During photoinhibition of photosystem II (PSII) in cyanobacteria, salt stress inhibits the repair of photodamaged PSII and, in particular, the synthesis of the D1 protein (D1). We investigated the effects of salt stress on the repair of PSII and the synthesis of D1 in wild-type tobacco (Nicotiana tabacum 'Xanthi') and in transformed plants that harbored the katE gene for catalase from Escherichia coli. Salt stress due to NaCl enhanced the photoinhibition of PSII in leaf discs from both wild-type and katE-transformed plants, but the effect of salt stress was less significant in the transformed plants than in wild-type plants. In the presence of lincomycin, which inhibits protein synthesis in chloroplasts, the activity of PSII decreased rapidly and at similar rates in both types of leaf disc during photoinhibition, and the observation suggests that repair of PSII was protected by the transgene-coded enzyme. Incorporation of [(35)S]methionine into D1 during photoinhibition was inhibited by salt stress, and the transformation mitigated this inhibitory effect. Northern blotting revealed that the level of psbA transcripts was not significantly affected by salt stress or by the transformation. Our results suggest that salt stress enhanced photoinhibition by inhibiting repair of PSII and that the katE transgene increased the resistance of the chloroplast's translational machinery to salt stress by scavenging hydrogen peroxide.

Seasonal dynamics of net photosynthesis (Anet) in 2-year-old seedlings of Pinus brutia Ten., Pinus pinea L. and Pinus pinaster Ait. were investigated. Seedlings were grown in the field in two light regimes: sun (ambient light) and shade (25% of photosynthetically active radiation (PAR)). Repeated measures analyses over a 12-month period showed that Anet varied significantly among species and from season to season. Maximum Anet in sun-acclimated seedlings was low in winter (yet remained positive) and peaked during summer. Maximum Anet was observed in June in P. pinea (12 micromol m-2 s-1), July in P. pinaster (23 micromol m-2 s-1) and August in P. brutia (20 micromol m-2 s-1). Photosynthetic light response curves saturated at a PAR of 200-300 micromol m-2 s-1 in winter and in shade-acclimated seedlings in summer. Net photosynthesis in sun-acclimated seedlings did not saturate at PAR up to 1900 micromol m-2 s-1 in P. brutia and P. pinaster. Minimum air temperature of the preceding night was apparently one of the main factors controlling Anet during the day. In shade-acclimated seedlings, photosynthetic rates were reduced by 50% in P. brutia and P. pinaster and by 20% in P. pinea compared with those in sun-acclimated seedlings. Stomatal conductance was generally lower in shaded seedlings than in seedlings grown in the sun, except on days with a high vapor pressure deficit. Total chlorophyll concentration per unit leaf area, specific leaf area (SLA) and height significantly increased in P. pinea in response to shade, but not in P. pinaster or P. brutia. In response to shade, P. brutia showed a significant increase in total chlorophyll concentration but not SLA. Photosynthetic and growth data indicate that P. pinaster and P. brutia are more light-demanding than P. pinea.

Application of the widely used Farquhar model of photosynthesis in interpretation of gas exchange data assumes that photosynthetic properties are homogeneous throughout the leaf. Previous studies showed that heterogeneity in stomatal conductance (g(s)) across a leaf could affect the shape of the measured leaf photosynthetic CO(2) uptake rate (A) versus intercellular CO(2) concentration (C(i)) response curve and, in turn, estimation of the critical biochemical parameters of this model. These are the maximum rates of carboxylation (V(c,max)), whole-chain electron transport (J(max)), and triose-P utilization (V(TPU)). The effects of spatial variation in V(c,max,) J(max), and V(TPU) on estimation of leaf averages of these parameters from A-C(i) curves measured on a whole leaf have not been investigated. A mathematical model incorporating defined degrees of spatial variability in V(c,max) and J(max) was constructed. One hundred and ten theoretical leaves were simulated, each with the same average V(c,max) and J(max), but different coefficients of variation of the mean (CV(VJ)) and varying correlation between V(c,max) and J(max) (Omega). Additionally, the interaction of variation in V(c,max) and J(max) with heterogeneity in V(TPU), g(s), and light gradients within the leaf was also investigated. Transition from V(c,max)- to J(max)-limited photosynthesis in the A-C(i) curve was smooth in the most heterogeneous leaves, in contrast to a distinct inflection in the absence of heterogeneity. Spatial variability had little effect on the accuracy of estimation of V(c,max) and J(max) from A-C(i) curves when the two varied in concert (Omega = 1.0), but resulted in underestimation of both parameters when they varied independently (up to 12.5% in V(c,max) and 17.7% in J(max) at CV(VJ) = 50%; Omega = 0.3). Heterogeneity in V(TPU) also significantly affected parameter estimates, but effects of heterogeneity in g(s) or light gradients were comparatively small. If V(c,max) and J(max) derived from such heterogeneous leaves are used in models to project leaf photosynthesis, actual A is overestimated by up to 12% at the transition between V(c,max)- and J(max)-limited photosynthesis. This could have implications for both crop production and Earth system models, including projections of the effects of atmospheric change.

