TRPM8 voltage sensor mutants reveal a mechanism for integrating thermal and chemical stimuli


TRPM8, a member of the transient receptor potential (TRP) channel superfamily, is expressed in thermosensitive neurons, in which it functions as a cold and menthol sensor. TRPM8 and most other temperature-sensitive TRP channels (thermoTRPs) are voltage gated; temperature and ligands regulate channel opening by shifting the voltage dependence of activation. The mechanisms and structures underlying gating of thermoTRPs are currently poorly understood. Here we show that charge-neutralizing mutations in transmembrane segment 4 (S4) and the S4-S5 linker of human TRPM8 reduce the channel's gating charge, which indicates that this region is part of the voltage sensor. Mutagenesis-induced changes in voltage sensitivity translated into altered thermal sensitivity, thereby establishing the strict coupling between voltage and temperature sensing. Specific mutations in this region also affected menthol affinity, which indicates a direct interaction between menthol and the TRPM8 voltage sensor. Based on these findings, we present a Monod-Wyman-Changeux–type model explaining the combined effects of voltage, temperature and menthol on TRPM8 gating.

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Figure 1: Charge neutralizations in S4 and the S4-S5 linker of TRPM8.
Figure 2: Determination of the total gating charge with the limited slope method.
Figure 3: Charge neutralizations in S4 and the S4-S5 linker alter the temperature dependence of TRPM8.
Figure 4: Residues in S4 and the S4-S5 linker determine the menthol sensitivity of TRPM8.
Figure 5: TRPM8-TRPM5 chimeras.
Figure 6: A quantitative model describing the combined effects of temperature, voltage and menthol on TRPM8.


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We thank G. Droogmans, F. Mahieu, J. Vriens, R. Vennekens and J. Prenen for helpful suggestions and criticisms, and A. Patapoutian and M. Bandell for the Y745H and L1009R mutants. This work was supported by grants from the Human Frontiers Science Program (RGP32/2004), the Belgian Federal Government (IUAP P5/05), the Research Foundation–Flanders (G.0136.00, G.0172.03 and G.0565.07) and the Research Council of the KU Leuven (GOA2004/07 and EF/95/010).

Author information




T.V. designed, performed and analyzed the electrophysiological experiments. G.O. developed the radioligand binding assay and conducted the biochemical experiments and molecular modeling. A.J. designed and performed TRPM8 mutagenesis. T.V. and K.T. performed the mathematical modeling. T.V. wrote the manuscript; G.O., K.T. and B.N. edited the manuscript; and T.V. and B.N. supervised the project.

Corresponding author

Correspondence to Thomas Voets.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Examples of the method used to determine Popen. (PDF 131 kb)

Supplementary Fig. 2

Fits of two published models describing the temperature dependence of TRPM8 to experimental data. (PDF 119 kb)

Supplementary Fig. 3

[3H]menthol displacement experiments on nontransfected HEK 293 cells and on cells expressing TRPM2, WT TRPM8 and TRPM8 mutants. (PDF 83 kb)

Supplementary Fig. 4

Plasma membrane expression and current densities for different TRPM8-TRPM5 chimeras. (PDF 249 kb)

Supplementary Fig. 5

Additional predictions of the MWC model. (PDF 99 kb)

Supplementary Table 1

Parameters that describe the expression level, voltage dependence, temperature dependence and menthol sensitivity of WT and mutant TRPM8. (PDF 218 kb)

Supplementary Methods

Procedure for determination of activation curves, comparison of two models describing the temperature dependence of TRPM8, and mathematical description of the MWC and HH models. (PDF 264 kb)

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Voets, T., Owsianik, G., Janssens, A. et al. TRPM8 voltage sensor mutants reveal a mechanism for integrating thermal and chemical stimuli. Nat Chem Biol 3, 174–182 (2007).

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