Ageing of chloroplasts in vitro — III. Comparison of the effects of ageing in vitro and senescence on the red absorption band and primary photochemical reactions of sunflower chloroplasts

A comparison of changes in absorption properties and electron transport activities of chloroplasts ageing in vivo and in vitro is made. Chloroplasts from sunflower leaves senescing in vivo during 7 days in dark do not show a blue shift of the red absorption band; in contrast, the shift becomes apparent within 24 h of in vitro ageing of isolated organelles. Photosynthetic activity by chloroplasts is lost much faster during in vitro than in vivo ageing. During in vitro ageing, the rate of degradation of thylakoid membranes as characterised by the shift in the red absorption band and loss in Hill reaction is further accelerated in chloroplasts isolated from dark-induced senescing leaves, suggesting the influence of the in vivo status of the chloroplasts on their in vitro stability.Abbreviations DCPIP 2,6-dichlorophenol indophenol - PSI Photosystem I - Chl Chlorophyll     

Ageing of chloroplasts in vitro — III. Comparison of the effects of ageing in vitro and senescence on the red absorption band and primary photochemical reactions of sunflower chloroplasts

A comparison of changes in absorption properties and electron transport activities of chloroplasts ageing in vivo and in vitro is made. Chloroplasts from sunflower leaves senescing in vivo during 7 days in dark do not show a blue shift of the red absorption band; in contrast, the shift becomes apparent within 24 h of in vitro ageing of isolated organelles. Photosynthetic activity by chloroplasts is lost much faster during in vitro than in vivo ageing. During in vitro ageing, the rate of degradation of thylakoid membranes as characterised by the shift in the red absorption band and loss in Hill reaction is further accelerated in chloroplasts isolated from dark-induced senescing leaves, suggesting the influence of the in vivo status of the chloroplasts on their in vitro stability.Abbreviations DCPIP 2,6-dichlorophenol indophenol - PSI Photosystem I - Chl+ Chlorophyll     

Photoprotective effect of kinetin on pigment content and photochemical activities of wheat chloroplasts agingin vitro

The effects of kinetin (Kn) on pigment content and electron transport activities (ETA) in wheat leavesin vivo and chloroplastsin vitro aging in light was investigated. Excised wheat leaves were infiltrated with Kn for 3 h under irradiation. The treatment increased zeaxanthin (Zx) content by 40% and also increased chlorophyll (Chia, Chib) and major carotenoid (Car) contents in the leaves (per fresh mass unit). Chloroplasts isolated from Kn treated leaves, when incubated in light for 4 h showed relatively lower pigment loss and slower loss of ETA compared to the chloroplasts of untreated leaves. These observations suggest photoprotective action of Kn. The photoprotection was more prominent when Kn was applied directly to the irradiated chloroplastsin vitro. Moreover, chloroplasts agingin vitro under irradiation without Kn treatment lost pigments and ETA. Within 3 h of irradiation, both whole chain (H₂O to methylviologen) electron transport as well as photosystem (PS) 2 activity were completely lost. However, in the chloroplasts treated with Kn, the loss of pigments was slow and even after 4 h of irradiation the chloroplasts retained 15 % of PS 2 and 9 % of whole chain ETA. In the untreated chloroplasts, the loss of Zx after 4 h of irradiation was 49 % whereas in Kn treated samples its level was 1.3 times higher than that of control. Since a higher level of Zx was maintained in Kn treated chloroplasts, photoprotective action of Kn is possibly mediated through Zx. One of us (NKC) thanks Sambalpur University for study leave and Department of Biological Sciences, Mankato State University, Mankato for labortory facilities.     

Two molecular forms of pea ferredoxin in the electron transport chain of chloroplasts

The effects of two molecular forms of water-soluble ferredoxin (Fd I and Fd II) on the kinetics of electron transport in bean chloroplasts (class B) were studied. The light-induced redox transitions of the photosystem I reaction center P700 were measured by the intensity of the EPR signal I produced by P700+. Both forms of ferredoxin, Fd I and Fd II, when added to the chloroplasts in catalytic amounts, stimulate the light-induced electron transfer from P700 to NADP+. Nevertheless, Fd I is a better mediator of the back reactions from NADPH to P700+. This electron transfer pathway is sensitive to the cyclic electron transport inhibitor, antimycin A, and to DCMU inhibitor of electron transport between photosystem II and plastoquinone. It may be concluded that the two molecular forms of ferredoxin, Fd I and Fd II, differ in their ability to catalyze cyclic electron transport in photosystem I. The role of Fd I and Fd II in regulation of electron transport at the acceptor site of photosystem I is discussed.     

