The MORN domain of Junctophilin2 regulates functional interactions with small-conductance Ca2+ -activated potassium channel subtype2 (SK2).

ABSTRACT

Small-conductance Ca2+ -activated K+ channel subtype2 (SK2) are stable macromolecular complexes that regulate myocardial excitability and Ca2+ homeostasis. Junctophilin-2 (JP2) is a membrane-binding protein, which provides functional crosstalk by physically linking with the cell-surface and intracellular ion channels. We previously demonstrated that the MORN domain of JP2 interacts with SK2 channels. However, the roles of the JP2 MORN domain in regulating the precise subcellular localization and molecular modulation of SK2 have not yet been incompletely understood. In the present study, in vitro and in vivo assays were used to confirm the physical interactions between the SK2 channel and JP2 in H9c2 and HEK293 cells, with a concentration on the association between the C-terminus of SK2 channels and the MORN domain of JP2. Furthermore, the membrane expression of SK2 were found to be significantly impaired by the mutation or knockdown of JP2. Using immunofluorescence staining along with Golgi/early endosome markers, we studied the mechanisms of JP2-regulated SK2 membrane trafficking, which indicates that the JP2 MORN domain is probably necessary for the retrograde trafficking of SK2 channels. The functional study demonstrates that whole cell SK2 current densities recorded from the HEK293 cells co-expressing the JP2-MORN domain with SK2 were significantly augmented, compared with cells expressing SK2 alone. Our findings suggest that the MORN domain of JP2 directly modulates SK2 channel current amplitude and trafficking, through its interaction with an overlapping region of the JP2 MORN domain on the SK2 C-terminus.