Robert M. Smillie

The effect of temperature of the rate of photosynthetic electron transfer in chloroplasts of chilling-sensitive and chilling-resistant plants

Abstract 1. Photochemical activities as a function of temperature have been compared in chloroplasts isolated from chilling-sensitive (below approximately 12 °C) and chilling-resistant plants. 2. An Arrhenius plot of the photoreduction of NADP + from water by chloroplasts isolated from tomato ( Lycopersicon esculentum var. Gross Lisse), a chilling-sensitive plant, shows a change in slope at about 12 °C. Between 25 and 14 °C the activation energy for this reaction is 8.3 kcal·mole −1 . Between 11 and 3 °C the activation energy increases to 22 kcal·mole −1 . Photoreduction of NADP + by chloroplasts from another chilling-sensitive plant, bean ( Phaseolus vulgaris var. brown beauty), shows an increase in activation energy from 5.9 to 17.5 kcal·mole −1 below about 12 °C. 3. The photoreduction of NADP + by chloroplasts isolated from two chilling-resistant plants, lettuce ( Lactuca sativa var. winter lake) and pea ( Pisum sativum var. greenfeast), shows constant activation energies of 5.4 and 8.0 kcal·mole −1 , respectively, over the temperature range 3–25 °C. 4. The effect of temperature on photosynthetic electron transfer in the chloroplasts of chilling-sensitive plants is localized in Photosystem I region of photosynthesis. Both the photoreduction of NADP + from reduced 2,6-dichlorophenol-indophenol and the ferredoxin-NADP + reductase (EC 1.6.99.4) activity of choroplasts of chilling-sensitive plants show increases in activation energies at approximately 12 °C whereas Photosystem II activity of chloroplasts of chilling-sensitive plants shows a constant activation energy over the temperature range 3–25 °C. The photoreduction of Diquat (1,1′-ethylene-2,2′-dipyridylium dibromide) from water by bean chloroplasts, however, does not show a change in activation energy over the same temperature range. The activation energies of each of these reactions in chilling-resistant plants is constant between 3 and 25 °C. 5. The effect of temperature on the activation energy of these reactions in chloroplasts from chilling-sensitive plants is reversible. 6. In chilling-sensitive plants, the increased activation energies below approximately 12 °C, with consequent decreased rates of reaction for the photoreduction of NADP + , would result in impaired photosynthetic activity at chilling temperatures. This could explain the changes in chloroplast structure and function when chilling-sensitive plants are exposed to chilling temperatures.

Multi-temperature effects on Hill reaction activity of barley chloroplasts

1. The relationship between temperature and Hill reaction activity has been investigated in chloroplasts isolated from barley (Hordeum vulgare L. cv. Abyssinian). 2. An Arrhenius plot of the photoreduction of 2,6-dichlorophenolindophenol (DCIP) showed no change in slope over the temperature range 2--38degreesC. The apparent Arrhenius activation energy (Ea) for the reaction was 48.1 kJ/mol. 3. In the presence of an uncoupler of photophosphorylation, methylamine, the Ea for DCIP photoreduction went through a series of changes as the temperature was increased. Changes were found at 9, 20, 29 and 36degreesC. The Ea was highest below 9degreesC at 63.7 kJ/mol. Between 9 and 20degreesC the Ea decreased to 40.4 kJ/mol and again to 20.2 kJ/mol between 20 and 29degreesC. Between 29 and 36degreesC there was no further increase in activity with increasing temperature. The temperature-induced changes at 9, 20 and 29degreesC were reversible. At temperatures above 36degreesC (2 min) a thermal and largely irreversible inactivation of the Hill reaction occurred. 4. Temperature-induced changes in Ea were also found when ferricyanide was substituted for DCIP or gramicidin D for methylamine. The addition of an uncoupler of photophosphorylation was not required to demonstrate temperature-induced changes in DCIP photoreduction following the exposure of the chloroplasts to a low concentration of cations. 5. The photoreduction of the lipophilic acceptor, oxidized 2, 3, 5, 6-tetramethyl-p-phenylenediamine, also showed changes in Ea in the absence of an uncoupler. 6. The temperature-induced changes in Hill activity at 9 and 29degreesC coincided with temperature-induced changes in the fluidity of chloroplast thylakoid membranes as detected by measurements of electron spin resonance spectra. It is suggested that the temperature-induced changes in the properties and activity of chloroplast membranes are part of a control mechanism for regulation of chloroplast development and photosynthesis by temperature.

