Identification and quantification of full‐length BK channel variants in the developing mouse cochlea

Abstract

Maxi K+ (BK) channel diversity is attributed to alternative splicing in the kcnma1 gene. The resultant variants manifest themselves in different cell types, tissues and functions such as excitation, metabolism, and signaling. Immuno-electron microscopy revealed immunogold particle labeling of BK in apical and basal regions of inner and outer hair cells, respectively. Additional labeling occurs in Deiters’ cells and the inner mitochondrial membrane. Identification of full-length sequences reveals 27 BK variants from embryonic and postnatal mouse inner ear, per classification by tail motif, VYR, DEC, and ERL, and by exon usage. Three predicted start codons are found encoding MAN, MSS, and MDA, of which MDA shows the greatest expression through all stages in development, whereas MAN is undetectable. Complex splice sites occur between exons 9 and 10, and 21 and 23. Spliced-in/out exons between 8 and 10 reveal a short fragment composed of exons 8+10, detectable on postnatal-day (PD) 14 and PD30, and a longer fragment composed of exons 8+9+10 that upregulates on embryonic-day (ED) 14. Spliced-in exons 22 or 23 are expressed on ED14 but decrease over time; however, exon 22 increases again on PD34. Using tail specific primers, qRT-PCR from ED14, PD4, 14, and 30, show that BK-VYR and -ERL dominate expression on ED14, whereas DEC dominates after birth in all cochlear regions. The localization of BK and the changes in expression of its exons and tail types, by alternative splicing during development, may contribute to cochlear organization, acquisition of hearing, and intracellular signaling.

Calcium activated K+ channels in the electroreceptor of the skate confirmed by cloning. Details of subunits and splicing

“…As shown in Fig 6a, a sequence of 26-28 homologous amino acids that can be identified as exon 29 is present in the chick cochlea [22] and the skate electroreceptor. A virtually identical sequence was identified as alternative exon B in an early description of the mouse B K channel [7] and later in the mouse cochlea [38] and the rat [17,35]. Fluorescence imaging studies have shown that exon 29 occurs in inner hair cells of the mouse cochlea, which are the primary auditory receptor cells [5].…”

Transduction of Voltage and Ca2+Signals by Slo1 BK Channels

“…A short REVIEWS PHYSIOLOGY • Volume 28 • March 2013 • www.physiologyonline.org stretch of ϳ20 residues connects the transmembrane helix S6 and RCK1 (S6-RCK1 linker), whereas ϳ100 or more residues bridge RCK1 and RCK2 (RCK1-RCK2 linker). Several splice variants that differ in the S6-RCK1 linker, the RCK1-RCK2 linker, and/or the distal COOH-terminal areas are known (125).…”

“…For example, although only one gene codes for the pore-forming subunit (KCNMA1, Slo1), its transcript is extensively spliced to create a vastly large number of variant polypeptides (1,36,40,141). Nearly 1,000 distinct full-length polypeptides may be theoretically available to form tetrameric BK channels in mice (125). Coassembly with the auxiliary subunits ␤1, ␤2, ␤3, ␤4, and leucine-rich repeat-containing proteins (LRRCs; ␥ subunits) also increases functional diversity by altering the channel's apparent sensitivity to Ca 2ϩ and V m as well as their kinetic properties including activation, deactivation, and in some cases conferring inactivation (11,17,156,161,165,170,176,177).…”

Conserved BK Channel-Protein Interactions Reveal Signals Relevant to Cell Death and Survival

“…NMDA2B receptors are present in spiral ganglion cells and adjacent to inner and outer hair cells of human cochlea [39]. Similarly, BK channels are found in ganglion cells and at the base of outer hair cells [19].…”

“…Both BK mito and K ATP channels are thought to protect against ischemia by inhibiting the PTP, either by direct physical interaction or by inhibiting the factors that initiate the opening of the PTP [40]. Recent evidence suggests the presence of BK mito in the cochlea [9], [19], although its relation to ATP synthase and the PTP requires further studies.…”