Apical sorting of a voltage- and Ca2+-activated K+ channel alpha-subunit in Madin-Darby canine kidney cells is independent of N-glycosylation

The voltage- and Ca2+-acttiated K+ (K-V,K-Ca) channel is expressed in a var iety of polarized epithelial cells seemingly displaying a tissue-dependent apical-to-basolateral regionalization, as revealed by electrophysiology. Us ing domain-specific biotinylation and immunofluorescence we show that the h uman channel K-V,K-Ca alpha -subunit (human Slowpoke channel, hSlo) is pred ominantly found in the apical plasma membrane domain of permanently transfe cted Madin-Darby canine kidney cells. Both the wild-type and a mutant hSlo protein lacking its only potential N-glycosylation site were efficiently tr ansported to the cell surface and concentrated in the apical domain even wh en they were overexpressed to levels 200- to 300-fold higher than the densi ty of intrinsic Slo channels. Furthermore, tunicamycin treatment did not pr event apical segregation of hSlo, indicating that endogenous glycosylated p roteins (e.g., K-V,K-Ca beta -subunits) were not required. hSlo seems to di splay properties for lipid-raft targeting, as judged by its buoyant distrib ution in sucrose gradients after extraction with either detergent or sodium carbonate. The evidence indicates that the hSlo protein possesses intrinsi c information for transport to the apical cell surface through a mechanism that may involve association with lipid rafts and that is independent of gl ycosylation of the channel itself or an associated protein. Thus, this part icular polytopic model protein shows that glycosylation-independent apical pathways exist for endogenous membrane proteins in Madin-Darby canine kidne y cells.