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PERSPECTIVES |
1 Department of Biology, Clarkson University, Potsdam, NY 13676, USA
Email: emoczydl{at}clarkson.edu
The large conductance, voltage- and Ca2+-activated K+ channel is known by many aliases: maxiK, BK, Slo, KCa1.1, even ‘the mammoth’ (Ledoux et al. 2006). This tetrameric K+ channel protein invites casual comparison to a hulking beast by virtue of an impressive unitary conductance (
250 pS) and hefty molecular mass of its 
-subunit (125 kDa). Given that K+ channels are one of the most diverse gene families in the human genome and comprise at least 75 recognized genes belonging to seven distinct subfamilies (Gutman et al. 2003), it may seem odd that maxiK is the product of only a single gene, KCNMA1 or HSlo. Not all is as it seems, however, since genetic simplicity belies the complexity of maxiK's diverse physiological roles and regulatory guises (Lu et al. 2006).
In smooth muscle and neurons, maxiK functions as a negative feedback regulator. Wielding potent repolarizing power, maxiK channels attenuate excitatory signals that increase intracellular Ca2+ by opening in response to local Ca2+ concentration at sites near the intracellular face of the plasma membrane. Due to allosteric coupling of its Ca2+-sensing and voltage-sensing functions, the opening of maxiK channels requires 100-fold less Ca2+ at a membrane potential of +50 mV than at –50 mV. This unique feature of the maxiK gating mechanism allows the cell to appropriately suppress overly exuberant Ca2+ entry through voltage-gated Ca2+ channels. Thus, voltage-activated and Ca2+-dependent processes of smooth muscle contraction and neurotransmitter release at nerve terminals are both moderated by the sobering influence of maxiK. Gene knockout experiments and mapping of human genetic diseases have recently implicated maxiK in a variety of physiological control mechanisms and neurological disorders including bladder control, penile erection, hypertension, cerebellar ataxia and epilepsy (see Ledoux et al. 2006; Lu et al. 2006).
As the protein product of the HSlo gene, maxiK is subject to an extraordinary array of cellular regulatory mechanisms and signalling pathways. Too numerous to cite here, previously described mechanisms that modify the structure, gating and pharmacology of maxiK include: alternative mRNA splicing, oligomerization with a unique family of four accessory subunits (ß1–ß4), protein phosphorylation and dephosphorylation, control of cellular localization, and membrane clustering by formation of macromolecular complexes with a variety of receptor and scaffolding proteins. Some specific examples of the many signalling mechanisms and mediator pathways that are known to modulate maxiK function are: haem binding, diffusible gases (O2, CO, NO), second messengers (cAMP, cGMP) acting via respective protein kinases (PKA, PKG), Src tyrosine kinase, redox modulation via reactive oxygen species, fatty acids, arachidonic acid metabolites, sex steroids, and endogenous cannabinoids. In view of these many options for fine-tuning maxiK function to the exact requirements of tissue-specific regulation, one might be tempted to ask: ‘What more could a cell need?’
Korovkina et al. (2006) offer a rather startling riposte to this question in this issue of The Journal of Physiology with their description of a new mechanism for reactivating channel function from two silent parts of an endoproteolytically cleaved maxiK channel. The story began with the earlier identification of an alternative splice variant, mK44, of maxiK expressed in human myometrial cells, which contains an insertion of a unique 44-amino acid sequence within the first intracellular loop between S0 and S1 transmembrane spans (Korovkina et al. 2001). When cellular localization of protein corresponding to the mK44 splice variant was examined in pregnant myometrium or cultured human myometrial smooth muscle cells, mK44 was found in a predominantly non-functional state within the cytoplasm. Korovkina et al. (2006) hypothesized that human mK44 might be recruited to the plasma membrane under certain conditions. They confirmed this idea by demonstrating that treatment of myometrial cells with 20 mM caffeine in a Ca2+-free medium resulted in translocation of mK44 to the plasma membrane. Use of caffeine to trigger this process is based on the idea that maxiK channels in smooth muscle are activated by Ca2+ released from endoplasmic reticulum (ER) via caffeine-sensitive Ca2+-release channels.
Sequence analysis of the 44-residue insertion of mK44 further led Korovkina et al. (2006) to hypothesize that the insertion may be a site of endoproteolytic cleavage and N-myristoylation of the newly formed N-terminus at the cleavage site. Using a vector-expressed construct of mK44 containing an N-terminal c-myc epitope, a 10 kDa N-terminal fragment of mK44 was found to be constitutively located in the plasma membrane with the larger C-terminal portion restricted to the cytoplasmic membrane compartment. This same approach was used to show that treatment of myometrial cells with 20 mM caffeine actually results in the translocation of the large C-terminal domain of truncated mK44 to the plasma membrane.
The big question in this novel endoproteolysis/N-myristoylation–translocation scheme is whether myometrial cells actually use this mechanism to functionally activate quiescent intracellular maxiK channels. In support of this intriguing concept, the authors show that coexpression of separate 1–62 N-terminal and 69–1157 C-terminal fragments of mK44 (presumably similar to fragments generated by endoproteolytic cleavage) do indeed reconstitute functional maxiK current in HEK293 cells. Cellular physiologists who have worked to demystify the machinations of maxiK are certainly used to surprises. However, this astonishing revelation of proteolytic processing and molecular rendezvous is likely to provide maximal motivation for further studies of the role of proteases and membrane trafficking in the regulation of this important K+ channel.
References
Gutman GA, Chandy KG, Adelman JP, Aiyar J, Bayliss DA, Clapham DE et al. (2003). Pharmacol Rev 55, 583–586.
Korovkina VP, Brainard AM & England SK (2006). J Physiol, 573, 329–341.
Korovkina VP, Fergus DJ, Holdiman AJ & England SK (2001). Am J Physiol Cell Physiol 281, C361–C367.
Ledoux J, Werner ME, Brayden JE & Nelson MT (2006). Physiology (Bethesda) 21, 69–79.[CrossRef][Medline]
Lu R, Alioua A, Kumar Y, Eghbali M & Stefani E (2006). J Physiol 570, 65–72.
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