Bioenergetics in aging: mitochondrial proton leak in aging rat liver, kidney and heart

Aging is associated with a decline in performance in many organs and loss of physiological performance can be due to free radicals. Mitochondria are incompletely coupled: during oxidative phosphorylation some of the redox energy is dissipated as natural proton leak across the inner membrane. To verify whether proton leak occurs in mitochondria during aging, we measured the mitochondrial respiratory chain activity, membrane potential and proton leak in liver, kidneys and heart of young and old rats. Mitochondria from old rats showed normal rates of Complex I and Complex II respiration. However, they had a lower membrane potential compared to mitochondria from younger rats. In addition, they exhibited an increased rate of proton conductance which partially dissipated the mitochondrial membrane potential when the rate of electron transport was suppressed. This could compromise energy homeostasis in aging cells in conditions that require additional energy supply and could minimize oxidative damage to DNA.     

Interaction between chloroplast and cytoplasmic factors in the inhibition of l-phenylalanine ammonia-lyase activity

Components of the system inactivating cytoplasmic l -phenylalanine ammonia-lyase (PAL; EC 4.3.1.5.) in barley seedlings ( Hordeiim vulgare L. cv. Polon) were investigated. In vitro light-activated chloroplasts and two cytoplasmic factors participate in the enzyme inhibition. One role of the chloroplasts is to mediate the reaction between PAL and the cytoplasmic factors, resulting in inactivation of the enzyme. Uncoupling (carbonyl cyanide m-chlorophenyl hydrazone, valinomycin) or inhibiting [3-(3,4-dichlorophenyl)-l,l-dimethylurea] photosynthetic electron transport showed that electron transport is necessary to enable chloroplasts to mediate PAL inhibition. Preliminary results also indicate that mitochondria are able to mediate the inhibition. The role of this inhibiting system in the regulation of PAL activity in vivo is discussed.     

Electron transport pathways in spinach chloroplasts. Reduction of the primary acceptor of photosystem II by reduced nicotinamide adenine dinucleotide phosphate in the dark.

Addition of NADPH to osmotically lysed spinach chloroplasts results in a reduction of the primary acceptor (Q) of photosystem II. This reduction of Q reaches a maximum of 50% in chloroplasts maintained under weak illumination and requires added ferredoxin and Mg2+. The reaction is inhibited by (I) an antibody to ferredoxin-NADP+ reductases (EC 1.6.7.1), (ii) treatment of chloroplasts with N-ethylmaleimide in the presence of NADPH, (iii) disulfodisalicylidenepropanediamine, (iv) antimycin, and (v) acceptors of non-cyclic electron transport. Uncouplers of phosphorylation do not affect NADPH-driven reduction of Q. It is proposed that electron flow from NADPH to Q may occur in the dark by a pathway utilising portions of the normal cyclic and non-cyclic electron carrier sequences. The possible in vivo role for such a pathway in redox poising of cyclic electron transport and hence in controlling the ATP/NADPH supply ratio is discussed.     

Characterization of factors affecting the activity of photosystem I cyclic electron transport in chloroplasts

PSI cyclic electron transport is essential for photosynthesis and photoprotection. In higher plants, the antimycin A-sensitive pathway is the main route of electrons in PSI cyclic electron transport. Although a small thylakoid protein, PGR5 (PROTON GRADIENT REGULATION 5), is essential for this pathway, its function is still unclear, and there are numerous debates on the rate of electron transport in vivo and its regulation. To assess how PGR5-dependent PSI cyclic electron transport is regulated in vivo, we characterized its activity in ruptured chloroplasts isolated from Arabidopsis thaliana. The activity of ferredoxin (Fd)-dependent plastoquinone (PQ) reduction in the dark is impaired in the pgr5 mutant. Alkalinization of the reaction medium enhanced the activity of Fd-dependent PQ reduction in the wild type. Even weak actinic light (AL) illumination also markedly activated PGR5-dependent PSI cyclic electron transport in ruptured chloroplasts. Even in the presence of linear electron transport [11 mumol O2 (mg Chl)(-1) h(-1)], PGR5-dependent PSI electron transport was detected as a difference in Chl fluorescence levels in ruptured chloroplasts. In the wild type, PGR5-dependent PSI cyclic electron transport competed with NADP+ photoreduction. These results suggest that the rate of PGR5-dependent PSI cyclic electron transport is high enough to balance the production ratio of ATP and NADPH during steady-state photosynthesis, consistently with the pgr5 mutant phenotype. Our results also suggest that the activity of PGR5-dependent PSI cyclic electron transport is regulated by the redox state of the NADPH pool.     

