The Interaction between the I-II Loop and the III-IV Loop of Cav2.1 Contributes to Voltage-dependent Inactivation in a β-Dependent Manner*

Abstract

We have investigated the molecular mechanisms whereby the I-II loop controls voltage-dependent inactivation in P/Q calcium channels. We demonstrate that the I-II loop is localized in a central position to control calcium channel activity through the interaction with several cytoplasmic sequences; including the III-IV loop. Several experiments reveal the crucial role of the interaction between the I-II loop and the III-IV loop in channel inactivation. First, point mutations of two amino acid residues of the I-II loop of Cav2.1 (Arg-387 or Glu-388) facilitate voltage-dependent inactivation. Second, overexpression of the III-IV loop, or injection of a peptide derived from this loop, produces a similar inactivation behavior than the mutated channels. Third, the III-IV peptide has no effect on channels mutated in the I-II loop. Thus, both point mutations and overexpression of the III-IV loop appear to act similarly on inactivation, by competing off the native interaction between the I-II and the III-IV loops of Cav2.1. As they are known to affect inactivation, we also analyzed the effects of β subunits on these interactions. In experiments in which the β4 subunit is co-expressed, the III-IV peptide is no longer able to regulate channel inactivation. We conclude that (i) the contribution of the I-II loop to inactivation is partly mediated by an interaction with the III-IV loop and (ii) the β subunits partially control inactivation by modifying this interaction. These data provide novel insights into the mechanisms whereby the β subunit, the I-II loop, and the III-IV loop altogether can contribute to regulate inactivation in high voltage-activated calcium channels.

  • Abbreviations:
    AID
    α1 interaction domain
    GST
    glutathioneS-transferase
    Fmoc
    N-(9-fluorenyl)methoxycarbonyl
    NMP
    N-methylpyrrolidinone
    HPLC
    high performance liquid chromatography
    HIT
    half-inactivation time
    HIP
    half-inactivation potential
    HAP
    half-activation potential
    F
    fahrad(s)
    • Received July 5, 2001.
    • Revision received December 11, 2001.
    Table of Contents

    This Article

    1. The Journal of Biological Chemistry 277, 10003-10013.
    1. All Versions of this Article:
      1. M106231200v1
      2. 277/12/10003 (most recent)

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