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Structural determinants of gating in the TRPV1 channel

Abstract

Transient receptor potential vanilloid 1 (TRPV1) channels mediate several types of physiological responses. Despite the importance of these channels in pain detection and inflammation, little is known about how their structural components convert different types of stimuli into channel activity. To localize the activation gate of these channels, we inserted cysteines along the S6 segment of mutant TRPV1 channels and assessed their accessibility to thiol-modifying agents. We show that access to the pore of TRPV1 is gated by S6 in response to both capsaicin binding and increases in temperature, that the pore-forming S6 segments are helical structures and that two constrictions are present in the pore: one that impedes the access of large molecules and the other that hampers the access of smaller ions and constitutes an activation gate of these channels.

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Figure 1: TRPV1-C157A can be used as a template for SCAM analysis of the channel pore.
Figure 2: Residue Leu681 restricts the entrance of large cations.
Figure 3: Ag+ modification reveals the presence of an intracellular gate for small ions at Tyr671.
Figure 4: CuP induces state-dependent disulfide bonds in 18TRPV1-L681C and 18TRPV1-Y671C.
Figure 5: Gating of the TRPV1 channel by temperature.
Figure 6: The S6 segment has an α-helical character.
Figure 7: Homology model of TRPV1 based on the Kv1.2 structure.

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Acknowledgements

We thank D. Julius at the University of San Francisco for providing the TRPV1 cDNA, and S. Simon at Duke University and M. Rosenbaum at Instituto de Ciencias Nucleares of Universidad Nacional Autónoma de México for thoughtful discussion of this manuscript. We also thank L. Ongay, S. Rojas, A. Aguilera Jiménez, J. Barbosa, A.M. Escalante and F. Sierra at Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, for expert technical support. This work was supported by grants from La Dirección General de Asuntos del Personal Académico (DGAPA)-Universidad Nacional Autónoma de México (UNAM) IN200308-3 to T.R. and IN202006–3 to L.D.I., and CONACyT No. 58038 to T.R. and No. 48990 to L.D.I. This study was performed in partial fulfillment of the requirements for the Doctorate degree in Biomedical Sciences of H.S. at the UNAM.

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H.S., A.J.-O., E.H.-G. and T.R. performed experiments; I.L. did molecular biology and cell culture; M.S.-G. and I.I.A.-O. performed molecular modeling; L.D.I. and T.R. designed experiments, analyzed data and wrote the paper; all authors discussed the data.

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Correspondence to Tamara Rosenbaum.

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Salazar, H., Jara-Oseguera, A., Hernández-García, E. et al. Structural determinants of gating in the TRPV1 channel. Nat Struct Mol Biol 16, 704–710 (2009). https://doi.org/10.1038/nsmb.1633

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