Proteomic Analysis Of Ethylene Response Factor LeERF2Confers Cold Tolerance In Rice Seedlings

Low temperature is one of the major abiotic stresses limiting plant development and agricultural production. It has been established that ethylene response factor (ERF) proteins regulate a variety of biotic and abiotic stress responses, and plant development as well. Previously, our researches have revealed that transgenic rice over-expressing LeERF2(encoding a tomato ERF protein) showed enhanced tolerance to cold stress.To gain a better understanding of cold stress responses in LeERF2transgenic rice, two-week-old rice seedlings were treated at4℃for12h, then total crude proteins were extracted from leaf blades, stems and roots, separated by two-dimensional electrophoresis. Through gels quantitative2-DE image analysis and mass spectrometry identification,54proteins of the70protein spots from leaf blades and stems, and91proteins of the109protein spots from roots were changed by qualitative or quantitative method.Our results showed that18proteins from leaf blades and stems were changed between WT and transgenic lines under normal conditions, with5proteins increasing, and8proteins decreasing,5proteins only expressed in WT. These proteins were involved in metabolic process, catalytic activity and cell part. However, the protein expression were changed between WT and transgenic plants after cold treatment, with2proteins up-regulated,4proteins only expressed in OEC and1proteins expressed only in WTC. Protein function analysis informed that5proteins of them were related to metabolic process,2involved in catalytic activity, as well as a protein linked to6-phosphofructokinase complex. In doing so, we highlight some of the important differences between WT and transgenic lines after cold treatment.Comparative proteomic assays revealed that22proteins from roots were changed compared the transgenic lines with WT plants after cold treatment, with3proteins increasing,4proteins decreasing,6proteins only expressed in OEC, and9in WT. These cold responsive proteins, besides10proteins of metabolic and cellular process function, include3establishment of localization proteins,4response to stimulus proteins,5proteins related to oxidoreductase activity and hydrolase activity, and as well as some of them involved in electron carrier activity, eroxidase activity, ransporter activity, transferase activity, transferring phosphorus-containing groups.Gene expression analysis of65different proteins by quantitative real-time PCR revealed that protein and mRNA levels are not correlated. It was concluded that proteins identified in LeERF2-transgenic seedlings related to energy metabolism were up-regulated, and defense-related proteins were down-regulated during chilling stress. The functional proteomes and the subcellular localization analysis demonstrate the facts that LeERF2might have important regulatory roles in tolerance to cold stress, by several types of proteins, especially in metabolisms of carbon and energy.