Proton cellular influx as a probable mechanism of variation potential influence on photosynthesis in pea.

Electrical signals (action potential and variation potential, VP) caused by environmental stimuli are known to induce various physiological responses in plants, including changes in photosynthesis; however, their functional mechanisms remain unclear. In this study, the influence of VP on photosynthesis in pea (Pisum sativum L.) was investigated and the proton participation in this process analysed. VP, induced by local heating, inactivated photosynthesis and activated respiration, with the initiation of the photosynthetic response connected with inactivation of the photosynthetic dark stage; however, direct VP influence on the light stage was also probable. VP generation was accompanied with pH increases in apoplasts (0.17-0.30 pH unit) and decreases in cytoplasm (0.18-0.60 pH unit), which probably reflected H(+) -ATPase inactivation and H(+) influx during this electrical event. Imitation of H(+) influx using the protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP) induced a photosynthetic response that was similar with a VP-induced response. Experiments on chloroplast suspensions showed that decreased external pH also induced an analogous response and that its magnitude depended on the magnitude of pH change. Thus, the present results showed that proton cellular influx was the probable mechanism of VP's influence on photosynthesis in pea. Potential means of action for this influence are discussed.

Influence of variation potential on resistance of the photosynthetic machinery to heating in pea.

Electrical signals [action potentials (APs) and variation potentials (VPs)] induced by local stimuli are a mechanism that underlies rapid plant response to environmental factors. Such signals induce a number of functional responses, including changes in photosynthesis. Ultimately, these responses are considered to increase plant resistance to stress factors, but this question has been poorly investigated. We studied the influence of VP on photosynthesis and resistance of the photosynthetic machinery to heating in leaves of pea (Pisum sativum). Localized burning induced a VP that decreased photosynthesis parameters [CO(2) assimilation rate and quantum yields of photosystem I (PSI) and photosystem II (PSII)]. The photosynthetic response was initiated by a decrease in photosynthesis dark-stage activity, which in turn increased resistance of PSI to heating. Three results supported this hypothesized mechanism: (1) the magnitude of VP-induced decrease in CO(2) assimilation and enhanced PSI resistance to heating were highly correlated; (2) the VP influence on PSI resistance to heating was suppressed under a low external CO(2) concentration and (3) decreasing external CO(2) concentration imitated the VP-induced photosynthetic response and increased PSI resistance to heating.

Chlorophyll Fluorescence in Wheat Breeding for Heat and Drought Tolerance.

The constantly growing need to increase the production of agricultural products in changing climatic conditions makes it necessary to accelerate the development of new cultivars that meet the modern demands of agronomists. Currently, the breeding process includes the stages of genotyping and phenotyping to optimize the selection of promising genotypes. One of the most popular phenotypic methods is the pulse-amplitude modulated (PAM) fluorometry, due to its non-invasiveness and high information content. In this review, we focused on the opportunities of using chlorophyll fluorescence (ChlF) parameters recorded using PAM fluorometry to assess the state of plants in drought and heat stress conditions and predict the economically significant traits of wheat, as one of the most important agricultural crops, and also analyzed the relationship between the ChlF parameters and genetic markers.

Prediction of biomass accumulation and tolerance of wheat seedlings to drought and elevated temperatures using hyperspectral imaging.

Early prediction of important agricultural traits in wheat opens up broad prospects for the development of approaches to accelerate the selection of genotypes for further breeding trials. This study is devoted to the search for predictors of biomass accumulation and tolerance of wheat to abiotic stressors. Hyperspectral (HS) and chlorophyll fluorescence (ChlF) parameters were analyzed as predictors under laboratory conditions. The predictive ability of reflectance and normalized difference indices (NDIs), as well as their relationship with parameters of photosynthetic activity, which is a key process influencing organic matter production and crop yields, were analyzed. HS parameters calculated using the wavelengths in Red (R) band and the spectral range next to the red edge (FR-NIR) were found to be correlated with biomass accumulation. The same ranges showed potential for predicting wheat tolerance to elevated temperatures. The relationship of HS predictors with biomass accumulation and heat tolerance were of opposite sign. A number of ChlF parameters also showed statistically significant correlation with biomass accumulation and heat tolerance. A correlation between HS and ChlF parameters, that demonstrated potential for predicting biomass accumulation and tolerance, has been shown. No predictors of drought tolerance were found among the HS and ChlF parameters analyzed.