Heat tolerance and heat hardening in crop plants measured by chlorophyll fluorescence

Variable chlorophyll fluorescence (Fv), indicative of the capacity for electron flow through photosystem II, decreased as leaves became heat injured. In leaves heated at a rate of 1°C per minute, Fv became undetectable above 43.3°C in barley, 44.2°C in pea, 45.2°C in bean, 45.3°C in tomato, 49.5°C in maize and 51.0°C in papaya. In heat hardened barley this temperature was increased by 6 to 9°C compared with unhardened barley. Measurement of Fv in vivo provides a rapid method to monitor the onset of cellular heat injury, to rank plants in order of heat resistance and to follow heat hardening. It appears to be suitable for developing a mass screening test for heat tolerance.

Heat tolerance and cold tolerance of cultivated potatoes measured by the chlorophyll-fluorescence method

Heat and cold tolerances were determined for 13 clones of the commonly cultivated potato, Solanum tuberosum L. Five clones were considered to be adapted to warm climates and the others to cool climates only in terms of their ability to produce tubers. The decrease in the rate of the induced rise in chlorophyll fluorescence after heating leaves at 41°C for 10 min was used to measure relative heat tolerance, and the decrease following chilling at 0°C was used to measure relative cold tolerance. The warm-adapted clones all showed enhanced heat tolerance compared with the cool-adapted clones. Higher heat tolerance was also correlated with a greater tolerance towards a cold stress of 0°C and it is suggested that the warm-adapted clones were selections showing an increased generalized capacity to withstand environmental stresses of several kinds rather than a specific genotypic adaptation to tolerate warm temperatures. Heat and cold tolerances were also determined for several other species of potato cultivated in the Andean region of South America. Of these, S. phureja, which is found at low altitudes on the eastern slopes of the Andes, showed a tolerance to heat comparable to that of the warm-adapted clones of the common potato, the two most heat tolerant of which contained some phureja in their parentage. Diploid and triploid species of cultivated potatoes were considerably more cold tolerant than the clones of the common potato, a tetraploid. The genetic variability for heat and cold tolerance in cultivated and wild potatoes is discussed in relation to increasing the tolerance of the potato to these stresses.

The distribution of galactolipids in mesophyll and bundle sheath chloroplasts of maize and sorghum

Abstract The galactolipid content of bundle sheath chloroplasts is significantly higher, on a chlorophyll basis, than that of mesophyll chloroplasts in both maize and sorghum. It is suggested that the molar ratio of galactolipid to chlorophyll reflects the degree of grana formation in a chloroplast.

Tolerances of wild potato species from different altitudes to cold and heat.

The ability of wild potatoes (Solanum spp.) to adapt to potentially stressful environmental temperatures was investigated by measuring the cold and heat tolerances of plants grown near sea-level in Lima following collection of tubers from plants growing naturally at altitudes ranging from 450 to 4,200 m. Relative cold tolerance was measured in leaves stored at 0°C by the decrease in the induced rise of chlorophyll fluorescence. Similarly, changes in chlorophyll fluorescence were used to determine the relative heat tolerance of leaves heated at 41°C for 10 min. With increasing altitude, the cold tolerance of different species tended to increase and conversely, heat tolerance decreased. However, these two genotypic adaptations were not closely correlated and appear to vary independently of each other in response to climate.

