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Nitric oxide activates TRP channels by cysteine S-nitrosylation

Nature Chemical Biology volume 2, pages 596–607 (2006)Cite this article

Abstract

Transient receptor potential (TRP) proteins form plasma-membrane cation channels that act as sensors for diverse cellular stimuli. Here, we report a novel activation mechanism mediated by cysteine S-nitrosylation in TRP channels. Recombinant TRPC1, TRPC4, TRPC5, TRPV1, TRPV3 and TRPV4 of the TRPC and TRPV families, which are commonly classified as receptor-activated channels and thermosensor channels, induce entry of Ca2+ into cells in response to nitric oxide (NO). Labeling and functional assays using cysteine mutants, together with membrane sidedness in activating reactive disulfides, show that cytoplasmically accessible Cys553 and nearby Cys558 are nitrosylation sites mediating NO sensitivity in TRPC5. The responsive TRP proteins have conserved cysteines on the same N-terminal side of the pore region. Notably, nitrosylation of native TRPC5 upon G protein–coupled ATP receptor stimulation elicits entry of Ca2+ into endothelial cells. These findings reveal the structural motif for the NO-sensitive activation gate in TRP channels and indicate that NO sensors are a new functional category of cellular receptors extending over different TRP families.

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Figure 1: NO activates TRPC5-mediated Ca2+ response.
Figure 2: The TRPC5-activating action of NO can be probed by reactive disulfides.
Figure 3: Chemical characterization and membrane sidedness of the action of NO and reactive disulfide in activating TRPC5 channels.
Figure 4: Cysteine modification activates TRPC5 channels via a pathway independent of receptor-activated PLC signaling cascades.
Figure 5: S-nitrosylation in TRPC5 channel protein complexes.
Figure 6: Molecular conservation of NO-induced activation in TRP channels.
Figure 7: Regulation of TRPV channel activation by nitrosylation.
Figure 8: Physiological S-nitrosylation of native TRPC5 proteins in cultured endothelial cells.
Figure 9: Native TRPC5 channels are essential for NO-induced Ca2+ influx in endothelial cells.
Figure 10: Model for TRP channel activation by NO and reactive disulfides.

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Acknowledgements

We thank D.E. Clapham and C. Strubing for TRPC5-DN, T. Furukawa and M. Nishida for helpful discussions, E. Mori and M. Sasaki for expert experiments and T. Kurosaki for IP3 receptor–deficient DT40 cells. This study was supported by research grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan, the Japan Society for the Promotion of Science, and from the Mitsubishi Foundation.

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Authors and Affiliations

  1. Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Kyoto, 615-8510, Japan

    Takashi Yoshida, Nobuaki Takahashi, Shinichiro Yamamoto, Yuji Hara & Yasuo Mori

  2. Center for Integrative Bioscience, National Institute for Physiological Sciences, Okazaki, 444-8585, Aichi, Japan

    Takashi Yoshida & Makoto Tominaga

  3. School of Life Science, The Graduate University for Advanced Studies, Okazaki, 444-8585, Aichi, Japan

    Takashi Yoshida & Makoto Tominaga

  4. Department of Physiology, Fukuoka University, Fukuoka, 814-0180, Japan

    Ryuji Inoue

  5. Institute of Advanced Energy, Kyoto University, Uji, 611-0011, Kyoto, Japan

    Takashi Morii

  6. Department of Pathophysiology, School of Pharmaceutical Sciences, Showa University, Tokyo, 142-8555, Japan

    Shunichi Shimizu

  7. Division of Cellular and Gene Therapy Products, National Institute of Health Sciences, Tokyo, 158-8510, Japan

    Yoji Sato

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T.Y., acquisition, analysis and interpretation of data, and drafting and revision of the manuscript; N.T., S.Y., Y.H., R.I., T.M., M.T., S.S. and Y.S., acquisition, analysis and interpretation of data; Y.M., analysis and interpretation of data, and drafting and critical review of the manuscript.

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Correspondence to Yasuo Mori.

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Yoshida, T., Inoue, R., Morii, T. et al. Nitric oxide activates TRP channels by cysteine S-nitrosylation. Nat Chem Biol 2, 596–607 (2006). https://doi.org/10.1038/nchembio821

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