Physiologia Plantarum 140: 46–56. 2010 Copyright © Physiologia Plantarum 2010, ISSN 0031-9317 Heat stress results in loss of chloroplast Cu/Zn superoxide dismutase and increased damage to Photosystem II in combined drought-heat stressed Lotus japonicus Martha Sainz, Pedro D´ ıaz, Jorge Monza and Omar Borsani ∗ Laboratorio de Bioqu´ ımica, Departamento de Biolog´ ıa Vegetal, Facultad de Agronom´ ıa, Universidad de la Rep ´ ublica, Av. Garz ´ on 780, CP 12900, Montevideo, Uruguay Correspondence *Corresponding author, e-mail: oborsani@fagro.edu.uy Received 23 April 2010; revised 3 May 2010 doi:10.1111/j.1399-3054.2010.01383.x Drought and heat stress have been studied extensively in plants, but most reports involve analysis of response to only one of these stresses. Studies in which both stresses were studied in combination have less commonly been reported. We report the combined effect of drought and heat stress on Photosystem II (PSII) of Lotus japonicus cv. Gifu plants. Photochemistry of PSII was not affected by drought or heat stress alone, but the two stresses together decreased PSII activity as determined by fluorescence emission. Heat stress alone resulted in degradation of D1 and CP47 proteins, and D2 protein was also degraded by combined drought – heat stress. None of these proteins were degraded by drought stress alone. Drought alone induced accumulation of hydrogen peroxide but the drought–heat combination led to an increase in superoxide levels and a decrease in hydrogen peroxide levels. Furthermore, combined drought–heat stress was correlated with an increase in oxidative damage as determined by increased levels of thiobarbituric acid reactive substances. Heat also induced degradation of chloroplast Cu/Zn superoxide dismutase (SOD: EC 1.15.1.1) as shown by reduced protein levels and isozyme-specific SOD activity. Loss of Cu/Zn SOD and induction of catalase (CAT: EC 1.11.1.6) activity would explain the altered balance between hydrogen peroxide and superoxide in response to drought vs combined drought–heat stress. Degradation of PSII could thus be caused by the loss of components of chloroplast antioxidant defence systems and subsequent decreased function of PSII. A possible explanation for energy dissipation by L. japonicus under stress conditions is discussed. Introduction Drought is one of the most important environmental fac- tors limiting photosynthesis. The rate of photosynthesis declines during low water stress because photoinhibi- tion increases, and this may result as a consequence of drought stress in combination with other abiotic stresses (Takahashi and Murata 2008). Limitation of Abbreviations – CAT, catalase; DAB, 3, 3’-diaminobenzidine; NBT, nitro blue tetrazolium; NPQ, non-photochemical quenching; PSII, Photosystem II; ROS, reactive oxygen species; SOD, superoxide dismutase; TBARS, thiobarbituric acid reactive substances. photosynthesis by strong light was initially showed over 50 years ago, and it has become clear that the pri- mary target of photoinhibition is Photosystem II (PSII) (Kok 1956). Plants are often subjected to a combination of stresses such as drought, heat and high irradiance; thus, analysis of plant responses to the combination of different types of stress may be critical for better 46 Physiol. Plant. 140, 2010