Comparison of the Efficiency of Hyperspectral and Pulse Amplitude Modulation Imaging Methods in Pre-Symptomatic Virus Detection in Tobacco Plants.

Early detection of pathogens can significantly reduce yield losses and improve the quality of agricultural products. This study compares the efficiency of hyperspectral (HS) imaging and pulse amplitude modulation (PAM) fluorometry to detect pathogens in plants. Reflectance spectra, normalized indices, and fluorescence parameters were studied in healthy and infected areas of leaves. Potato virus X with GFP fluorescent protein was used to assess the spread of infection throughout the plant. The study found that infection increased the reflectance of leaves in certain wavelength ranges. Analysis of the normalized reflectance indices (NRIs) revealed indices that were sensitive and insensitive to infection. NRI700/850 was optimal for virus detection; significant differences were detected on the 4th day after the virus arrived in the leaf. Maximum (Fv/Fm) and effective quantum yields of photosystem II (ΦPSII) and non-photochemical fluorescence quenching (NPQ) were almost unchanged at the early stage of infection. ΦPSII and NPQ in the transition state (a short time after actinic light was switched on) showed high sensitivity to infection. The higher sensitivity of PAM compared to HS imaging may be due to the possibility of assessing the physiological changes earlier than changes in leaf structure.

Analysis of chlorophyll fluorescence parameters as predictors of biomass accumulation and tolerance to heat and drought stress of wheat (Triticum aestivum) plants.

Agricultural technologies aimed at increasing yields require the development of highly productive and stress-tolerant cultivars. Phenotyping can significantly accelerate breeding; however, no reliable markers have been identified to select the most promising cultivars at an early stage. In this work, we determined the light-induced dynamic of chlorophyll fluorescence (ChlF) parameters in young seedlings of 10 wheat (Triticum aestivum L.) cultivars and evaluated potency of these parameters as predictors of biomass accumulation and stress tolerance. Dry matter accumulation positively correlated with the effective quantum efficiency of photosystem II (Φ PSIIef ) and negatively correlated with the half-time of Φ PSIIef reaching (t 1/2 (Φ PSIIef )). There was a highly significant correlation between t 1/2 (Φ PSIIef ) and dry matter accumulation with increasing prediction period. Short-term heating and drought caused an inhibition of biomass accumulation and photosynthetic activity depending on the stressor intensity. The positive correlation between the Φ PSII dark level (Φ PSIId ) in young seedlings and tolerance to a rapidly increasing short-term stressor (heating) was shown. In the case of a long-term stressor (drought), we revealed a strong negative relationship between tolerance and the level of non-photochemical fluorescence quenching (NPQ). In general, the results show the potency of the ChlF parameters of young seedlings as predictors of biomass accumulation and stress tolerance.

Participation of calcium ions in induction of respiratory response caused by variation potential in pea seedlings.

Electrical signals in plants caused by external stimuli are capable of inducing various physiological responses. The mechanisms of transformation of a long-distance electrical signal (ES) into a functional response remain largely unexplored and require additional research. In this work, we investigated the role of calcium ions in the development of ES-induced respiratory response. Gradual heating of the leaf causes the propagation of variation potential (VP) in the pea seedling. The propagation of VP leads to a transient activation of respiration in an unaffected leaf. During the VP generation, a transient increase in the intracellular calcium concentration takes place. A calcium channel blocker inhibits the respiratory response, and a calcium ionophore induces the activation of respiration. Inhibitory analysis has showed that the VP-induced increase in respiration activity is probably associated with calcium-mediated activation of rotenone-insensitive alternative NADPH dehydrogenases in mitochondria.