Photosynthetic Electron Transport During Senescence of the Primary Leaves of Phaseolus vulgaris L.: II. THE ACTIVITY OF PHOTOSYSTEMS ONE AND TWO, AND A NOTE ON THE SITE OF REDUCTION OF FERRICYANIDE

The activity of photosystems one and two (PS I and PS II) wasmeasured in chloroplasts isolated from the primary leaves ofPhaseolus vulgaris. During foliar senescence, the rates of electrontransport through PS I and PS II declined by approximately 25%and 33% respectively. These losses of activity could not accountfor the decrease of 80% in the rate of coupled, non-cyclic electrontransport during senescence. It is therefore suggested thatan impairment of electron flow between the photosystems limitednon-cyclic electron transport in chloroplasts from older leaves.In this study the activity of PS II was measured using oxidizedp-phenylenediamine as the electron acceptor, and trifluralinas an inhibitor of electron transport between PS II and PS I.In chloroplasts from young leaves the reduction of ferricyanidewas a measure of non-cyclic electron transport, but in preparationsfrom older leaves ferricyanide received a large proportion ofelectrons from PS II.     

Structural and functional stability of isolated intact chloroplasts

The effect of in vitro ageing on the ultrastructure, electron transport, thermoluminescence and flash-induced 515 nm absorbance change of isolated intact (type A) chloroplasts compared with non-intact (types B and C) chloroplasts was studied.When stored in the dark for 18 h at 5°C, the structural characteristics of intact and non-intact chloroplasts were only slightly altered. The most conspicuous difference between the two was in the coupling of the electron transport which was tighter and more stable in intact chloroplasts. Under dark-storage the activity of PS 2* decreased and the -20°C peak of thermoluminescence increased at the expense of the emission at +25°C. These changes were less pronounced in the intact chloroplasts. PS 1 activity and the flash-induced 515 nm absorbance change were not affected by dark-storage.When kept in the light (80 W m^-2 (400–700 nm) for 1 h at 5°C), the thylakoid system of chloroplasts rapidly became disorganized. Although the initial activity of electron transport was much higher in intact chloroplasts, after a short period of light-storage the linear electron transport and the electron transport around PS 2 decreased in both types of preparations to the same low level. These changes were accompanied by an overall decrease of the intensity of thermoluminescence. PS 1 was not inhibited by light-storage, while the flash-induced 515 nm absorbance change was virtually abolished both in preparations of intact and non-intact chloroplasts.The data show that in stored chloroplast preparations intactness cannot be estimated reliably either by the FeCy test or by inspection under the electron microscope. These tests should be cross-checked on the level and coupling of the electron transport.     

The coleoptile chloroplast: Distinct distribution of xanthophyll cycle pigments,and enrichment in Photosystem II