Chilling resistance of somatic hybrids of tomato and potato

Chilling resistance was investigated in four somatic hybrid plants of tomato and potato. Tomato (Lycopersicon esculentum) is very susceptible to injury at chilling temperatures, whereas the other hybrid parent, potato (Solanum tuberosum), is relatively chilling resistant. Leaves of the four somatic hybrid plants and leaves of tomato and potato plants were detached and stored at 0°C. Chilling resistance was determined by the rate of change in photosynthetic electron transfer activity in the stored leaves, as indicated by measurements of chloroplast cytochromef reduction. The chilling resistance of all four tomato-potato hybrids was intermediate between the chilling resistances of tomato and potato. Somatic hybrids produced by fusion of tomato and potato protoplasts may be useful for transferring genes for chilling resistance into the domestic tomato.

Oxidation-reduction properties of phytoflavin, a flavoprotein from blue-green algae

Abstract Phytoflavin from the blue-green alga Anacystis nidulans is a protein which can replace the low potential electron carrier, ferredoxin, in photosynthetic electron transport by isolated chloroplasts. Phytoflavin contains one molecule of FMN per molecule of protein, and was found to accept two reducing equivalents during reduction. It forms a stable semiquinone and can be reduced in two stages. Molar extinction coefficients have been determined for the absorption spectra of the three oxidation states of the protein. Oxidation-reduction potentials have been determined for each stage in reduction over a range of pH values. At pH 7.0, the potentials are −0.221 V and −0.447 V. The second potential is very low for a flavoprotein, and lower than the potentials determined for ferredoxin from A. nidulans and higher plants. The potentials determined provide a quantitative measure of the stability of the semiquinone of phytoflavin. The results presented help to explain the mechanism of reduction and oxidation and how phytoflavin can substitute for ferredoxin in photosynthetic electron transport by utilization of the oxidation-reduction couple, phytoflavin semiquinone/hydroquinone.

A photosystem I mutant in barley (Hordeum vulgare L.).

The nuclear gene mutantviridis-n34 in barley has been characterized as a photosystem I mutant. The slow component of the light dependent absorption change at 518 nm and the photooxidation of cytochromef were greatly reduced in mutant leaves compared with wild-type leaves. The oxidation of cytochromef in mutant leaves irradiated with far-red light was only 7% of the value obtained with wild-type. The fast component of the 518 nm absorbance change was similar in wild-type and mutant leaves. The rate of photosystem I electron transport in the mutant is approximately 10% of the rate observed with wild-type thylakoids whereas photosystem II electron transport appears normal. Little or noP700 was detected in the mutant. The levels of cytochromes and ferredoxin-NADP+ oxidoreductase were normal. Mutant thylakoids were unable to generate a proton gradient upon illumination. Analysis of the polypeptide composition of thylakoids ofviridis-n34 revealed that these are depleted in chlorophylla-protein 1 and three polypeptides believed to be iron-sulfur proteins. These four polypeptides are components of photosystem I particles isolated from wild-type barley.

Tolerance of Borya nitida, a poikilohydrous angiosperm, to heat, cold and high-light stress in the hydrated state.

Borya nitida Labill., a plant able to colonize rock outcrops and shallow sands in areas of high incident solar radiation in Western Australia, was examined for its tolerance to extremes of temperature, and to intense visible radiation. Stress injury to the leaves from heat, chilling or photoinhibitory light was followed by the decrease in in-vivo variable chlorophyll fluorescence. Heat injury was also ascertained by an increase in the “constant” fluorescence. Borya nitida leaves were extremely heat tolerant when heated at 1° C min-1. In-vivo variable chlorophyll fluorescence was detectable up to 55° C, several degrees higher than either maize or barley which are, respectively, adapted to warm and cool climates. An increase in “constant” fluorescence occurred above 50° C in B. nitida. This compares with values in the literature of 48–49° C for three desert plants from Death Valley, California, and 44–48° C for ten species of tropical plants. Unlike the Death-Valley plants, the high degree of heat tolerance found in B. nitida did not require prior acclimation by growth at high temperatures. Borya nitida was also tolerant of a chilling temperature of 0° C. Plants grown at a low photon fluence rate (120 μmol m-2s-1) were irreversibly photoinhibited by light at 650 μmol m-2s-1. Plants grown in sunlight resisted photoinhibition; however, the capacity to withstand photoinhibition was no greater than that of plants from less extreme environments.