Comparative Proteomic Analysis Provides New Insights into Chilling Stress Responses in Rice*

Low temperature is one of the major abiotic stresses limiting the productivity and the geographical distribution of many important crops. To gain a better understanding of chilling stress responses in rice (Oryza sativa L. cv. Nipponbare), we carried out a comparative proteomic analysis. Three-week-old rice seedlings were treated at 6 °C for 6 or 24 h and then recovered for 24 h. Chilling treatment resulted in stress phenotypes of rolling leaves, increased relative electrolyte leakage, and decreased net photosynthetic rate. The temporal changes of total proteins in rice leaves were examined using two-dimensional electrophoresis. Among ∼1,000 protein spots reproducibly detected on each gel, 31 protein spots were down-regulated, and 65 were up-regulated at least at one time point. Mass spectrometry analysis allowed the identification of 85 differentially expressed proteins, including well known and novel cold-responsive proteins. Several proteins showed enhanced degradation during chilling stress, especially the photosynthetic proteins such as Rubisco large subunit of which 19 fragments were detected. The identified proteins are involved in several processes, i.e. signal transduction, RNA processing, translation, protein processing, redox homeostasis, photosynthesis, photorespiration, and metabolisms of carbon, nitrogen, sulfur, and energy. Gene expression analysis of 44 different proteins by quantitative real time PCR showed that the mRNA level was not correlated well with the protein level. In conclusion, our study provides new insights into chilling stress responses in rice and demonstrates the advantages of proteomic analysis.