) are still unclarified. In this work, we determined the temperature dependence of g(i) in spinach leaves grown at 30/25 degrees C (high temperature; HT) and 15/10 degrees C (low temperature; LT), using the concurrent measurements of the gas exchange rate and stable carbon isotope ratio. Moreover, we quantified the effects of g(i) on the temperature dependence of the photosynthetic rate. We measured g(i) and the photosynthetic rate at a CO(2) concentration of 360 microl l(-1) under saturating light (A(360)) at different leaf temperatures. The optimum temperature for A(360) was 28.5 degrees C in HT leaves and 22.9 degrees C in LT leaves. The optimum temperatures for g(i) were almost similar to those of A(360) in both HT and LT leaves. There was a strong linear relationship between A(360) and g(i). The photosynthetic rates predicted from the C(3) photosynthesis model taking account of g(i) agreed well with A(360) in both HT and LT leaves. The temperature coefficients (Q(10)) of g(i) between 10 and 20 degrees C were 2.0 and 1.8 in HT and LT leaves, respectively. This suggests that g(i) was determined not only by physical diffusion but by processes facilitated by protein(s). The limitation of the photosynthetic rate imposed by g(i) increased with leaf temperature and was greater than the limitation of the stomatal conductance at any temperature, in both HT and LT leaves. This study suggests that g(i) substantially limits the photosynthetic rate, especially at higher temperatures.

The subject of this paper, sun leaves are thicker and show higher photosynthetic rates than the shade leaves, is approached in two ways. The first seeks to answer the question: why are sun leaves thicker than shade leaves? To do this, CO2 diffusion within a leaf is examined first. Because affinity of Rubisco for CO2 is low, the carboxylation of ribulose 1,5-bisphosphate is competitively inhibited by O2, and the oxygenation of ribulose 1,5-bisphosphate leads to energy-consuming photorespiration, it is essential for C3 plants to maintain the CO2 concentration in the chloroplast as high as possible. Since the internal conductance for CO2 diffusion from the intercellular space to the chloroplast stroma is finite and relatively small, C3 leaves should have sufficient mesophyll surfaces occupied by chloroplasts to secure the area for CO2 dissolution and transport. This explains why sun leaves are thicker. The second approach is mechanistic or 'how-oriented'. Mechanisms are discussed as to how sun leaves become thicker than shade leaves, in particular, the long-distance signal transduction from mature leaves to leaf primordia inducing the periclinal division of the palisade tissue cells. To increase the mesophyll surface area, the leaf can either be thicker or have smaller cells. Issues of cell size are discussed to understand plasticity in leaf thickness.

Background and Aims For a comprehensive understanding of the mechanisms of changing plant photosynthetic capacity during plant evolutionary history, knowledge of leaf gas exchange and optical properties are essential, both of which relate strongly to mesophyll anatomy. Although ferns are suitable for investigating the evolutionary history of photosynthetic capacity, comprehensive research of fern species has yet to be undertaken in this regard. Methods We investigated leaf optical properties, gas exchange and mesophyll anatomy of fern species with a wide range of divergence time, using 66 ferns from natural habitats and eight glasshouse-grown ferns. We used a spectroradiometer and an integrating sphere to measure light absorptance and reflectance by the leaves. Key Results The more newly divergent fern species had a thicker mesophyll, a larger surface area of chloroplasts facing the intercellular airspaces (S c), thicker cell walls and large light absorptance. Although no trend with divergence time was obtained in leaf photosynthetic capacity on a leaf-area basis, when the traits were expressed on a mesophyll-thickness basis, trends in leaf photosynthetic capacity became apparent. On a mesophyll-thickness basis, the more newly divergent species had a low maximum photosynthesis rate, accompanied by a low S c. Conclusions We found a strong link between light capture, mesophyll anatomy and photosynthesis rate in fern species for the first time. The thick mesophyll of the more newly divergent ferns does not necessarily relate to the high photosynthetic capacity on a leaf-area basis. Rather, the thick mesophyll accompanied by thick cell walls allowed the ferns to adapt to a wider range of environments through increasing leaf toughness, which would contribute to the diversification of fern species. [ABSTRACT FROM AUTHOR]

