Abstract
Chloroplast thylakoid membranes were isolated from leaves of unhardened and cold-acclimated spinach (Spinacia oleracea L.). For freezethaw treatment, the membranes were suspended in complex media composed to simulate the solute concentrations in the chloroplast stroma in the unhardened and hardened states of the leaves. In particular, high concentrations of amino acids were applied for simulating the hardened state. After frost treatment, photosynthetic activities and chlorophyll fluorescence parameters of the thylakoids were tested to determine the degree of freezing damage. The results revealed a pattern of freezing injury similar to that observed upon frost treatment of thylakoids in situ. A major manifestation of damage was the inhibition of photosynthetic electron transport. Uncoupling of photophosphorylation, which is the dominating effect of freezing of thylakoids suspended in binary solutions (e.g., containing one sugar and one inorganic salt), was also visible but less pronounced in the complex media. Thylakoids obtained from cold-acclimated leaves did not exhibit an increased frost tolerance in vitro, as compared with thylakoids from unhardened plants. The results, furthermore, indicated a strong protective effect of free amino acids at the concentrations and composition found in chloroplasts of hardened leaves. The presence of inorganic salts in the complex media slightly stabilized rather than damaged the membranes during freezing. It is concluded that inactivation of thylakoids in situ may be understood as the destabilizing action of the combined solutes surrounding the thylakoids, occurring when solute concentration is raised due to freezing of water.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Abbreviations
- Chl:
-
chlorophyll
- DCMU:
-
3-(3′,4′-dichlorophenyl)-1,1-dimethylurea
- Hepes:
-
4-(2-hydroxyethyl)-1-piper-azineethanesulfonic acid
- PSI:
-
photosystem I
- PSII:
-
photosystem II
References
Arnon, D.I. (1949) Copper enzymes in isolated chloroplasts. Polyphenoloxidase in Beta vulgaris. Plant Physiol. 24, 1–15
Barényi, B., Krause, G.H. (1985) Inhibition of photosynthetic reactions by light. A study with isolated spinach chloroplasts. Planta 163, 218–226
Ben-Hayyim, G., Krause, G.H. (1980) Transport of mono-and divalent cations across chloroplast membranes mediated by the ionophore A 23187. Arch. Biochem. Biophys. 202, 546–557
Bennoun, P., Li, V.S. (1973) New results on the mode of action of 3,-(3,4-dichlorphenyl)-1,1-dimethylurea in spinach chloroplasts. Biochim. Biophys. Acta 292, 162–168
Garber, M.P., Steponkus, P.L. (1976a) Alterations in chloroplast thylakoids during an in vitro freeze-thaw cycle. Plant Physiol. 57, 673–680
Garber, M.P., Steponkus, P.L. (1976b) Alterations in chloroplast thylakoids during cold acclimation. Plant Physiol. 57, 681–686
Giersch, Ch., Heber, U., Krause, G.H. (1980) ATP transfer from chloroplasts to the cytosol of leaf cells during photosynthesis and its effect on leaf metabolism. In: Plant membrane transport: Current conceptual issues, pp. 65–80, Spanswick, R.M., Lucas, W.J., Dainty, J., eds. Elsevier/North Holland Biomedical Press, Amsterdam
Heber, U., Schmitt, J.M., Krause, G.H., Klosson, R.J., Santarius, K.A. (1981) Freezing damage to thylakoid membranes in vitro and in vivo. In: Effects of low temperatures on biological membranes, pp 263–283, Morris, G.J., Clarke, A., eds. Academic Press, London New York
Heber, U., Tyankova, L., Santarius, K.A. (1971) Stabilization and inactivation of biological membranes during freezing in the presence of amino acids. Biochim. Biophys. Acta 241, 578–592
Heber, U., Tyankova, L., Santarius, K.A. (1973) Effects of freezing on biological membranes in vivo and in vitro. Biochim. Biophys. Acta 291, 23–37
Izawa, S., Good, N. (1966) Effects of salts and electron transport on the conformation of isolated chloroplasts. II. Electron microscopy. Plant Physiol. 41, 544–552
Jagendorf, A.T. (1975) Mechanism of photophosphorylation. In: Bioenergetics of photosynthesis, pp. 413–492, Govindjee, ed. Academic Press, New York London
Jensen, R.G., Bassham, J.A. (1966) Photosynthesis by isolated chloroplasts. Proc. Natl. Acad. Sci. USA 56, 1095–1101