Recent studies have shown that coleoptile chloroplasts operate the xanthophyll cycle, and that their zeaxanthin concentration co-varies with their sensitivity to blue light. The present study characterized the distribution of photosynthetic pigments in thylakoid pigment–protein complexes from dark-adapted and light-treated coleoptile and mesophyll chloroplasts, the low temperature fluorescence emission spectra, and the rates of PS I and PS II electron transport in both types of chloroplasts from 5-day-old corn seedlings. Pigments were extracted from isolated PS I holocomplex, LHC IIb trimeric and LHC II monomeric complexes and analyzed by HPLC. Chlorophyll distribution in coleoptile thylakoids showed 31% of the total collected Chl in PS I and 65% in the light harvesting complexes of PS II. In mesophyll thylakoids, the values were 44% and 54%, respectively. Mesophyll and coleoptile PS I holocomplexes differed in their Chl t a/Chl t b ratios (8.1 and 6.1, respectively) and -carotene content. In contrast, mesophyll and coleoptile LHC IIb trimers and LHC II monomers had similar Chl t a/Chl t b ratios and -carotene content. The three analyzed pigment–protein complexes from dark-adapted coleoptile chloroplasts contained zeaxanthin, whereas there was no detectable zeaxanthin in the complexes from dark-adapted mesophyll chloroplasts. In both chloroplast types, zeaxanthin and antheraxanthin increased markedly in the three pigment–protein complexes upon illumination, while violaxanthin decreased. In mesophyll thylakoids, zeaxanthin distribution as a percentage of the xanthophyll cycle pool was: LHC II monomers > LHC IIb trimers > PS I holocomplex, and in coleoptile thylakoids, it was: LHC IIb trimers > LHC II monomers = PS I holocomplex. Low temperature (77 K) fluorescence emission spectra showed that the 686 nm emission of coleoptile chloroplasts was approximately 50% larger than that of mesophyll chloroplasts when normalized at 734 nm. The pigment and fluorescence analysis data suggest that there is relatively more PS II per PS I and more LHC I per CC I in coleoptile chloroplasts than in mesophyll chloroplasts. Measurements of t in vitro uncoupled photosynthetic electron transport showed approximately 60% higher rates of electron flow through PS II in coleoptile chloroplasts than in mesophyll chloroplasts. Electron transport rates through PS I were similar in both chloroplast types. Thus, when compared to mesophyll chloroplasts, coleoptile chloroplasts have a distinct PS I pigment composition, a distinct chlorophyll distribution between PS I and PS II, a distinct zeaxanthin percentage distribution among thylakoid pigment–protein complexes, a higher PS II-related fluorescence emission, and higher PS II electron transport capacity. These characteristics may be associated with a sensory transducing role of coleoptile chloroplasts.     

Adaptation of photosynthesis under iron deficiency in maize

This paper explores the effects of high light stress on Fe-deficient plants. Maize (Zea mays) plants were grown under conditions of Fe deficiency and complete nutrition. Attached, intact leaves of Fe-deficient and control plants were used for gas exchange experiments under suboptimal, optimal and photoinhibitory illumination. Isolated chloroplasts were used to study photosynthetic electron transport system, compromised by the induction of Fe deficiency. The reaction centers of PS II (measured as reduction of Q, the primary electron acceptor of P 680) and PS I (measured as oxidation of P 700) were estimated from the amplitude of light induced absorbance change at 320 and 700 nm, respectively. Plants were subjected to photoinhibitory treatment for different time periods and isolated chloroplasts from these plants were used for electron transport studies. Carbon dioxide fixation in control as well as in Fe-deficient plants decreased in response to high light intensities. Total chlorophyll, P 700 and Q content in Fe-deficient chloroplasts decreased, while Chl a/b ratio and Q/P 700 ratio increased. However, electron transport through PS II suffered more after photoinhibitory treatment as compared to electron transport through PS I or whole chain. Electron transfer through PS I+PS II, excluding the water oxidation complex showed a decrease in Fe-deficient plants. However, electron transport through this part of the chain did not suffer much as a result of photoinhibition, suggesting a defect in the oxidising side of PS II.     

Effect of salts and electron transport on the conformation of isolated chloroplasts. I. Light-scattering and volume changes

Whole chloroplasts isolated from the leaves of spinach (Spinacia oleracea L.) exhibit 2 types of conformational change during electron transport. Amine-uncoupled chloroplasts swell and atebrin-uncoupled chloroplasts shrink. Chloroplasts uncoupled by carbonylcyanide phenylhydrazones and by treatment with ethylenediamine tetraacetic acid do not change their volumes or light-scattering properties during electron transport. Phosphorylating chloroplasts shrink only slightly.The rate and extent of the conformational change parallel the rate of electron transport; both the decrease in turbidity with methylamine and the increase in turbidity with atebrin are rougly proportional to the Hill reaction rate. Consequently the great volume and light-scattering changes which occur in the presence of these uncouplers can be attributed, in part, to the very high rates of uncoupled electron transport. However, for a given rate of electron transport the atebrin-induced scattering increase is very much greater than the increase observed during photophosphorylation.When uncouplers are combined, the carbonylcyanide phenylhydrazone effect (no change) supercedes both the methylamine effect (swelling) and the atebrin effect (shrinking). The methylamine effect supercedes the atebrin (shrinking) and ethylenediamine tetracetic acid (no change) effects. The atebrin effect supercedes the ethylenediamine tetraacetic acid effect. A similar hierarchy of effects is observed with regard to the rate of the uncoupled electron transport.These light-scattering changes of whole chloroplasts reflect similar changes which occur in very small digitonin particles of chloroplasts. Therefore one must look among chloroplast substructures for the basic mechanism of swelling and shrinking.Many salts (including methylamine hydrochloride) cause the chloroplasts to shrink. This phenomenon is not osmotic since comparable osmolarities of sucrose are without effect. Magnesium chloride and calcium chloride are most effective but all salts tested gave major volume decrease when less than 0.05 m. The salt-shrunken chloroplasts show greater light-scattering changes during electron transport than do low-salt chloroplasts.     

