Preassociated apocalmodulin mediates Ca2+-dependent sensitization of activation and inactivation of TMEM16A/16B Ca2+-gated Cl- channels

Proc Natl Acad Sci U S A. 2014 Dec 23;111(51):18213-8. doi: 10.1073/pnas.1420984111. Epub 2014 Dec 8.

Abstract

Ca(2+)-activated chloride currents carried via transmembrane proteins TMEM16A and TMEM16B regulate diverse processes including mucus secretion, neuronal excitability, smooth muscle contraction, olfactory signal transduction, and cell proliferation. Understanding how TMEM16A/16B are regulated by Ca(2+) is critical for defining their (patho)/physiological roles and for rationally targeting them therapeutically. Here, using a bioengineering approach--channel inactivation induced by membrane-tethering of an associated protein (ChIMP)--we discovered that Ca(2+)-free calmodulin (apoCaM) is preassociated with TMEM16A/16B channel complexes. The resident apoCaM mediates two distinct Ca(2+)-dependent effects on TMEM16A, as revealed by expression of dominant-negative CaM1234. These effects are Ca(2+)-dependent sensitization of activation (CDSA) and Ca(2+)-dependent inactivation (CDI). CDI and CDSA are independently mediated by the N and C lobes of CaM, respectively. TMEM16A alternative splicing provides a mechanism for tuning apoCaM effects. Channels lacking splice segment b selectively lost CDI, and segment a is necessary for apoCaM preassociation with TMEM16A. The results reveal multidimensional regulation of TMEM16A/16B by preassociated apoCaM and introduce ChIMP as a versatile tool to probe the macromolecular complex and function of Ca(2+)-activated chloride channels.

Keywords: TMEM16A; anoctamin1; calcium-activated chloride channel; calmodulin.

Publication types

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

MeSH terms

  • Anoctamin-1
  • Anoctamins
  • Calcium / metabolism*
  • Calmodulin / physiology*
  • Chloride Channels / antagonists & inhibitors
  • Chloride Channels / genetics
  • Chloride Channels / metabolism*
  • Chloride Channels / physiology
  • Humans
  • Ion Channel Gating / physiology*
  • Membrane Proteins / genetics
  • Membrane Proteins / physiology*
  • Neoplasm Proteins / genetics
  • Neoplasm Proteins / physiology*
  • RNA Splicing

Substances

  • ANO1 protein, human
  • ANO2 protein, human
  • Anoctamin-1
  • Anoctamins
  • Calmodulin
  • Chloride Channels
  • Membrane Proteins
  • Neoplasm Proteins
  • Calcium