Pre-Symptomatic Detection of Viral Infection in Tobacco Leaves Using PAM Fluorometry.

Chlorophyll fluorescence imaging was used to study potato virus X (PVX) infection of Nicotiana benthamiana. Infection-induced changes in chlorophyll fluorescence parameters (quantum yield of photosystem II photochemistry (ΦPSII) and non-photochemical fluorescence quenching (NPQ)) in the non-inoculated leaf were recorded and compared with the spatial distribution of the virus detected by the fluorescence of GFP associated with the virus. We determined infection-related changes at different points of the light-induced chlorophyll fluorescence kinetics and at different days after inoculation. A slight change in the light-adapted steady-state values of ΦPSII and NPQ was observed in the infected area of the non-inoculated leaf. In contrast to the steady-state parameters, the dynamics of ΦPSII and NPQ caused by the dark-light transition in healthy and infected areas differed significantly starting from the second day after the detection of the virus in a non-inoculated leaf. The coefficients of correlation between chlorophyll fluorescence parameters and virus localization were 0.67 for ΦPSII and 0.76 for NPQ. In general, the results demonstrate the possibility of reliable pre-symptomatic detection of the spread of a viral infection using chlorophyll fluorescence imaging.

Inactivation of H+-ATPase Participates in the Influence of Variation Potential on Photosynthesis and Respiration in Peas.

Local damage (e.g., burning, heating, or crushing) causes the generation and propagation of a variation potential (VP), which is a unique electrical signal in higher plants. A VP influences numerous physiological processes, with photosynthesis and respiration being important targets. VP generation is based on transient inactivation of H+-ATPase in plasma membrane. In this work, we investigated the participation of this inactivation in the development of VP-induced photosynthetic and respiratory responses. Two- to three-week-old pea seedlings (Pisum sativum L.) and their protoplasts were investigated. Photosynthesis and respiration in intact seedlings were measured using a GFS-3000 gas analyzer, Dual-PAM-100 Pulse-Amplitude-Modulation (PAM)-fluorometer, and a Dual-PAM gas-exchange Cuvette 3010-Dual. Electrical activity was measured using extracellular electrodes. The parameters of photosynthetic light reactions in protoplasts were measured using the Dual-PAM-100; photosynthesis- and respiration-related changes in O2 exchange rate were measured using an Oxygraph Plus System. We found that preliminary changes in the activity of H+-ATPase in the plasma membrane (its inactivation by sodium orthovanadate or activation by fusicoccin) influenced the amplitudes and magnitudes of VP-induced photosynthetic and respiratory responses in intact seedlings. Decreases in H+-ATPase activity (sodium orthovanadate treatment) induced fast decreases in photosynthetic activity and increases in respiration in protoplasts. Thus, our results support the effect of H+-ATPase inactivation on VP-induced photosynthetic and respiratory responses.

Exogenous Abscisic Acid Can Influence Photosynthetic Processes in Peas through a Decrease in Activity of H+-ATP-ase in the Plasma Membrane.

