Calmodulin regulates Cav3 T-type channels at their gating brake

Jean Chemin, Valentina Taiakina, Arnaud Monteil, Michael Piazza, Wendy Guan, Robert F. Stephens, Ashraf Kitmitto, Zhiping Pang, Annette C. Dolphin, Edward Perez-Reyes, Thorsten Dieckmann, Joseph Guy Guillemette, J. David Spafford

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Calcium (Cav1 and Cav2) and sodium channels possess homologous CaM-binding motifs, known as IQ motifs in their C termini, which associate with calmodulin (CaM), a universal calcium sensor. Cav3 T-type channels, which serve as pacemakers of the mammalian brain and heart, lack a C-terminal IQ motif. We illustrate that T-type channels associate with CaM using co-immunoprecipitation experiments and single particle cryoelectron microscopy. We demonstrate that protostome invertebrate (LCav3) and human Cav3.1, Cav3.2, and Cav3.3 T-type channels specifically associate with CaM at helix 2 of the gating brake in the I–II linker of the channels. Isothermal titration calorimetry results revealed that the gating brake and CaM bind each other with high-nanomolar affinity. We show that the gating brake assumes a helical conformation upon binding CaM, with associated conformational changes to both CaM lobes as indicated by amide chemical shifts of the amino acids of CaM in 1H-15N HSQC NMR spectra. Intact Ca2-binding sites on CaM and an intact gating brake sequence (first 39 amino acids of the I–II linker) were required in Cav3.2 channels to prevent the runaway gating phenotype, a hyperpolarizing shift in voltage sensitivities and faster gating kinetics. We conclude that the presence of high-nanomolar affinity binding sites for CaM at its universal gating brake and its unique form of regulation via the tuning of the voltage range of activity could influence the participation of Cav3 T-type channels in heart and brain rhythms. Our findings may have implications for arrhythmia disorders arising from mutations in the gating brake or CaM.

Original languageEnglish (US)
Pages (from-to)20010-20031
Number of pages22
JournalJournal of Biological Chemistry
Volume292
Issue number49
DOIs
StatePublished - Jan 1 2017

Fingerprint

Calmodulin
Brakes
Brain
Binding Sites
Calcium
Homeless Youth
Cryoelectron Microscopy
Amino Acids
Pacemakers
Calorimetry
Sodium Channels
Chemical shift
Electric potential
Invertebrates
Titration
Immunoprecipitation
Amides
Conformations
Cardiac Arrhythmias
Microscopic examination

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Chemin, J., Taiakina, V., Monteil, A., Piazza, M., Guan, W., Stephens, R. F., ... Spafford, J. D. (2017). Calmodulin regulates Cav3 T-type channels at their gating brake. Journal of Biological Chemistry, 292(49), 20010-20031. https://doi.org/10.1074/jbc.M117.807925
Chemin, Jean ; Taiakina, Valentina ; Monteil, Arnaud ; Piazza, Michael ; Guan, Wendy ; Stephens, Robert F. ; Kitmitto, Ashraf ; Pang, Zhiping ; Dolphin, Annette C. ; Perez-Reyes, Edward ; Dieckmann, Thorsten ; Guillemette, Joseph Guy ; Spafford, J. David. / Calmodulin regulates Cav3 T-type channels at their gating brake. In: Journal of Biological Chemistry. 2017 ; Vol. 292, No. 49. pp. 20010-20031.
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Chemin, J, Taiakina, V, Monteil, A, Piazza, M, Guan, W, Stephens, RF, Kitmitto, A, Pang, Z, Dolphin, AC, Perez-Reyes, E, Dieckmann, T, Guillemette, JG & Spafford, JD 2017, 'Calmodulin regulates Cav3 T-type channels at their gating brake', Journal of Biological Chemistry, vol. 292, no. 49, pp. 20010-20031. https://doi.org/10.1074/jbc.M117.807925

Calmodulin regulates Cav3 T-type channels at their gating brake. / Chemin, Jean; Taiakina, Valentina; Monteil, Arnaud; Piazza, Michael; Guan, Wendy; Stephens, Robert F.; Kitmitto, Ashraf; Pang, Zhiping; Dolphin, Annette C.; Perez-Reyes, Edward; Dieckmann, Thorsten; Guillemette, Joseph Guy; Spafford, J. David.