Kaiser, W.M. (1984) Sites and mechanisms for the inhibition of photosynthesis by water stress. In: Advances in photosynthesis, vol. 4, pp. 341–348, Sybesma, C., ed. Martinus Nijhoff/Junk, The Hague
Kaiser, W.M., Weber, H., Sauer, M. (1983) Photosynthetic capacity, osmotic response and solute content of leaves and chloroplasts from Spinacia oleracea under salt stress. Z. Pflanzenphysiol. 113, 15–27
Klosson, R.J., Krause, G.H. (1981a) Freezing injury in cold-acclimated and unhardened spinach leaves. I. Photosynthetic reactions of thylakoids isolated from frost-damaged leaves. Planta 151, 339–346
Klosson, R.J., Krause, G.H. (1981b) Freezing injury in coldacclimated and unhardened spinach leaves. II. Effects of freezing on chlorophyll fluorescence and light scattering reactions. Planta 151, 347–352
Krause, G.H., Klosson, R.J., Tröster, U. (1982a) On the mechanism of freezing injury and cold acclimation of spinach leaves. In: Plant cold hardiness and freezing stress, pp. 55–75, Li, P.H., Sakai, A., eds. Academic Press, New York London
Krause, G.H., Klosson, R.J., Justenhoven, A., Ahrer-Steller, V. (1984) Effects of low temperatures on the photosynthetic system in vivo. In: Advances in photosynthesis research vol. 4, pp. 349–358, Sysbesma, C., ed. Martinus Nijhoff/Junk, The Hague
Krause, G.H., Köster, S., Wong, S.C. (1985) Photoinhibition of photosynthesis under anaerobic conditions studied with leaves and chloroplasts of Spinacia oleracea L. Planta 165, 430–438
Krause, G.H., Vernotte, C., Briantais, J.-M. (1982b) Photoinduced quenching of chlorophyll fluorescence in intact chloroplasts and algae. Resolution into two components. Biochim. Biophys. Acta 679, 116–124
Krueger, R.W., Miles, D. (1981) Photosynthesis in Fescue. I. High rate of electron transport and phosphorylation in chloroplasts of hexaploid plants. Plant Physiol. 67, 763–767
Melis, A., Homann, P.H. (1975) Kinetic analysis of the fluorescence induction in 3-(3,4-dichlorphenyl)-1,1-dimethylurea poisened chloroplasts. Photochem. Photobiol. 21, 431–437
Nishimura, M., Ito, T., Chance, B. (1962) Studies on bacterial photophosphorylation. III. A sensitive and rapid method of determination of photophosphorylation. Biochim. Biophys. Acta 59, 177–182
Rumich-Bayer, S., Krause, G.H. (1986) Freezing damage and frost tolerance of the photosynthetic apparatus studied with isolated mesophyll protoplasts of Valerianella locusta L. Photosynth. Res. (in press)
Santarius, K.A. (1971) The effect of freezing on thylakoid membranes in the presence of organic acids. Plant Physiol. 48, 156–162
Santarius, K.A. (1984) Effective cryoprotection of thylakoid membranes by ATP. Planta 161, 555–561
Santarius, K.A. (1986) Freezing of isolated thylakoid membranes in complex media. I. The effect of potassium and sodium chloride, nitrate and sulfate. Cryobiol. 23, 168–176
Santarius, K.A., Heber, U. (1972) Physiological and biochemical aspects of frost damage and winter hardiness in higher plants. In: Proc. of a colloquium on the winter hardiness of cereals, pp. 7–29, Agricultural Research Institute of the Hungarian Academy of Sciences, Martonvásár
Santarius, K.A., Milde, H. (1977) Sugar compartmentation in frost-hardy and partially dehardened cabbage leaves. Planta 136, 163–166
Schuldiner, S., Rottenberg, H., Avron, M. (1972) Determination of Δ pH in chloroplasts. 2. Fluorescent amines as a probe for the determination of Δ pH in chloroplasts. Eur. J. Biochem. 25, 64–70
Trebst, A. (1974) Energy conservation in photosynthetic electron transport of chloroplasts. Annu. Rev. Plant Physiol. 25, 423–458
Volger, H.G., Heber, U. (1975) Cryoprotective leaf proteins. Biochim. Biophys. Acta 412, 335–349
Volger, H., Heber, U., Berzborn, R.J. (1978) Loss of function of biomembranes and solubilization of membrane proteins during freezing. Biochim. Biophys. Acta 511, 455–469
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Grafflage, S., Krause, G.H. Simulation of in situ freezing damage of the photosynthetic apparatus by freezing in vitro of thylakoids suspended in complex media. Planta 168, 67–76 (1986). https://doi.org/10.1007/BF00407011
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00407011