G protein-gated inhibitory module of N-type (ca(v)2.2) ca2+ channels

Neuron. 2005 Jun 16;46(6):891-904. doi: 10.1016/j.neuron.2005.05.011.

Abstract

Voltage-dependent G protein (Gbetagamma) inhibition of N-type (CaV2.2) channels supports presynaptic inhibition and represents a central paradigm of channel modulation. Still controversial are the proposed determinants for such modulation, which reside on the principal alpha1B channel subunit. These include the interdomain I-II loop (I-II), the carboxy tail (CT), and the amino terminus (NT). Here, we probed these determinants and related mechanisms, utilizing compound-state analysis with yeast two-hybrid and mammalian cell FRET assays of binding among channel segments and G proteins. Chimeric channels confirmed the unique importance of NT. Binding assays revealed selective interaction between NT and I-II elements. Coexpressing NT peptide with Gbetagamma induced constitutive channel inhibition, suggesting that the NT domain constitutes a G protein-gated inhibitory module. Such inhibition was limited to NT regions interacting with I-II, and G-protein inhibition was abolished within alpha1B channels lacking these NT regions. Thus, an NT module, acting via interactions with the I-II loop, appears fundamental to such modulation.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Calcium Channels, L-Type / chemistry
  • Calcium Channels, L-Type / physiology*
  • Calcium Channels, N-Type
  • Cell Line
  • Electric Stimulation
  • GTP-Binding Protein beta Subunits / metabolism
  • GTP-Binding Protein gamma Subunits / metabolism
  • GTP-Binding Proteins / metabolism*
  • Humans
  • Ion Channel Gating / physiology*
  • Membrane Potentials / physiology
  • Molecular Biology / methods
  • Neural Inhibition / physiology*
  • Patch-Clamp Techniques / methods
  • Protein Binding / physiology
  • Protein Structure, Tertiary / physiology
  • Protein Subunits / metabolism
  • Rabbits
  • Radioligand Assay / methods
  • Rats
  • Recombinant Fusion Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Structure-Activity Relationship
  • Transfection / methods
  • Two-Hybrid System Techniques
  • Yeasts

Substances

  • CACNA1B protein, human
  • Calcium Channels, L-Type
  • Calcium Channels, N-Type
  • GTP-Binding Protein beta Subunits
  • GTP-Binding Protein gamma Subunits
  • Protein Subunits
  • Recombinant Fusion Proteins
  • GTP-Binding Proteins