Various aspects of the biochemistry of photosynthetic carbon assimilation in C3 plants are integrated into a form compatible with studies of gas exchange in leaves. These aspects include the kinetic properties of ribulose bisphosphate carboxylase-oxygenase; the requirements of the photosynthetic carbon reduction and photorespiratory carbon oxidation cycles for reduced pyridine nucleotides; the dependence of electron transport on photon flux and the presence of a temperature dependent upper limit to electron transport. The measurements of gas exchange with which the model outputs may be compared include those of the temperature and partial pressure of CO2(p(CO2)) dependencies of quantum yield, the variation of compensation point with temperature and partial pressure of O2(p(O2)), the dependence of net CO2 assimilation rate on p(CO2) and irradiance, and the influence of p(CO2) and irradiance on the temperature dependence of assimilation rate.

Blue light regulates many molecular and physiological activities in a large number of organisms. In Neurospora crassa, a eukaryotic model system for studying blue-light responses, the transcription factor and blue-light photoreceptor WHITE COLLAR-1 (WC-1) and its partner WC-2 are central to blue-light sensing. Neurospora's light responses are transient, that is, following an initial acute phase of induction, light-regulated processes are down-regulated under continuous illumination, a phenomenon called photoadaptation. The molecular mechanism(s) of photoadaptation are not well understood. Here we show that a common mechanism controls the light-induced transcription of immediate early genes (such as frq, al-3, and vvd) in Neurospora, in which light induces the binding of identical large WC-1/WC-2 complexes (L-WCC) to the light response elements (LREs) in their promoters. Using recombinant proteins, we show that the WC complexes are functional without the requirement of additional factors. In vivo, WCC has a long period photocycle, indicating that it cannot be efficiently used for repeated light activation. Contrary to previous expectations, we demonstrate that the light-induced hyperphosphorylation of WC proteins inhibits bindings of the L-WCC to the LREs. We show that, in vivo, due to its rapid hyperphosphorylation, L-WCC can only bind transiently to LREs, indicating that WCC hyperphosphorylation is a critical process for photoadaptation. Finally, phosphorylation was also shown to inhibit the LRE-binding activity of D-WCC (dark WC complex), suggesting that it plays an important role in the circadian negative feedback loop.

We report here the striking anisotropy of fluorescence exhibited by crystals of native green fluorescence protein (GFP). The crystals were generated by water dialysis of highly purified GFP obtained from the jellyfish Aequorea. We find that the fluorescence becomes six times brighter when the excitation, or emission, beam is polarized parallel (compared with perpendicular) to the crystal long axis. Thus, the major dipoles of the fluorophores must be oriented very nearly parallel to the crystal long axis. Observed in a polarizing microscope between parallel polars instead of either a polarizer or analyzer alone, the fluorescence polarization ratio rises to an unexpectedly high value of about 30:1, nearly the product of the fluorescence excitation and emission ratios, suggesting a sensitive method for measuring fluorophore orientations, even of a single fluorophore molecule. We have derived equations that accurately describe the relative fluorescence intensities of crystals oriented in various directions, with the polarizer and analyzer arranged in different configurations. The equations yield relative absorption and fluorescence coefficients for the four transition dipoles involved. Finally, we propose a model in which the elongated crystal is made of GFP molecules that are tilted 60 degrees to align the fluorophores parallel to the crystal long axis. The unit layer in the model may well correspond to the arrangement of functional GFP molecules, to which resonant energy is efficiently transmitted from Ca2+-activated aequorin, in the jellyfish photophores.