Plants respond to environmental stressors, such as an oligotrophic environments, by altering the morphological and physiological functions of their leaves. Sex affects these functions because of the asymmetric cost of reproduction in dioecious plants. We compared the leaf mass per leaf area (LMA), ratio of intercellular air space in leaf mesophyll tissue (mesophyll porosity), palisade thickness, and carbon isotope ratio (δ13C) of leaves of the dioecious shrub Myrica gale based on sex and gradients of soil water chemistry across habitats in the field. The PCA showed that the first three principal components accounted for 84.5% of the variation. PC1 to PC3 were associated with the origin of soil water, nitrogen status of habitats, and sea–salt contributions, respectively. LMA varied from 5.22 to 7.13 μg/cm2, and it was positively related to PC2 and negatively related to PC3, but not to PC1 or sex, suggesting that LMA was low under poor nitrogen conditions and varied with salinity. Mesophyll porosity values were over 50% for all habitats. Mesophyll porosity was positively affected by PC3 and smaller in females than in males. This suggests that M. gale exhibits differences in mesophyll anatomy according to sex. Palisade thickness ranged from 0.466 to 0.559 mm/mm. The leaves of females had thinner palisade layers per mesophyll layer than those of males; however, the habitat did not affect the thickness of the palisade layer per mesophyll layer. The δ13C values of leaves varied from −32.14 to −30.51 ‰. We found that δ13C values were positively related to PC2 but not to PC1, PC3, and sex. Under poor nitrogen conditions, the δ13C of M. gale leaves decreased, suggesting that nutrient deficiency would decrease more under the long-term averaged ratio of photosynthesis than stomatal conductance, leading to low water use efficiency. [ABSTRACT FROM AUTHOR]

For 383 Poaceae species harvested over the Japanese islands and stored as herbarium specimens along several decades, we determined C3 and C4 types of photosynthesis from leaf stable carbon isotope ratio (δ13C). Then, we sought the relationships between C4 species richness and climatic factors or habitat types. Except for the two Panicum species ( P. lanuginosum and P. scoparium) having the possibility of C3–C4 intermediate, 227 and 154 species were classified into C3 and C4. The C4 species richness increased from northern to southern islands in Japan, positively correlated with mean annual air temperature. Greater C4 species richness in the seashore habitats, and smaller C4 species richness in the shaded, wet and highland habitats would be related to the photosynthetic responses to local environmental factors such as irradiance level and temperature regime. No difference of leaf δ-value of C3 Poaceae was obtained between the habitats with different soil water availability, suggesting the less importance of soil water availability on leaf water-use efficiency in C3 Poaceae species in Japan having humid climate. Additionally, possible effects of human activity around the harvested time or site on leaf δ-value were estimated, because the habitat includes the sites with high human activity. Leaf δ-value was decreased with sampling year, and it was higher in the densely inhabited district for both C3 and C4. They are probably due to a historical decrease in the atmospheric δ-value via increasing human activity, and high gas emission at the districts of high human density. [ABSTRACT FROM AUTHOR]

To evaluate the effects of air pollution on the decline of Pinus densiflora forests, various research has been conducted around Mt. Gokurakuji (34° 23′N, 132° 19′E, 693 m a.s.l.) north of the Seto Inland Sea, west Japan. To investigate the mechanisms responsible for decreases in photosynthesis ( Pn) and stomatal conductance ( gl), δ13C of needles and seasonal changes in the water content (WC) and abscisic acid concentration (ABA) of needles were measured in various stands. The δ13C values were less negative in declining stands and younger needles. ABA and WC were not correlated with each other. WC decreased consistently with needle age while the ABA showed a minimum in August and a smaller content in older needles. Monthly precipitation and the daily maximum vapor pressure were not correlated with ABA and WC. In declining stands, WC and ABA tended to be higher and lower, respectively, than in nondeclining stands. These results suggest that the trees in declining stands received less water stress than those in nondeclining stands and the differences in gl and δ13C are not caused by the difference in water stress. The possibilities of the effects of air pollution and the infection of pine-wood nematode on the physiological decline on the pine needles are discussed. [ABSTRACT FROM AUTHOR]

The ecophysiological effects of understory vegetation on an overstory tree Pinus densiflora were examined in secondary pine forest stands, west Japan. Three pairs of pine forests were compared. Each pair contained an unmanaged stand and a managed stand in which the understory vegetation was clear cut. The two stands in each pair were adjacent to each other and had similar light conditions, ground matrix, precipitation and aerial conditions. Total fine root biomass in the soil surface layer was significantly larger in the unmanaged stands than in the managed stands because of the invasion of other understory trees. In the unmanaged stands, the maximum net photosynthesis (Pmax) and stomatal conductance (glmax) were smaller and δ13C values of needles were larger than those in the managed stands. These results suggested that water for pine needles was more restricted and water use efficiency (WUE) was higher in the unmanaged stand than in the managed stands. Nitrogen contents in the needles and photosynthetic nitrogen use efficiency (NUE) in the unmanaged stands were lower than those in the managed stands. There was no significant difference between the managed and the unmanaged stands in the emission of stress ethylene from needles. Our results showed that the understory vegetation which invaded during the process of secondary succession had negative physiological effects on the overstory pine needles through the competition of roots for water and nutrients. The traditional style of forest floor management in Japan had significant positive effects on the health condition of P. densiflora plantations. [Copyright &y& Elsevier]

The original article can be found online. [ABSTRACT FROM AUTHOR]