ABSTRACT
Transient Receptor Potential (TRP) channels are a large and diverse family of tetrameric cation selective channels that are activated by many different types of stimuli, including noxious heat or cold, organic ligands such as vanilloids or cooling agents, or intracellular Ca2+. Structures available for all subtypes of TRP channels reveal that the transmembrane domains are closely related despite their unique sensitivity to activating stimuli. Here we use computational and electrophysiological approaches to explore the conservation of the cooling agent binding pocket identified within the S1-S4 domain of the Melastatin subfamily member TRPM8, the mammalian sensor of noxious cold, with other TRPM channel subtypes. We find that a subset of TRPM channels, including TRPM2, TRPM4 and TRPM5, contain pockets very similar to the cooling agent binding pocket in TRPM8. We then show how the cooling agent icilin modulates activation of TRPM4 to intracellular Ca2+, enhancing the sensitivity of the channel to Ca2+ and diminishing outward-rectification to promote opening at negative voltages. Mutations known to promote or diminish activation of TRPM8 by cooling agents similarly alter activation of TRPM4 by icilin, suggesting that icilin binds to the cooling agent binding pocket to promote opening of the channel. These findings demonstrate that TRPM4 and TRPM8 channels share related ligand binding pockets that are allosterically coupled to opening of the pore.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
We revised the text to address the comments of the reviewers and we have added data to Fig. 4 and 6 to document the influence of icilin on slowing of channel closure following removal of internal Ca2+. We have also added data showing that icilin does not have any apparent effects on TRPM3 (Fig. 4 Fig. Supp. 1) and we have increased the n values by including additional data to Fig. 3, 4, 5, 6, 7 and 8.