Abscisic acid (ABA) is an important hormone in plants that participates in their acclimation to the action of stressors. Treatment by exogenous ABA and its synthetic analogs are a potential way of controlling the tolerance of agricultural plants; however, the mechanisms of influence of the ABA treatment on photosynthetic processes require further investigations. The aim of our work was to investigate the participation of inactivation of the plasma membrane H+-ATP-ase on the influence of ABA treatment on photosynthetic processes and their regulation by electrical signals in peas. The ABA treatment of seedlings was performed by spraying them with aqueous solutions (10-5 M). The combination of a Dual-PAM-100 PAM fluorometer and GFS-3000 infrared gas analyzer was used for photosynthetic measurements; the patch clamp system on the basis of a SliceScope Pro 2000 microscope was used for measurements of electrical activity. It was shown that the ABA treatment stimulated the cyclic electron flow around photosystem I and decreased the photosynthetic CO2 assimilation, the amplitude of burning-induced electrical signals (variation potentials), and the magnitude of photosynthetic responses relating to these signals; in contrast, treatment with exogenous ABA increased the heat tolerance of photosynthesis. An investigation of the influence of ABA treatment on the metabolic component of the resting potential showed that this treatment decreased the activity of the H+-ATP-ase in the plasma membrane. Inhibitor analysis using sodium orthovanadate demonstrated that this decrease may be a mechanism of the ABA treatment-induced changes in photosynthetic processes, their heat tolerance, and regulation by electrical signals.

Variation potential propagation decreases heat-related damage of pea photosystem I by 2 different pathways.

Local burning is known to generate and propagate variation potential (VP) in plants. VP affects different physiological processes, including reducing heat-related damage to photosystem I (PSI). We investigated mechanisms of the process. Photosynthesis parameters were measured with Dual-PAM-100 and GFS-3000. VP was induced by burning the first mature leaf and then waiting 5, 10, 15, or 20 min to initiate heating of the second mature leaf. Photosystems activities in the second leaf were investigated at 15 and 135 min after heating. In the absence of VP induction, when incubation in hot water (5 min) was used for heating the intact second leaf, PSI and PSII activities decreased after incubation at both exposure temperatures (45°C and 50°C). When local burning of the first leaf induced VP propagation into the second leaf, reduced photosynthesis (PSI) was observed. Arrival of VP in the second leaf prior to hot water incubation at 50°C decreased heating-induced suppression of PSI activity when measured 15 and 135 min later. Dependence of PSI activity on the time interval (5, 10, 15, or 20 min) between VP induction and heating of the second leaf was dissimilar at 15 and 135 min. Heat-induced suppression of PSII activity in the second leaf was stimulated after VP induction. In contrast, the effect of VP on PSI and PSII damage was weak when leaf 2 was heated at 45°C. VP-induced decrease of PSI activity suppression at 15 min after heating was correlated with stimulation of PSII activity suppression, but increase of PSI activity at 135 min after heating was not related to PSII activity. Thus, our results suggest the possibility of 2 different pathways of VP-induced decrease of heat-related PSI damage.

Variation potential-induced photosynthetic and respiratory changes increase ATP content in pea leaves.

Local damage induces a physiological response in higher plants by means of generation and propagation of variation potential (VP). The response includes changes in photosynthesis and respiration. The aim of the present study was to investigate the effect of these changes on adenosine triphosphate (ATP) content in pea leaves. VP was induced by local heating of the first mature leaf and registered using extracellular and intracellular electrodes. Photosynthesis and respiration were measured using Dual-PAM-100 and GFS-3000. ATP content was determined using a bioluminescence-based ATP determination kit. Two non-stimulated leaves (second and fourth) were investigated. We showed that heating induced VP that propagated into the second mature leaf, but only a slight electrical reaction was registered in the fourth mature leaf. VP-induced inactivation of photosynthesis developed in the second leaf and included two stages: short- and long-term inactivation. Local heating also caused a two-stage increase in ATP content in the second leaf, which was connected with the photosynthetic responses. Changes in photosynthesis and ATP content were not observed in the fourth leaf. The effect of VP on respiration was investigated under dark conditions. We found that variation potential induced short-term activation of respiration in the second leaf. Local heating induced ATP content increase which included only one stage under dark conditions. Changes in ATP and respiration were absent in the fourth leaf under dark conditions. Thus, VP-induced photosynthetic and respiratory changes are likely to increase ATP content in pea leaves.

