Characterization and knockdown of a family of ice-binding proteins from freeze-tolerant grasses
Plants are exposed to environmental stresses that threaten their growth, reproduction, and survival. At sub-zero temperatures, the recrystallization of ice in the apoplast results in cellular dehydration, disruption of the plasma membrane and cell death. Certain plants from temperate regions produce ice-binding proteins (IBPs), which protect plants by adsorbing to ice crystals and modifying their growth. Using the model crop, Brachypodium distachyon, we have identified seven novel IBPs and characterized their in vitro ice-binding activity, in planta induction, and post-translational modification. We have also shown the ability of these proteins to attenuate bacterial ice-nucleation activity. Additionally, we have generated the first IBP-knockdown in any organism, allowing us to finally confirm the importance of these proteins in membrane protection and freeze-survival. Expression of a number of IBPs from the perennial ryegrass, Lolium perenne, in Arabidopsis thaliana has localized IBPs to the apoplast, with the identification of one IBP isoform that is non-classically secreted. IBPs provided freeze protection to A. thaliana in a localization-dependent manner, with an enhanced phenotype associated with the expression of multiple isoforms. This research highlights the potential utility of IBPs for the generation of freeze-hardy crops, where previous transgenic studies have shown limited success.