Respiration In Protoplasts Research Articles

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.

Dark Respiration Protects Photosynthesis Against Photoinhibition in Mesophyll Protoplasts of Pea (Pisum sativum)

The optimal light intensity required for photosynthesis by mesophyll protoplasts of pea (Pisum sativum) is about 1250 microeinsteins per square meter per second. On exposure to supra-optimal light intensity (2500 microeinsteins per square meter per second) for 10 min, the protoplasts lost 30 to 40% of their photosynthetic capacity. Illumination with normal light intensity (1250 microeinsteins per square meter per second) for 10 min enhanced the rate of dark respiration in protoplasts. On the other hand, when protoplasts were exposed to photoinhibitory light, their dark respiration also was markedly reduced along with photosynthesis. The extent of photoinhibition was increased when protoplasts were incubated with even low concentrations of classic respiratory inhibitors: 1 micromolar antimycin A, 1 micromolar sodium azide, and 1 microgram per milliliter oligomycin. At these concentrations, the test inhibitors had very little or no effect directly on the process of photosynthetic oxygen evolution. The promotion of photoinhibition by inhibitors of oxidative electron transport (antimycin A, sodium azide) and phosphorylation (oligomycin) was much more pronounced than that by inhibitors of glycolysis and tricarboxylic acid cycle (sodium fluoride and sodium malonate, respectively). We suggest that the oxidative electron transport and phosphorylation in mitochondria play an important role in protecting the protoplasts against photoinhibition of photosynthesis. Our results also demonstrate that protoplasts offer an additional experimental system for studies on photoinhibition.

Beneficial interaction between photosynthesis and respiration in mesophyll protoplasts of pea during short light‐dark cycles

The respiratory uptake or photosynthetic evolution of oxygen by mesophyll protoplasts of pea (Pisum sativum L. cv. Arkel) were monitored during successive short. (3–5 min) cycles of darkness and illumination. The rate of respiration was nearly doubled after 3–4 short periods of illumination while there was a 15–20% enhancement in photosynthesis with cycles of illumination and darkness preceding illumination. Such interaction between photosynthesis and respiration was statistically significant when bicarbonate was present in the reaction medium. The inhibitors of photosynthesis [3(3,4–dichlorophenyl)‐l,l‐dimethylurea (DCMU), glyceraldehyde] decreased respiration after periods of illumination, whereas inhibitors of respiratory electron transport (Rotenone, antimycin A, NaN3) suppressed photosynthesis, as well. We suggest that a rapid beneficial interaction exists between photosynthesis and respiration in protoplasts, even during short cycles of light and darkness.

The respiration of protoplasts from leaves of barley infected by powdery mildew ( Erysiphe graminis F.SP. Hordei)

Infection of barley leaves by powdery mildew ( Erysiphe graminis f.sp. hordei) causes increased dark respiration, par tof which is associated with active host responses to infection, and a consequence of which is reduced plant growth. The pathogen cannot be grown separately from the host. Therefore, in order to examine those changes in respiratory activity peculiar to the host, attempts were made to isolate protoplasts from infected tissues, and from healthy controls. Isolation of useful numbers ( > 10 6 cm −6 ) of viable mesophyll protoplasts from infected tissues was possible with one among several batches of commercial ‘Cellulysin’ tested; on analysis by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), this batch contained a low molecular weight protein at 20.1 kDa not found in other batches. In all isolated protoplasts, total respiration increased with the age of the source-leaf, but within 24 h of inoculation respiration was stimulated by infection. Protoplasts from infected leaves showed a small initial increase in the cyanide (CN)-sensitive component (cytochrome pathway) of total respiration, but an increase in the CN-insensitive component (alternative pathway) was the major cause of the stimulation of total respiration.

FACTORS INFLUENCING RESPIRATORY MEASUREMENTS USING OXYGEN ELECTRODES AND CARTESIAN‐DIVER MICRORESPIROMETRY

SummaryThe dark respiration rates of primary leaf mesophyll protoplasts of barley (Hordeum vulgare L. cv. Patty) were measured by a Clark‐type oxygen electrode and Cartesian‐diver microrespirometry, in order to compare the two techniques. Factors found to be important in determinations of respiratory rates of protoplasts were the numbers of protoplasts per unit volume of suspension, the age of protoplast preparations and the stirring speed of the magnetic follower when using the oxygen electrode. Increases in numbers of protoplasts per unit volume caused a marked decrease in respiration using both techniques. Below 2000 protoplasts μl−1 both techniques provided rates of dark respiration within the same range until 1000 protoplasts μl−1, below which a marked increase in respiration occurred. Respiratory rates of protoplasts declined by 32 % from initial values over a 30 h period after isolation in the dark. In addition, an increase in the stirring rate in the reaction vessel of the oxygen electrode had the effect of reducing both the viability of protoplasts and their rates of oxygen uptake. It is suggested that consistent standardization of procedures, especially in terms of numbers of protoplasts per unit volume, is necessary for assay techniques for respiration in protoplasts. The advantages and disadvantages of using Cartesian‐diver microrespirometry compared to oxygen electrodes are also discussed.

Inhibition of Respiration in Protoplasts from Meristematic Tissues3

Inhibition of Respiration in Protoplasts from Meristematic Tissues3

Calcium, Calmodulin and the Control of Respiration in Protoplasts Isolated from Meristematic Tissues8

Calcium, Calmodulin and the Control of Respiration in Protoplasts Isolated from Meristematic Tissues8

Influences of Light Quality, Growth Regulators and Inhibitors of Protein Synthesis on Respiration of Protoplasts from Meristematic Cells in Barley Seedlings

Influences of Light Quality, Growth Regulators and Inhibitors of Protein Synthesis on Respiration of Protoplasts from Meristematic Cells in Barley Seedlings