The long-term aim of this project is to understand the structural components of BK-type Ca2+- and voltage- activated K+ channels and their functional properties and physiological roles. BK channels are widely expressed and couple changes in submembrane Ca2+ concentrations to changes in membrane potential and excitability. How they do this varies in a tissue-specific fashion dependent in part on molecular composition. They consist minimally of four pore-forming 1 subunits. In addition, there are four distinct genes that encode auxiliary 2 subunits. Two of the 2 subunit family members, 22 and splice variants of 23, produce rapid inactivation in which the cytosolic N-terminus of the 2 subunits moves into a blocking position within the BK channel pore. However, in contrast to rapid inactivation of voltage-dependent K+ channels, inactivation of BK channels occurs by a two-step mechanism, in which a fully-conducting, pre-inactivated open state precedes the inactivated state. An implication of this mechanism is that it enables BK 2 subunits to regulate afterhyperpolarizations following development of inactivation. In fact, the 23a subunit results in a profound increase in the net current flux through BK channels following repolarization. This effect represents an entirely new mechanism by which slow use-dependent afterhyperpolarizations can be generated and will have profound effects on excitability of cells in which it is found. This project will be pursued in two parts. In part one, two aims are devoted to understanding the mechanism and structural basis of inactivation mediated by these 22 and 23 subunits. In part two, the potential physiological consequences of this inactivation mechanism and the tissue specific localization of the key 2 subunits will be determined. For the mechanistic studies and examination of physiological consequences, methods of electrophysiology combined with molecular biology will be employed. Methods of PCR and immunohistochemistry will be used to identify loci of expression of 22 and 23 subunits. Together these results will provide insight into a new type of regulatory mechanism that may have profound significance for regulation of excitability in cells containing BK 2 subunit. BK channels are of broad importance in the normal functioning of a variety of excitable cells. Among different tissues, BK channels contribute to regulation of neuronal excitability, smooth muscle relaxation, synaptic transmission and hormone release. Better understanding the composition and functional role of BK channel variants is of potential medical importance, not only because the channels may serve as specific therapeutic targets but also because altered function of particular variants may contribute to currently unrecognized pathological conditions. Calcium and voltage-regulated potassium channels (BK-type) are widely distributed among a range of cell types and contribute to regulation of neuronal excitability, smooth muscle relaxation, synaptic transmission and hormone release. These channels have been implicated in pathological conditions as diverse as hypertension and epilepsy. Better understanding the composition and functional role of BK channel variants is of potential medical importance, not only because the channels may serve as specific therapeutic targets but also because altered function of particular BK variants may contribute to currently unrecognized pathological conditions.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM081748-16
Application #
8026873
Study Section
Special Emphasis Panel (ZRG1-MDCN-N (03))
Program Officer
Hagan, Ann A
Project Start
1993-05-21
Project End
2012-09-27
Budget Start
2011-01-01
Budget End
2012-09-27
Support Year
16
Fiscal Year
2011
Total Cost
$388,325
Indirect Cost
Name
Washington University
Department
Anesthesiology
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Lingle, Christopher J (2015) NAVigating a transition from single action potential firing to bursting in chromaffin cells. J Physiol 593:761-2
Zeng, Xu-Hui; Yang, Chengtao; Xia, Xiao-Ming et al. (2015) SLO3 auxiliary subunit LRRC52 controls gating of sperm KSPER currents and is critical for normal fertility. Proc Natl Acad Sci U S A 112:2599-604
Martinez-Espinosa, Pedro L; Wu, Jianping; Yang, Chengtao et al. (2015) Knockout of Slo2.2 enhances itch, abolishes KNa current, and increases action potential firing frequency in DRG neurons. Elife 4:
Gonzalez-Perez, Vivian; Xia, Xiao-Ming; Lingle, Christopher J (2015) Two classes of regulatory subunits coassemble in the same BK channel and independently regulate gating. Nat Commun 6:8341
Brenker, Christoph; Zhou, Yu; Müller, Astrid et al. (2014) The Ca2+-activated K+ current of human sperm is mediated by Slo3. Elife 3:e01438
Gonzalez-Perez, Vivian; Xia, Xiao-Ming; Lingle, Christopher J (2014) Functional regulation of BK potassium channels by ?1 auxiliary subunits. Proc Natl Acad Sci U S A 111:4868-73
Martinez-Espinosa, Pedro L; Yang, Chengtao; Gonzalez-Perez, Vivian et al. (2014) Knockout of the BK ?2 subunit abolishes inactivation of BK currents in mouse adrenal chromaffin cells and results in slow-wave burst activity. J Gen Physiol 144:275-95
Zeng, Xu-Hui; Navarro, Betsy; Xia, Xiao-Ming et al. (2013) Simultaneous knockout of Slo3 and CatSper1 abolishes all alkalization- and voltage-activated current in mouse spermatozoa. J Gen Physiol 142:305-13
Ajith Karunarathne, W K; O'Neill, Patrick R; Martinez-Espinosa, Pedro L et al. (2012) All G protein ?? complexes are capable of translocation on receptor activation. Biochem Biophys Res Commun 421:605-11
Zhou, Yu; Zeng, Xu-Hui; Lingle, Christopher J (2012) Barium ions selectively activate BK channels via the Ca2+-bowl site. Proc Natl Acad Sci U S A 109:11413-8

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