The leucine-rich repeat domains of BK channel auxiliary ? subunits regulate their expression, trafficking, and channel-modulation functions

As high-conductance calcium-and voltage-dependent potassium channels, BK channels consist of pore-forming, voltage-, and Ca2+-sensing alpha and auxiliary subunits. The leucine-rich repeat (LRR) domain-containing auxiliary gamma subunits potently modulate the voltage dependence of BK channel activation. Despite their dominant size in whole protein masses, the function of the LRR domain in BK channel gamma subunits is unknown. We here investigated the function of these LRR domains in BK channel modulation by the auxiliary gamma 1-3 (LRRC26, LRRC52, and LRRC55) subunits. Using cell surface protein immunoprecipitation, we validated the predicted extracellular localization of the LRR domains. We then refined the structural models of mature proteins on the membrane via molecular dynamic simulations. By replacement of the LRR domain with extracellular regions or domains of non-LRR proteins, we found that the LRR domain is nonessential for the maximal channel-gating modulatory effect but is necessary for the all-or-none phenomenon of BK channel modulation by the gamma 1 subunit. Mutational and enzymatic blockade of N-glycosylation in the gamma 1-3 subunits resulted in a reduction or loss of BK channel modulation by gamma subunits. Finally, by analyzing their expression in whole cells and on the plasma membrane, we found that blockade of N-glycosylation drastically reduced total expression of the gamma 2 subunit and the cell surface expression of the gamma 1 and gamma 3 subunits. We conclude that the LRR domains play key roles in the regulation of the expression, cell surface trafficking, and channel-modulation functions of the BK channel gamma subunits.

Automated summary

In this study, the function of the LRR domains in modulation of BK channels by auxiliary gamma subunits was investigated. The results showed that the LRR domain is essential for the all-or-none phenomenon of BK channel modulation by the gamma 1 subunit. Furthermore, N-glycosylation plays a crucial role in the modulation of BK channels by the gamma subunits.