Abstract
Kir2 subfamily members of inward rectifier K+ channel are known to co- assemble to form heteromultimers, and it is also known for Kir3. Here we examined whether Kir2.1 (IRK1) and Kir3.4 (GIRK4) belonging to different subfamilies can assemble each other or not. First, we examined the association by co-immunoprecipitation experiments using FLAG or myc tagged constructs co-transfected in HEK 293 cells. We observed "GIRK4-FLAG and IRK1-myc" as well as "IRK1-FLAG and GIRK4-myc" co-immunoprecipitated at a comparable level with a positive control pair, IRK1-FLAG/IRK1-myc. This clear co-immunoprecipitation was not observed in a negative control pair, P2X2 receptor-FLAG/GIRK4-myc. As a next step, we anlayzed electrophysiologically using Xenopus oocyte expression system the effect of co-injection of GIRK4 or GIRK4/GIRK1 cRNA on IRK1 current. We could not record a clear emergence of current with unique features which reflects formation of functional heteromultimers, but observed a decrease in the amplitude of IRK1 current, suggesting a suppression effect possibly by hetero-multimerization. Finally, we carried out FRET analysis of IRK1-CFP/GIRK4-YFP pair and GIRK1-CFP/IRK1-YFP pair expressed in CHO cells, and obtained preliminary data supporting association of these two subunits. Taken together, these results suggest that IRK1 and GIRK4 subunits have a capability to form heteromultimers in heterologous expression systems. [J Physiol Sci. 2006;56 Suppl:S151]