Changes in the Ability of Photophosphorylation and Activities of Surface-Bound Adenosine Triphosphatase and Ribulose Diphosphate Carboxylase of Chloroplasts Isolated from the Barley Leaves Senescing in Darkness

Dark-induced aging of detached primary leaves of 11-day-old barley seedlings brings about a significant decline in the rates of ferricyanide [Fe(CN)₆]^3? reduction and photophosphorylations of isolated chloroplasts. Ferricyanide-supported noncyclic photophosphorylation is somewhat more susceptible to leaf aging than phenazine methosulfate (PMS)-supported cyclic phosphorylation. Non-latent membrane-bound adenosine triphosphatase (ATPase) and ribulosediphosphate carboxylase (RuDPCase) lose about half of their initial activities after 24 h, while during this period the electron transport and photophosphorylation activities are much less affected. Also, the loss of RuDPCase is almost complete, while chloroplasts still exhibit a significant level of [Fe(CN)₆]^3? reduction and photophosphorylations after 7 days of dark incubation. This would suggest that the enzymatic dark reactions are more sensitive to aging stress than the primary photochemical reactions of chloroplasts. Studies on the effect of divalent cations such as Mg²⁺ and Ca²⁺ on non-latent ATPase activity revealed that the dark stressed aging of detached leaves brings about a time dependent alteration in the response of this enzyme to Mg²⁺, but not to Ca²⁺. The former showed inhibitory as well as stimulatory response, whereas the latter always caused the usual stimulation. Addition of kinetin (50 μM) ensured retention of [Fe(CN)₆]^3? reduction, photophosphorylations and RuDPCase activity in chloroplasts during leaf aging, but it failed to preserve the initial loss in the activity of the non-activated membrane-bound ATPase.     

Cyclic and noncyclic photophosphorylation during the ontogenesis of high-light and low-light leaves of Sinapis alba

Noncyclic electron transport to ferricyanide and photophosphorylation as well as the methylviologen mediated aerobic and anaerobic photophosphorylation with dichlorophenolindophenol-ascorbate as the electron donor of photosystem I were measured during the development of high-light and low-light adapted leaves of Sinapis alba. Anaerobic methylviologen-catalyzed phosphorylation is more than twice as high as aerobic phosphorylation. The difference between the rates of aerobic and anaerobic phosphorylation is sensitive to dibromothymoquinone. Thus, under anaerobic conditions, methylviologen mediates a cyclic phosphorylation including plastoquinone. All photochemical activities of high-light chloroplasts are about twice as high as that of low-light chloroplasts and show a permanent decline with increasing plant age. The lower activities of low-light chloroplasts correlate with a decrease of electron transport components, such as cytochrome f. This indicates that the number of electron transport chains is decreased under low-light conditions and more chlorophyll molecules interact with one electrontransport chain.Abbreviations Asc ascorbate - Chl chlorophyll a+b - DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - DCMU 3-(dichlorophenyl)-1,1-dimethylurea - DCPIP dichlorophenolindophenol - HL high light - LL low light - MV methylviologen - PhAR photosynthetically active radiation - PS photosystem     

