The crown is the critical region for survival of winter wheat exposed to low temperature stresses. When wheat is exposed to non-freezing low temperatures, they can increase their freezing tolerance (cold acclimation, ACC). Changes within the apoplast are thought to be crucial for acquisition of freezing tolerance. However, how individual tissues within the ccrown, namely the shoot apical meristem (SAM, responsible for new shoot growth) and vascular transition zone (VTZ, located at the base of the crown)enhance tolerance to freezing has not yet been characterized. In the present study, we conducted shotgun proteomic analysis of the apoplast fluid to investigate ACC-induced proteins in the SAM and VTZ.