Aims: 

It has been shown previously that G-Protein dependent inwardly rectifying K⁺-channels (GIRK`s), are regulated by cAMP dependent proteinkinase (PKA) and by proteinphosphatase 2A (PP2A; "heterologous facilitation"). Both the GIRK1 and the GIRK4 subunit have been identified as prominent targets for PKA phosphorylation and the structural determinants on the channel subunits were identified. Aim of the current study was to investigate the roles of the individual subunits in the heterologous facilitation of IK+ACh and in membrane trafficking of the channel complex.

Methods: 

Wild type and phosphorylation deficient GIRK1 and GIRK4 subunits were heterologously expressed in oocytes of Xenopus laevis and the effect of PKA phosphorylation on agonist induced as well as on basal currents was quatitated for different subunit compositions. Membrane trafficking of GFP tagged subunits, following activation and deactivation of PKA was studied by confocal microscopy.

Results: 

It was found, that phosphorylation of the GIRK4 subunit is most important for the instantaneous increase in the K⁺ current observed (heterologous facilitation), while GIRK1 subunit phosphorylation contributes little. Instead phosphorylation of the GIRK1 subunit turned out to be crucial for membrane localization and trafficking of the channel complex: it was found, that channel complexes containing WT GIRK1 get internalized 1-2 h after cytosolic injection of cAMP or Sp-cAMPS. In turn cytosolic injection of the PKA antagonist Rp-cAMPS produced a marked increase in surface localization.

Conclusion: 

These results demonstrate the importance of individual S/T's on both the GIRK1 and the GIRK4 subunit in regulation of channel activation as well as in membrane trafficking.