Glycosylation Of Beta 1 Subunit Plays A Pivotal Role In The Toxin Sensitivity And Activation Of Bk Channels

Background: The accessory beta 1 subunits, regulating the pharmacological and biophysical properties of BK channels, always undergo post-translational modifications, especially glycosylation. To date, it remains elusive whether the glycosylation contributes to the regulation of BK channels by beta 1 subunits.Methods: Herein, we combined the electrophysiological approach with molecular mutations and biochemical manipulation to investigate the function roles of N-glycosylation in beta 1 subunits.Results: The results show that deglycosylation of beta 1 subunits through double-site mutations (beta 1 N80A/N142A or beta 1 N80Q/N142Q) could significantly increase the inhibitory potency of iberiotoxin, a specific BK channel blocker. The deglycosylated channels also have a different sensitivity to martentoxin, another BK channel modulator with some remarkable effects as reported before. On the contrary to enhancing effects of martentoxin on glycosylated BK channels under the presence of cytoplasmic Ca2+, deglycosylated channels were not affected by the toxin. However, the deglycosylated channels were surprisingly inhibited by martentoxin under the absence of cytoplasmic Ca2+, while the glycosylated channels were not inhibited under this same condition. In addition, wild type BK (alpha+beta 1) channels treated with PNGase F also showed the same trend of pharmacological results to the mutants. Similar to this modulation of glycosylation on BK channel pharmacology, the deglycosylated forms of the channels were activated at a faster speed than the glycosylated ones. However, the V-1/2 and slope were not changed by the glycosylation.Conclusion: The present study reveals that glycosylation is an indispensable determinant of the modulation of beta 1-subunit on BK channel pharmacology and its activation. The loss of glycosylation of beta 1 subunits could lead to the dysfunction of BK channel, resulting in a pathological state.