Molecular determinants of PKA-dependent inhibition of TRPC5 channel

Am J Physiol Cell Physiol. 2011 Oct;301(4):C823-32. doi: 10.1152/ajpcell.00351.2010. Epub 2011 Jul 6.


Canonical transient receptor potential (TRPC) channels are Ca(2+)-permeable, nonselective cation channels that are widely expressed in numerous cell types. Here, we demonstrate a new mechanism of TPRC isofom 5 (TRPC5) regulation, via cAMP signaling via Gα(s). Monovalent cation currents in human embryonic kidney-293 cells transfected with TRPC5 were induced by G protein activation with intracellular perfusion of GTPγS or by muscarinic stimulation. This current could be inhibited by a membrane-permeable analog of cAMP, 8-bromo-cAMP, by isoproterenol, by a constitutively active form of Gα(s) [Gα(s) (Q227L)], and by forskolin. These inhibitory effects were blocked by the protein kinase A (PKA) inhibitors, KT-5720 and H-89, as well as by two point mutations at consensus PKA phosphorylation sites on TRPC5 (S794A and S796A). Surface expression of several mutated versions of TRPC5, quantified using surface biotinylation, were not affected by Gα(s) (Q227L), suggesting that trafficking of this channel does not underlie the regulation we report. This mechanism of inhibition was also found to be important for the closely related channel, TRPC4, in particular for TRPC4α, although TRPC4β was also affected. However, this form of regulation was not found to be involved in TRPC6 and transient receptor potential vanilloid 6 function. In murine intestinal smooth muscle cells, muscarinic stimulation-induced cation currents were mediated by TRPC4 (>80%) and TRPC6. In murine intestinal smooth muscle cells, 8-bromo-cAMP, adrenaline, and isoproterenol decreased nonselective cation currents activated by muscarinic stimulation or GTPγS. Together, these results suggest that TRPC5 is directly phosphorylated by G(s)/cAMP/PKA at positions S794 and S796. This mechanism may be physiologically important in visceral tissues, where muscarinic receptor and β(2)-adrenergic receptor are involved in the relaxation and contraction of smooth muscles.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 8-Bromo Cyclic Adenosine Monophosphate / pharmacology
  • Animals
  • Carbachol / pharmacology
  • Cyclic AMP / metabolism
  • Cyclic AMP-Dependent Protein Kinases / metabolism*
  • GTP-Binding Protein alpha Subunits, Gs / genetics
  • GTP-Binding Protein alpha Subunits, Gs / metabolism*
  • Gene Expression Regulation / physiology*
  • Guanosine 5'-O-(3-Thiotriphosphate) / pharmacology
  • HEK293 Cells
  • Humans
  • Ion Channel Gating / drug effects
  • Membrane Potentials
  • Mice
  • Muscle, Skeletal / drug effects
  • Muscle, Skeletal / physiology
  • TRPC Cation Channels / genetics
  • TRPC Cation Channels / metabolism*


  • TRPC Cation Channels
  • TRPC5 protein, human
  • 8-Bromo Cyclic Adenosine Monophosphate
  • Guanosine 5'-O-(3-Thiotriphosphate)
  • Carbachol
  • Cyclic AMP
  • Cyclic AMP-Dependent Protein Kinases
  • GTP-Binding Protein alpha Subunits, Gs