The principles, equipment and procedures for measuring leaf and canopy gas exchange have been described previously as has chlorophyll fluorescence. Simultaneous measurement of the responses of leaf gas exchange and modulated chlorophyll fluorescence to light and CO2 concentration now provide a means to determine a wide range of key biochemical and biophysical limitations on photo synthesis in vivo. Here the mathematical frameworks and practical procedures for determining these parameters in vivo are consolidated. Leaf CO2 uptake (A) versus intercellular CO2 concentration (Ci) curves may now be routinely obtained from commercial gas exchange systems. The potential pitfalls, and means to avoid these, are examined. Calculation of in vivo maximum rates of ribulose-1,5-bisphosphate (RuBP) carboxylase/oxygenase (Rubisco) carboxylation (Vc,max), electron transport driving regeneration of RuBP (Jmax), and triose-phosphate utilization (VTPU) are explained; these three parameters are now widely assumed to represent the major limitations to light-saturated photosynthesis. Precision in determining these in intact leaves is improved by the simultaneous measurement of electron transport via modulated chlorophyll fluorescence. The A/Ci response also provides a simple practical method for quantifying the limitation that stomata impose on CO2 assimilation. Determining the rate of photorespiratory release of oxygen (Rl) has previously only been possible by isotopic methods, now, by combining gas exchange and fluorescence measurements, Rl may be determined simply and routinely in the field. The physical diffusion of CO2 from the intercellular air space to the site of Rubisco in C3 leaves has long been suspected of being a limitation on photosynthesis, but it has commonly been ignored because of the lack of a practical method for its determination. Again combining gas exchange and fluorescence provides a means to determine mesophyll conductance. This method is described and provides insights into the magnitude and basis of this limitation.

Transcription of the Neurospora crassa gene con-10 is induced during conidiation and following exposure of vegetative mycelia to light, but light activation is transient due to photoadaptation. We describe mutational analyses of photoadaptation using a N. crassa strain bearing a translational fusion of con-10, including its regulatory region, to a selectable bacterial gene conferring hygromycin resistance (hph). Growth of this strain was sensitive to hygromycin, upon continuous culture in the light. Five mutants were isolated that were resistant to hygromycin when cultured under constant light. Three mutant strains displayed elevated, sustained accumulation of con-10::hph mRNA during continued light exposure, suggesting that they bear mutations that reduce or eliminate the presumed light-dependent repression mechanism that blocks con-10 transcription upon prolonged illumination. These mutations altered photoadaptation for only a specific group of genes (con-10 and con-6), suggesting that regulation of photoadaptation is relatively gene specific. The mutations increased light-dependent mRNA accumulation for genes al-1, al-2, and al-3, each required for carotenoid biosynthesis, resulting in a threefold increase in carotenoid accumulation following continuous light exposure. Identification of the altered gene or genes in these mutants may reveal novel proteins that participate in light regulation of gene transcription in fungi.

BACKGROUND AND AIMS: Theory for optimal allocation of foliar nitrogen (ONA) predicts that both nitrogen concentration and photosynthetic capacity will scale linearly with gradients of insolation within plant canopies. ONA is expected to allow plants to efficiently use both light and nitrogen. However, empirical data generally do not exhibit perfect ONA, and light-use optimization per se is little explored. The aim was to examine to what degree partitioning of nitrogen or light is optimized in the crowns of three tropical canopy tree species. METHODS: Instantaneous photosynthetic photon flux density (PPFD) incident on the adaxial surface of individual leaves was measured along vertical PPFD gradients in tree canopies at a frequency of 0.5 Hz over 9-17 d, and summed to obtain the average daily integral of PPFD for each leaf to characterize its insolation regime. Also measured were leaf N per area (N(area)), leaf mass per area (LMA), the cosine of leaf inclination and the parameters of the photosynthetic light response curve [photosynthetic capacity (A(max)), dark respiration (R(d)), apparent quantum yield (phi) and curvature (theta)]. The instantaneous PPFD measurements and light response curves were used to estimate leaf daily photosynthesis (A(daily)) for each leaf. KEY RESULTS: Leaf N(area) and A(max) changed as a hyperbolic asymptotic function of the PPFD regime, not the linear relationship predicted by ONA. Despite this suboptimal nitrogen partitioning among leaves, A(daily) did increase linearly with PPFD regime through co-ordinated adjustments in both leaf angle and physiology along canopy gradients in insolation, exhibiting a strong convergence among the three species. CONCLUSIONS: The results suggest that canopy tree leaves in this tropical forest optimize photosynthetic use of PPFD rather than N per se. Tropical tree canopies then can be considered simple 'big-leaves' in which all constituent 'small leaves' use PPFD with the same photosynthetic efficiency.

