Transient receptor potential melastatin-8 (TRPM8) is a cation channel that is activated by cold and “cooling agents” such as menthol and icilin, which induce a cold sensation. Stimulation of TRPM8 activates an intracellular signaling cascade that ultimately leads to a change in the gene expression pattern of the cells. Here, we investigated the TRPM8-induced signaling pathway that links TRPM8 channel activation to gene transcription. Using a pharmacological approach, we show that inhibition of phosphatidylinositol 4-phosphate 5 kinase alpha (PIP5K), an enzyme essential for the biosynthesis of phosphatidylinositol 4,5-bisphosphate, attenuated TRPM8-induced gene transcription. Analyzing the link between TRPM8 and Gq proteins, we show that pharmacological inhibition of the beta gamma subunits impairs TRPM8 signaling. In addition, genetic studies showed that TRPM8 requires an activated Galpha subunit for signaling. In the nucleus, the TRPM8-induced signaling cascade triggers an activation of the transcription factor AP-1, a complex consisting of a dimer of basic region leucine zipper (bZIP) transcription factors. Here, we identified the bZIP protein c-Jun as an essential component of AP-1 within the TRPM8-induced signling cscade. In summary, with PIP5K, Gq subunits and c-Jun we have identified key molecules in the TRPM8-induced signaling from the plasma membrane to the nucleus.

The glycoprotein hormones luteinizing hormone, follicle-stimulating hormone, thyroid-stimulating hormone and chorionic gonadotropin consist of a common alpha-subunit and a hormone-specific beta-subunit. The alpha-subunit is expressed in thyrotrophs, gonadotrophs, and placental cells, and its expression is regulated by extracellular signal molecules. Here, we assessed the regulation of the gene encoding the alpha-subunit in placental JEG-3 cells, using a chromatin-embedded reporter gene under the control of the glycoprotein hormone alpha-subunit promoter. The results show that stimulation of the cells with forskolin, an activator of adenylate cyclase, induced transcription of a reporter gene controlled by the alpha-subunit promoter. Similarly, expression of a nuclear-targeted catalytic subunit of cAMP-dependent protein kinase increased reporter gene transcription. Expression experiments involving constitutively active mutants of MKK6 and MEKK-1 showed that the alpha-subunit promoter is responsive to activation of p38 protein kinase but only slightly responsive to activation of c-Jun N-terminal protein kinase. Finally, we show that stimulation of a Galphaq-coupled designer receptor triggers transcription of a reporter gene controlled by the glycoprotein hormone alpha-subunit promoter, mediated by the transcription factor CREB. Deletion mutagenesis highlights the importance of a tandem cAMP response element within the alpha-subunit promoter that acts as a point of convergence for different signaling pathways.