title: A New Splice Variant of Large Conductance Ca2+-activated K+ (BK) Channel α Subunit Alters Human Chondrocyte Function.
abstract: Large conductance Ca2+-activated K+(BK) channels play essential roles in both excitable and non-excitable cells. For example, in chondrocytes, agonist-induced Ca2+release from intracellular store activates BK channels, and this hyperpolarizes these cells, augments Ca2+entry, and forms a positive feed-back mechanism for Ca2+signaling and stimulation-secretion coupling. In the present study, functional roles of a newly identified splice variant in the BK channel α subunit (BKαΔe2) were examined in a human chondrocyte cell line, OUMS-27, and in a HEK293 expression system. Although BKαΔe2 lacks exon2, which codes the intracellular S0-S1 linker (Glu-127-Leu-180), significant expression was detected in several tissues from humans and mice. Molecular image analyses revealed that BKαΔe2 channels are not expressed on plasma membrane but can traffic to the plasma membrane after forming hetero-tetramer units with wild-type BKα (BKαWT). Single-channel current analyses demonstrated that BKα hetero-tetramers containing one, two, or three BKαΔe2 subunits are functional. These hetero-tetramers have a smaller single channel conductance and exhibit lower trafficking efficiency than BKαWT homo-tetramers in a stoichiometry-dependent manner. Site-directed mutagenesis of residues in exon2 identified Helix2 and the linker to S1 (Trp-158-Leu-180, particularly Arg-178) as an essential segment for channel function including voltage dependence and trafficking. BKαΔe2 knockdown in OUMS-27 chondrocytes increased BK current density and augmented the responsiveness to histamine assayed as cyclooxygenase-2 gene expression. These findings provide significant new evidence that BKαΔe2 can modulate cellular responses to physiological stimuli in human chondrocyte and contribute under pathophysiological conditions, such as osteoarthritis.
authors.affiliation: From the Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabedori, Mizuhoku, Nagoya 467-8603, Japan.
authors.forename: Yoshiaki
authors.identifier: http://orcid.org/0000-0002-7879-6059
authors.initials: Y
authors.lastname: Suzuki
authors.affiliation: From the Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabedori, Mizuhoku, Nagoya 467-8603, Japan.|the Department of Pharmacology, Division of Pathological Sciences, Kyoto Pharmaceutical University, Kyoto 607-8414, Japan, and.
authors.forename: Susumu
authors.initials: S
authors.lastname: Ohya
authors.affiliation: From the Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabedori, Mizuhoku, Nagoya 467-8603, Japan.
authors.forename: Hisao
authors.initials: H
authors.lastname: Yamamura
authors.affiliation: the Faculties of Kinesiology and Medicine, University of Calgary, Calgary, Alberta T2N 1N4, Canada.
authors.forename: Wayne R
authors.initials: WR
authors.lastname: Giles
authors.affiliation: From the Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabedori, Mizuhoku, Nagoya 467-8603, Japan, yimaizum@phar.nagoya-cu.ac.jp.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Imaizumi
mesh_terms: D017398:Alternative Splicing; D000818:Animals; D019902:Chondrocytes / Q000378:metabolism* / Q000473:pathology; D057809:HEK293 Cells; D006801:Humans; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D016297:Mutagenesis, Site-Directed; D010003:Osteoarthritis / Q000235:genetics / Q000378:metabolism* / Q000473:pathology
chemical_list: C494841:KCNMA1 protein, human; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits
keywords: alternative splicing; chondrocyte; fluorescence resonance energy transfer (FRET); molecular imaging; patch clamp; potassium channel; single molecule biophysics