Leaves of C(3) plants which exhibit a normal O(2) inhibition of CO(2) fixation at less than saturating light intensity were found to exhibit O(2)-insensitive photosynthesis at high light. This behavior was observed in Phaseolus vulgaris L., Xanthium strumarium L., and Scrophularia desertorum (Shaw.) Munz. O(2)-insensitive photosynthesis has been reported in nine other C(3) species and usually occurred when the intercellular CO(2) pressure was about double the normal pressure. A lack of O(2) inhibition of photosynthesis was always accompanied by a failure of increased CO(2) pressure to stimulate photosynthesis to the expected degree. O(2)-insensitive photosynthesis also occurred after plants had been water stressed. Under such conditions, however, photosynthesis became O(2) and CO(2) insensitive at physiological CO(2) pressures. Postillumination CO(2) exchange kinetics showed that O(2) and CO(2) insensitivity was not the result of elimination of photorespiration.It is proposed that O(2) and CO(2) insensitivity occurs when the concentration of phosphate in the chloroplast stroma cannot be both high enough to allow photophosphorylation and low enough to allow starch and sucrose synthesis at the rates required by the rest of the photosynthetic component processes. Under these conditions, the energy diverted to photorespiration does not adversely affect the potential for CO(2) assimilation.

Photosynthetic responses to carbon dioxide concentration can provide data on a number of important parameters related to leaf physiology. Methods for fitting a model to such data are briefly described. The method will fit the following parameters: V(cmax), J, TPU, R(d) and g(m)[maximum carboxylation rate allowed by ribulose 1.5-bisphosphate carboxylase/oxygenase (Rubisco), rate of photosynthetic electron transport (based on NADPH requirement), triose phosphate use, day respiration and mesophyll conductance, respectively]. The method requires at least five data pairs of net CO(2) assimilation (A) and [CO(2)] in the intercellular airspaces of the leaf (C(i)) and requires users to indicate the presumed limiting factor. The output is (1) calculated CO(2) partial pressure at the sites of carboxylation, C(c), (2) values for the five parameters at the measurement temperature and (3) values adjusted to 25 degrees C to facilitate comparisons. Fitting this model is a way of exploring leaf level photosynthesis. However, interpreting leaf level photosynthesis in terms of underlying biochemistry and biophysics is subject to assumptions that hold to a greater or lesser degree, a major assumption being that all parts of the leaf are behaving in the same way at each instant.

We examined the photosynthetic responses of four species of saplings growing in the understory of the Duke Forest FACE experiment during the seventh year of exposure to elevated CO2 concentration ([CO2]). Saplings of these same species were measured in the first year of the Duke Forest FACE experiment and at that time showed only seasonal fluctuations in acclimation of photosynthesis to elevated [CO2]. Based on observations from the Duke Forest FACE experiment, we hypothesized that after seven years of exposure to elevated [CO2] significant photosynthetic down-regulation would be observed in these tree species. To test our hypothesis, photosynthetic CO2-response and light-response curves, along with chlorophyll fluorescence, chlorophyll concentration and foliar N were measured twice during the summer of 2003. Exposure to elevated [CO2] continued to increase photosynthesis in all species measured after seven years of treatment with the greatest photosynthetic increase observed near saturating irradiances. In all species, elevated [CO2] increased electron transport efficiency but did not significantly alter carboxylation efficiency. Quantum yield estimated by light curves, chlorophyll concentration, and foliar nitrogen concentrations were unaffected by elevated [CO2]. Contrary to our hypothesis, there is little evidence of progressive N limitation of leaf-level processes in these understory tree species after seven years of exposure to elevated [CO2] in the Duke Forest FACE experiment.

Cyclic electron flow (CEF) around photosystem I has a role in avoiding photoinhibition of photosystem II (PSII), which occurs under conditions in which the rate of photodamage to PSII exceeds the rate of its repair. However, the molecular mechanism underlying how CEF contributes to photoprotection is not yet well understood. We examined the effect of impairment of CEF and thermal energy dissipation (qE) on photoinhibition using CEF (pgr5) and qE (npq1 and npq4) mutants of Arabidopsis (Arabidopsis thaliana) exposed to strong light. Impairment of CEF by mutation of pgr5 suppressed qE and accelerated photoinhibition. We found that impairment of qE, by mutations of pgr5, npq1, and npq4, caused inhibition of the repair of photodamaged PSII at the step of the de novo synthesis of the D1 protein. In the presence of the chloroplast protein synthesis inhibitor chloramphenicol, impairment of CEF, but not impairment of qE, accelerated photoinhibition, and a similar effect was obtained when leaves were infiltrated with the protonophore nigericin. These results suggest that CEF-dependent generation of DeltapH across the thylakoid membrane helps to alleviate photoinhibition by at least two different photoprotection mechanisms: one is linked to qE generation and prevents the inhibition of the repair of photodamaged PSII at the step of protein synthesis, and the other is independent of qE and suppresses photodamage to PSII.

