Role of the Alpha-Kinase Domain in Transient Receptor Potential Melastatin 6 Channel and Regulation by Intracellular ATP

J Biol Chem. 2008 Jul 18;283(29):19999-20007. doi: 10.1074/jbc.M800167200. Epub 2008 May 19.

Abstract

Transient receptor potential melastatin 6 (TRPM6) plays an essential role in epithelial Mg(2+) transport. TRPM6 and its closest homologue, TRPM7, both combine a cation channel with an alpha-kinase domain. However, the role of this alpha-kinase domain in TRPM6 channel activity remains elusive. The aim of this study was to investigate the regulation of TRPM6 channel activity by intracellular ATP and the involvement of its alpha-kinase domain. We demonstrated that intracellular Na- and Mg-ATP decreased the TRPM6 current in HEK293 cells heterogeneously expressing the channel, whereas Na-CTP or Na-GTP had no effect on channel activity. Whole cell recordings in TRPM6-expressing HEK293 cells showed that deletion of the alpha-kinase domain prevented the inhibitory effect of intracellular ATP without abrogating channel activity. Mutation of the conserved putative ATP-binding motif GXG(A)XXG (G1955D) in the alpha-kinase domain of TRPM6 inhibited the ATP action, whereas this effect remained preserved in the TRPM6 phosphotransferase-deficient mutant K1804R. Mutation of the TRPM6 autophosphorylation site, Thr(1851), into either an alanine or an aspartate, resulted in functional channels that could still be inhibited by ATP. In conclusion, intracellular ATP regulates TRPM6 channel activity via its alpha-kinase domain independently of alpha-kinase activity.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / metabolism*
  • Amino Acid Motifs
  • Amino Acid Sequence
  • Cell Line
  • Electrophysiology
  • Humans
  • Models, Molecular
  • Molecular Sequence Data
  • Mutation / genetics
  • Patch-Clamp Techniques
  • Protein Kinases / chemistry
  • Protein Kinases / genetics
  • Protein Kinases / metabolism*
  • Protein Structure, Tertiary
  • Substrate Specificity
  • TRPM Cation Channels / chemistry
  • TRPM Cation Channels / genetics
  • TRPM Cation Channels / metabolism*

Substances

  • TRPM Cation Channels
  • TRPM6 protein, human
  • Adenosine Triphosphate
  • Protein Kinases