The TRPM8 ion channel comprises direct Gq protein-activating capacity

The transient receptor potential (TRP) family of ion channels comprises receptors that are activated by a vast variety of physical as well as chemical stimuli. TRP channels interact in a complex manner with several intracellular signaling cascades, both up- and downstream of receptor activation. Investigating cascades stimulated downstream of the cold and menthol receptor TRPM8, we found evidence for both, functional and structural interaction of TRPM8 with Gαq. We demonstrated menthol-evoked increase in intracellular Ca2+ under extracellular Ca2+-free conditions, which was blocked by the PLC inhibitors U73122 or edelfosine. This metabotropic Ca2+ signal could be observed also in cells expressing a channel-dead (i.e. non-conducting) or a chloride-conducting TRPM8 pore mutant. However, this intracellular metabotropic Ca2+ signal could not be detected in Gαq deficient cells or in the presence of dominant-negative GαqX. Evidence for a close spatial proximity necessary for physical interaction of TRPM8 and Gαq was provided by acceptor bleaching experiments demonstrating FRET between TRPM8-CFP and Gαq-YFP. A Gαq-YFP mobility assay (FRAP) revealed a restricted diffusion of Gαq-YFP under conditions when TRPM8 is immobilized in the plasma membrane. Moreover, a menthol-induced and TRPM8-mediated G protein activation could be demonstrated by FRET experiments monitoring the dissociation of Gαq-YFP from a Gβ/Gγ-CFP complex, and by the exchange of radioactive [35S]GTPγS for GDP. Our observations lead to a view that extends the operational range of the TRPM8 receptor from its function as a pure ion channel to a molecular switch with additional metabotropic capacity.