Plant aging increases oxidative stress in chloroplasts

Aging has received considerable attention in biomedicine, but little is known about the regulatory mechanisms responsible for the aging not associated with senescence in plants. This study provides new insights into the relationship between oxidative stress and plant aging, and points out chloroplasts as one of the target organelles of age-associated oxidative stress in plants. We simultaneously analyzed lipid oxidation, photosynthesis, chlorophyll content, de-epoxidation state of the xanthophyll cycle, and levels of chloroplastic antioxidant defenses such as beta-carotene and alpha-tocopherol in leaves of the same age in 1-, 3- and 7-year-old Cistus clusii Dunal plants growing under Mediterranean field conditions. Enhanced formation of malondialdehyde in leaves (2.7-fold) and chloroplasts (2.8-fold), decreased photosynthetic activity (25%), and lower chlorophyll (ca. 20%) and chloroplastic antioxidant defense levels (ca. 25%-85%) were observed in 7-year-old plants, when compared with 1- and 3-year-old plants. The differences observed, which were associated with plant aging, were only noticeable in mature nonsenescing plants (7-year-old plants). No differences were observed between pre-reproductive (1-year-old plants) and young plants (3-year-old plants). This study shows that from a certain age, oxidative stress increases progressively in chloroplasts as plants age, whereas photosynthesis is reduced. The results indicate that the oxidative stress associated with the aging in plants accumulates progressively in chloroplasts, and that the contribution of oxidative stress to aging increases as plants age.     

A balanced PGR5 level is required for chloroplast development and optimum operation of cyclic electron transport around photosystem I

PSI cyclic electron transport contributes markedly to photosynthesis and photoprotection in flowering plants. Although the thylakoid protein PGR5 (Proton Gradient Regulation 5) has been shown to be essential for the main route of PSI cyclic electron transport, its exact function remains unclear. In transgenic Arabidopsis plants overaccumulating PGR5 in the thylakoid membrane, chloroplast development was delayed, especially in the cotyledons. Although photosynthetic electron transport was not affected during steady-state photosynthesis, a high level of non-photochemical quenching (NPQ) was transiently induced after a shift of light conditions. This phenotype was explained by elevated activity of PSI cyclic electron transport, which was monitored in an in vitro system using ruptured chloroplasts, and also in leaves. The effect of overaccumulation of PGR5 was specific to the antimycin A-sensitive pathway of PSI cyclic electron transport but not to the NAD(P)H dehydrogenase (NDH) pathway. We propose that a balanced PGR5 level is required for efficient regulation of the rate of antimycin A-sensitive PSI cyclic electron transport, although the rate of PSI cyclic electron transport is probably also regulated by other factors during steady-state photosynthesis.     

Correlation between thylakoid stacking and cation-induced decrease in photosystem I electron transport at saturating light intensity

A Mg²⁺-induced decrease of the rate of photosystem I (PS I) electron transport (DCIPH₂ → methyl viologen) in thylakoids under saturated light intensities has been reported earlier (S. Bose, J. E. Mullet, G. E. Hoch, and C. J. Arntzen, 1981, Photobiochem. Photobiophys.2, 45–52). A similar effect is observed with Na⁺, although the concentration required for half-maximal inhibition was higher by about two orders of magnitude. The cation effect was gradually abolished as the thylakoids were aged by incubation at 30 °C for 6 h. The loss of cation effect on PS I electron transport rate during aging was parallel to the corresponding loss of cation effect on thylakoid stacking. The cation concentration required for thylakoid stacking and the degree of inhibition as a function of cation concentration correlated strongly with the degree of thylakoid stacking. These observations indicated that the inhibition of the rate of PS I electron transport by cations is a consequence of cation-induced stacking of thylakoid membranes. The observed inhibition of the rate of PS I electron transport is discussed in terms of two hypotheses: (i) a fraction (20–30%) of the PS I complexes is trapped in the appressed region of grana and becomes unavailable to the electron donor (DCIPH₂) and (ii) the membrane structure is altered by the cations in such a manner that the rate constant of electron donation by the donor to the electron transport chain in the thylakoid is decreased.     

Temperature dependencies of the rate of electron flow and of the formation of the high energy state in spinach chloroplasts and leaves

The relationship between the rate of electron transport andthe rate of formation of the high energy state during illuminationwas studied by measuring temperature dependencies of oxygenuptake and of the induction of millisecond delayed fluorescenceof chlorophyll in isolated spinach chloroplasts and in spinachleaves. Measurements in chloroplasts with and without added electronacceptors showed that the decrease in the rate of formationof the high energy state caused by cooling was due to the decreasein the rate of electron transport linked to the Mehler reaction. Measurement of the induction of delayed fluorescence in leavessuggested the existence of a similar temperature-sensitive stepin the electron transport system. (Received February 17, 1977; )