Molecular basis of the facilitation of the heterooligomeric GIRK1/GIRK4 complex by cAMP dependent protein kinase

Fritz Treiber, Christian Rosker, Tal Keren-Raifman, Bibiane Steinecker, Astrid Gorischek, Nathan Dascal, Wolfgang Schreibmayer

Research output: Contribution to journalArticlepeer-review

Abstract

G-protein activated inwardly rectifying K+ channels (GIRKs) of the heterotetrameric GIRK1/GIRK4 composition mediate IK + ACh in atrium and are regulated by cAMP dependent protein kinase (PKA). Phosphorylation of GIRK1/GIRK4 complexes promotes the activation of the channel by the G-protein Gβγ-dimer ("heterologous facilitation"). Previously we reported that 3 serines/threonines (S/Ts) within the GIRK1 subunit are phosphorylated by the catalytic subunit of PKA (PKA-cs) in-vitro and are responsible for the acute functional effects exerted by PKA on the homooligomeric GIRK1F137S (GIRK1*) channel. Here we report that homooligomeric GIRK4WT and GIRK4S143T (GIRK4*) channels are clearly regulated by PKA phosphorylation. Heterooligomeric channels of the GIRK1S385CS401CT407C/GIRK4 WT composition, where the GIRK1 subunit is devoid of PKA mediated phosphorylation, exhibited reduced but still significant acute effects (reduction during agonist application was ≈ 49% compared to GIRK1 WT/GIRK4WT). Site directed mutagenesis of truncated cytosolic regions of GIRK4 revealed four serines/threonines (S/Ts) that were heavily phosphorylated by PKA-cs in vitro. Two of them were found to be responsible for the acute effects exerted by PKA in vivo, since the effect of cAMP injection was reduced by ≈ 99% in homooligomeric GIRK4 *T199CS412C channels. Coexpression of GIRK1WT/ GIRK4T199CS412C reduced the acute effect by ≈ 65%. Only channels of the GIRK1S385CS401CT407C/GIRK4T199CS412C composition were practically devoid of PKA mediated effects (reduction by ≈ 97%), indicating that both subunits contribute to the heterologous facilitation of IK + ACh.

Original languageEnglish
Pages (from-to)1214-1221
Number of pages8
JournalBiochimica et Biophysica Acta - Biomembranes
Volume1828
Issue number4
DOIs
StatePublished - Apr 2013

Keywords

  • GIRK1
  • GIRK4
  • Heterologous facilitation
  • PKA
  • cAMP

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