Changes in H(+)-ATP Synthase Activity, Proton Electrochemical Gradient, and pH in Pea Chloroplast Can Be Connected with Variation Potential.

Local stimulation induces generation and propagation of electrical signals, including the variation potential (VP) and action potential, in plants. Burning-induced VP changes the physiological state of plants; specifically, it inactivates photosynthesis. However, the mechanisms that decrease photosynthesis are poorly understood. We investigated these mechanisms by measuring VP-connected systemic changes in CO2 assimilation, parameters of light reactions of photosynthesis, electrochromic pigment absorbance shifts, and light scattering. We reveal that inactivation of photosynthesis in the pea, including inactivation of dark and light reactions, was connected with the VP. Inactivation of dark reactions decreased the rate constant of the fast relaxation of the electrochromic pigment absorbance shift, which reflected a decrease in the H(+)-ATP synthase activity. This decrease likely contributed to the acidification of the chloroplast lumen, which developed after VP induction. However, VP-connected decrease of the proton motive force across the thylakoid membrane, possibly, reflected a decreased pH in the stroma. This decrease may be another mechanism of chloroplast lumen acidification. Overall, stroma acidification can decrease electron flow through photosystem I, and lumen acidification induces growth of fluorescence non-photochemical quenching and decreases electron flow through photosystem II, i.e., pH decreases in the stroma and lumen, possibly, contribute to the VP-induced inactivation of light reactions of photosynthesis.

Variation potential induces decreased PSI damage and increased PSII damage under high external temperatures in pea.

Local burning of a leaf induces a unique electrical signal in plants: variation potential (VP), which can cause numerous functional responses, including changes in photosynthesis. The influences of VP on damage to photosynthetic machinery and thermal resistance of whole plant with heating were investigated in pea (Pisum sativum L.). Under high external temperature (53°C), VP induction was found to cause accelerated initial lowering of photosynthetic parameters, reduced transpiration rate, decreased PSI damage, and increased PSII damage. Increased PSI resistance was in good accordance with our previous data, but VP-induced stimulation of PSII damage under heating required further analysis. The magnitudes of leaf heating in experiments and controls were found to be different (~46-48°C after VP and ~44-45°C without it) under identical external temperatures and were connected with different transpiration rates under these conditions. Also PSII damage and the magnitude of leaf heating were strongly correlated. As a result, it was supposed that leaf temperature contributed to VP-induced stimulation of PSII damage with heating. Investigation of VP's influence on pea growth after heating showed that the electrical signal decreased plant growth suppression after heating (i.e. VP-induced increases in PSII damage did not noticeably influence thermal resistance in the whole plant).

Variation potential influence on photosynthetic cyclic electron flow in pea.

Cyclic electron flow is an important component of the total photosynthetic electron flow and participates in adaptation to the action of stressors. Local leaf stimulation induces electrical signals, including variation potential (VP), which inactivate photosynthesis; however, their influence on cyclic electron flow has not been investigated. The aim of this study was to investigate VP's influence on cyclic electron flow in pea (Pisum sativum L.). VP was induced in pea seedling leaves by local heating and measured in an adjacent, undamaged leaf by extracellular electrodes. CO2 assimilation was measured using a portable gas exchange measuring system. Photosystem I and II parameters were investigated using a measuring system for simultaneous assessment of P700 oxidation and chlorophyll fluorescence. Heating-induced VP reduced CO2 assimilation and electron flow through photosystem II. In response, cyclic electron flow rapidly decreased and subsequently slowly increased. Slow increases in cyclic flow were caused by decreased electron flow through photosystem II, which was mainly connected with VP-induced photosynthetic dark stage inactivation. However, direct influence by VP on photosystem I also participated in activation of cyclic electron flow. Thus, VP, induced by local leaf-heating, activated cyclic electron flow in undamaged leaves. This response was similar to photosynthetic changes observed under the direct action of stressors. Possible mechanisms of VP's influence on cyclic flow were discussed.