AbstractTemperature sensation initiates from the activation of cellular receptors when the cell is exposed to a decrease in temperature. Here, we applied a phosphoproteome profiling approach to the human lung epithelial cell line BEAS‐2B to elucidate cellular cold‐responsive processes. The primary aim of this study was to determine which intracellular changes of phosphorylation are accompanied by cold sensation. Eighteen protein spots that exhibited differentially phosphorylated changes in cells were identified. Most of the proteins that were phosphorylated after 5 or 10 min were returned to control levels after 30 or 60 min. Identified proteins were mainly RNA‐related (i.e., they were involved in RNA binding and splicing). Temperature (18 and 10°C) stimuli showed homologies that were detected for time course changes in phosphoproteome. The data indicated a time‐shift between two temperatures. The phosphorylation of putative cold responsive markers, such as ribosomal protein large P0 and heterochromatin‐associated proteins 1, were verified by Western blotting in cells transfected with TRPM8 or TRPA1. J. Cell. Biochem. 112: 633–642, 2011. © 2010 Wiley‐Liss, Inc.