• 2-D gel electrophoresis
• freezing stress
• MALDI–ToF
• proteomics

Plants are continuously exposed to environmental stress. Frost damage is a source of substantial agricultural losses. Many plants increase their tolerance to freezing in an adaptive process known as cold acclimation. The aim of this study is to explore the potential of proteomics as a molecular tool to investigate freezing stress in Arabidopsis thaliana.
We carried out analysis on 5 to 35 days-old plants and every age was tested from –4 to –12°C, during a time course of 24 hours. Freezing tolerance was estimated after recovery under unstressed conditions evaluating plasmatic membrane and photosynthetic apparatus damages. We focused our attention on 20 days-old plants, freezed at –10°C for 12 hours (the most resistant ones, previous acclimation or not).
We performed a proteome study based on 2-D gel electrophoresis. The results were analized through a dedicated software. Protein spots of interest were excised from the gel, subjected to in gel digestion with trypsin and analyzed by MALDI–ToF Mass Spectrometry. The MASCOTsearch engine software was used to search in the database.
Up to now amongst the identified proteins of Arabidopsis 19 were induced/inhibited by freezing stress.