Identification of a spotted leaf sheath Gene and Proteomic Analysis of Rice Seedlings Exposed to Cold and Heat Stresses

Abstract

During the life span, plants are exposed to many stresses. Since plants, unlike animals, are incapable of maintaining the optimum condition for their growth, a slight change, even transiently, may affect physiological and biochemical processes crucial for plant growth and its production. Therefore, understanding the molecular mechanisms in response to biotic and abiotic stresses is important to develop cultivars with tolerance to various stress constraints.<br/> <br/> In this study, we identified spotted leaf sheath (sles), a MAP kinase kinase kinase (MAPKKK) gene that is required for resistance against Magnaporthe oryzae. Two day after inoculation, the development of infectious hyphae (IH) were mostly restricted to primary infected cells with dark brown granules in sles mutant while IH actively grew in wild type. ROS content was elevated in sles mutant, corresponding to increased expression of genes encoding ROS-generating enzymes. Moreover, the sles mutant showed lesion mimic spots on the leaf sheath rather than on leaves which differed from that of other lesion mimic mutants (LMMs). The sles mutant also displayed early senescence, as shown, by color loss in the mesophyll cells, a decrease in chlorophyll content, and upregulation of chlorophyll degradation-related and senescence-associated genes. Taken together, our results revealed that SLES is involved in ROS homeostasis resulting resistance against pathogen infection and formation of lesion mimic spots on the leaf sheath.<br/> <br/> We also identified proteins differentially expressed in response to temperature stresses, including cold and heat, followed by recovery. Two rice cultivars with contrasting levels of tolerance to cold and heat stress, Koshihikari and Samnam, were used in this study. Proteomic responses of typical and healthy three-leaf old seedlings to sudden temperature changes were investigated. Rice seedling grown at 28/25°C (day/night) were subjected to 5 day exposure to 4°C (day/night) for cold stress, and 42°C (day/night) for heat stress, followed by 5 days of recovery. Mature leaves were harvested from plants from each treatment for protein extraction and subsequent triple TOF MS/MS analysis. Out of over 1192 proteins identified in one or more temperature treatment, more than 500 were found to be responsive to temperature stresses. Of these, 82, 159, 254, and 250 proteins were expressed in both cultivars at cold stress, recovery after cold stress, heat stress, and recovery after heat stress, respectively. In addition, 197 and 278 proteins were exclusively found in Koshihikari under cold stress and recovery and 104, and 155 proteins exclusively found in Samnam under heat stress and recovery, respectively. This study has provided a number of valuable molecular insights into temperature stress responses in rice and generated large number of candidate proteins that can form the basis of further detailed study.<br/>