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A channelopathy mechanism revealed by direct calmodulin activation of TrpV4

Stephen H Loukin1, Jinfeng Teng2, Ching Kung3

  • 1Laboratory of Molecular Biology, University of Wisconsin, Madison, WI 53706; ckung@wisc.edu shloukin@wisc.edu.

Proceedings of the National Academy of Sciences of the United States of America
|July 15, 2015
PubMed

Insights

Calcium-calmodulin (CaM) regulates Transient Receptor Potential vanilloid subtype 4 (TrpV4) channels. Mutations causing skeletal dysplasia (SD) disrupt this regulation, leading to constitutive channel activity and disease.

Area of Science:

  • Molecular biology
  • Ion channel physiology
  • Biophysics

Background:

  • Calcium-calmodulin (CaM) is a key regulator of numerous ion channels, including Transient Receptor Potential vanilloid subtype 4 (TrpV4).
  • Previous models suggest Ca(2+)-CaM binding to the C-terminal domain enhances TrpV4 channel activity.
  • Over 50 human mutations in TrpV4 are linked to heritable skeletal dysplasia (SD) and other diseases.

Purpose of the Study:

  • To investigate the regulatory mechanism of Ca(2+)-calmodulin (CaM) on Transient Receptor Potential vanilloid subtype 4 (TrpV4) channel activity.
  • To elucidate the role of the region upstream of the Ca(2+)-CaM binding domain (CBD) in TrpV4 channel gating.
  • To understand how skeletal dysplasia (SD)-associated mutations affect TrpV4 channel function and CaM regulation.

Main Methods:

  • Direct application of Ca(2+)-CaM protein to excised membrane patches from oocytes expressing TrpV4.
  • Analysis of TrpV4 channel activity in wild-type and mutant channels, including specific SD alleles (E797K, P799L).
  • Functional characterization of a TrpV4 construct with a deleted upstream peptide (Δ795-804).

Main Results:

  • Directly confirmed Ca(2+)-CaM binding enhances TrpV4 activity in excised patches.
  • SD-associated mutations E797K and P799L, located upstream of the CBD, showed increased basal activity but reduced Ca(2+)-CaM responsiveness.
  • Deletion of the upstream peptide (Δ795-804) resulted in constitutive TrpV4 channel activity and abolished Ca(2+)-CaM response.
  • Gain-of-function effects of SD mutations correlate with disease severity.

Conclusions:

  • The region upstream of the TrpV4 CBD acts as an autoinhibitory domain, maintaining the channel in a closed state.
  • Ca(2+)-CaM binding to the CBD likely relieves this autoinhibition through steric interference.
  • TrpV4 mutations causing skeletal dysplasia may lead to constitutive channel leakage by destabilizing this closed conformation, potentially through removal of autoinhibition.
Keywords:
TrpV1autoinhibitioncalciumion channelskeletal dysplasia

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