In: Journal of Biological Chemistry, Vol. 292, No. 49, 01.01.2017, p. 20010-20031.

Research output: Contribution to journalArticle

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T1 - Calmodulin regulates Cav3 T-type channels at their gating brake

AU - Chemin, Jean

AU - Taiakina, Valentina

AU - Monteil, Arnaud

AU - Piazza, Michael

AU - Guan, Wendy

AU - Stephens, Robert F.

AU - Kitmitto, Ashraf

AU - Pang, Zhiping

AU - Dolphin, Annette C.

AU - Perez-Reyes, Edward

AU - Dieckmann, Thorsten

AU - Guillemette, Joseph Guy

AU - Spafford, J. David

PY - 2017/1/1

Y1 - 2017/1/1

N2 - Calcium (Cav1 and Cav2) and sodium channels possess homologous CaM-binding motifs, known as IQ motifs in their C termini, which associate with calmodulin (CaM), a universal calcium sensor. Cav3 T-type channels, which serve as pacemakers of the mammalian brain and heart, lack a C-terminal IQ motif. We illustrate that T-type channels associate with CaM using co-immunoprecipitation experiments and single particle cryoelectron microscopy. We demonstrate that protostome invertebrate (LCav3) and human Cav3.1, Cav3.2, and Cav3.3 T-type channels specifically associate with CaM at helix 2 of the gating brake in the I–II linker of the channels. Isothermal titration calorimetry results revealed that the gating brake and CaM bind each other with high-nanomolar affinity. We show that the gating brake assumes a helical conformation upon binding CaM, with associated conformational changes to both CaM lobes as indicated by amide chemical shifts of the amino acids of CaM in 1H-15N HSQC NMR spectra. Intact Ca2-binding sites on CaM and an intact gating brake sequence (first 39 amino acids of the I–II linker) were required in Cav3.2 channels to prevent the runaway gating phenotype, a hyperpolarizing shift in voltage sensitivities and faster gating kinetics. We conclude that the presence of high-nanomolar affinity binding sites for CaM at its universal gating brake and its unique form of regulation via the tuning of the voltage range of activity could influence the participation of Cav3 T-type channels in heart and brain rhythms. Our findings may have implications for arrhythmia disorders arising from mutations in the gating brake or CaM.

AB - Calcium (Cav1 and Cav2) and sodium channels possess homologous CaM-binding motifs, known as IQ motifs in their C termini, which associate with calmodulin (CaM), a universal calcium sensor. Cav3 T-type channels, which serve as pacemakers of the mammalian brain and heart, lack a C-terminal IQ motif. We illustrate that T-type channels associate with CaM using co-immunoprecipitation experiments and single particle cryoelectron microscopy. We demonstrate that protostome invertebrate (LCav3) and human Cav3.1, Cav3.2, and Cav3.3 T-type channels specifically associate with CaM at helix 2 of the gating brake in the I–II linker of the channels. Isothermal titration calorimetry results revealed that the gating brake and CaM bind each other with high-nanomolar affinity. We show that the gating brake assumes a helical conformation upon binding CaM, with associated conformational changes to both CaM lobes as indicated by amide chemical shifts of the amino acids of CaM in 1H-15N HSQC NMR spectra. Intact Ca2-binding sites on CaM and an intact gating brake sequence (first 39 amino acids of the I–II linker) were required in Cav3.2 channels to prevent the runaway gating phenotype, a hyperpolarizing shift in voltage sensitivities and faster gating kinetics. We conclude that the presence of high-nanomolar affinity binding sites for CaM at its universal gating brake and its unique form of regulation via the tuning of the voltage range of activity could influence the participation of Cav3 T-type channels in heart and brain rhythms. Our findings may have implications for arrhythmia disorders arising from mutations in the gating brake or CaM.

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Chemin J, Taiakina V, Monteil A, Piazza M, Guan W, Stephens RF et al. Calmodulin regulates Cav3 T-type channels at their gating brake. Journal of Biological Chemistry. 2017 Jan 1;292(49):20010-20031. https://doi.org/10.1074/jbc.M117.807925