A series of experiments is presented investigating short term and long term changes of the nature of the response of rate of CO2 assimilation to intercellular p(CO2). The relationships between CO2 assimilation rate and biochemical components of leaf photosynthesis, such as ribulose-bisphosphate (RuP2) carboxylase-oxygenase activity and electron transport capacity are examined and related to current theory of CO2 assimilation in leaves of C3 species. It was found that the response of the rate of CO2 assimilation to irradiance, partial pressure of O2, p(O2), and temperature was different at low and high intercellular p(CO2), suggesting that CO2 assimilation rate is governed by different processes at low and high intercellular p(CO2). In longer term changes in CO2 assimilation rate, induced by different growth conditions, the initial slope of the response of CO2 assimilation rate to intercellular p(CO2) could be correlated to in vitro measurements of RuP2 carboxylase activity. Also, CO2 assimilation rate at high p(CO2) could be correlated to in vitro measurements of electron transport rate. These results are consistent with the hypothesis that CO2 assimilation rate is limited by the RuP2 saturated rate of the RuP2 carboxylase-oxygenase at low intercellular p(CO2) and by the rate allowed by RuP2 regeneration capacity at high intercellular p(CO2).

The calculated maximum net photosynthetic rate (P N) at saturation irradiance (I m) of 1 314.13 μmol m−2 s−1 was 25.49 μmol(CO2) m−2 s−1, and intrinsic quantum yield at zero irradiance was 0.103. The results fitted by nonrectangular hyperbolic model, rectangular hyperbolic method, binomial regression method, and the new model were compared. The maximum P N values calculated by nonrectangular hyperbolic model and rectangular hyperbolic model were higher than the measured values, and the I m calculated by nonrectangular hyperbolic model and rectangular hyperbolic model were less than measured values. Results fitted by new model showed that the response curve of P N to I was nonlinear at low I for Oryza sativa, P N increased nonlinearly with I below saturation value. Above this value, P N decreased nonlinearly with I.]]>

The model couples stomatal conductance (g s) and net photosynthetic rate (P N) describing not only part of the curve up to and including saturation irradiance (I max), but also the range above the saturation irradiance. Maximum stomatal conductance (g smax) and I max can be calculated by the coupled model. For winter wheat (Triticum aestivum) the fitted results showed that maximum P N (P max) at 600 μmol mol−1 was more than at 350 μmol mol−1 under the same leaf temperature, which can not be explained by the stomatal closure at high CO2 concentration because g smax at 600 μmol mol−1 was less than at 350 μmol mol−1. The irradiance-response curves for winter wheat had similar tendency, e.g. at 25 °C and 350 μmol mol−1 both P N and g s almost synchronously reached the maximum values at about 1 600 μmol m−2 s−1. At 25 °C and 600 μmol mol−1 the I max corresponding to P max and g smax was 2 080 and 1 575 μmol m−2 s−1, respectively.]]>

BACKGROUND AND AIMS: The stomata are a key channel of the water cycle in ecosystems, and are constrained by both physiological and environmental elements. The aim of this study was to parameterize stomatal conductance by extending a previous empirical model and a revised Ball-Berry model. METHODS: Light and CO(2) responses of stomatal conductance and photosynthesis of winter wheat in the North China Plain were investigated under ambient and free-air CO(2) enrichment conditions. The photosynthetic photon flux density and CO(2) concentration ranged from 0 to 2000 micro mol m(-2) s(-1) and from 0 to 1400 micro mol mol(-1), respectively. The model was validated with data from a light, temperature and CO(2) response experiment. RESULTS: By using previously published hyperbolic equations of photosynthetic responses to light and CO(2), the number of parameters in the model was reduced. These response curves were observed diurnally with large variations of temperature and vapour pressure deficit. The model interpreted stomatal response under wide variations in environmental factors. CONCLUSIONS: Most of the model parameters, such as initial photon efficiency and maximum photosynthetic rate (P(max)), have physiological meanings. The model can be expanded to include influences of other physiological elements, such as leaf ageing and nutrient conditions, especially leaf nitrogen content.