Identification of chloroplast envelope proteins with critical importance for cold acclimation

The ability of plants to cope with cold temperatures relies on their photosynthetic activity. This already demonstrates that the chloroplast is of utmost importance for cold acclimation and acquisition of freezing tolerance. During cold acclimation, the properties of the chloroplast change markedly. To provide the most comprehensive view of the protein repertoire of chloroplast envelope, we analysed this membrane system in Arabidopsis thaliana using MS-based proteomics. Profiling chloroplast envelope membranes was achieved by a cross comparison of protein intensities across plastid and the enriched membrane fraction both under normal and cold conditions. To address envelop localization, multivariable logistic regression models the probabilities for the classification problem. In total, we identified 38 envelope membrane intrinsic or associated proteins exhibiting altered abundance after cold acclimation. These proteins comprise several solute carries, such as the ATP/ADP antiporter NTT2 (substantially increased abundance) or the maltose exporter MEX1 (substantially decreased abundance). Remarkably, analysis of the frost recovery of ntt loss-of-function and mex1 overexpressor mutants confirmed that the comparative proteome is well suited to identify novel key factors involved in cold acclimation and acquisition of freezing tolerance. Moreover, for proteins with known physiological function we propose scenarios explaining their possible role in cold acclimation. Furthermore, spatial proteomics introduces a novel layer of complexity and enabled the identification of proteins differentially localized at the envelope membrane under the changing environmental regime.