Molecular mechanism underlying β1 regulation in voltage- and calcium-activated potassium (BK) channels

Being activated by depolarizing voltages and increases in cytoplasmic Ca2+, voltage- and calcium-activated potassium (BK) channels and their modulatory β-subunits are able to dampen or stop excitatory stimuli in a wide range of cellular types, including both neuronal and nonneuronal tissues. Minimal alterations in BK channel function may contribute to the pathophysiology of several diseases, including hypertension, asthma, cancer, epilepsy, and diabetes. Several gating processes, allosterically coupled to each other, control BK channel activity and are potential targets for regulation by auxiliary β-subunits that are expressed together with the α (BK)-subunit in almost every tissue type where they are found. By measuring gating currents in BK channels coexpressed with chimeras between β1 and β3 or β2 auxiliary subunits, we were able to identify that the cytoplasmic regions of β1 are responsible for the modulation of the voltage sensors. In addition, we narrowed down the structural determinants to the N terminus of β1, which contains two lysine residues (i.e., K3 and K4), which upon substitution virtually abolished the effects of β1 on charge movement. The mechanism by which K3 and K4 stabilize the voltage sensor is not electrostatic but specific, and the α (BK)-residues involved remain to be identified. This is the first report, to our knowledge, where the regulatory effects of the β1-subunit have been clearly assigned to a particular segment, with two pivotal amino acids being responsible for this modulation.

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Deletion of cytosolic gating ring decreases gate and voltage sensor coupling in BK channels

The Journal of General Physiology ◽

10.1085/jgp.201611646 ◽

2017 ◽

Vol 149 (3) ◽

pp. 373-387 ◽

Cited By ~ 13

Author(s):

Guohui Zhang ◽

Yanyan Geng ◽

Yakang Jin ◽

Jingyi Shi ◽

Kelli McFarland ◽

...

Keyword(s):

Bk Channels ◽

Bk Channel ◽

Voltage Sensor ◽

Voltage Sensitivity ◽

Gating Currents ◽

Voltage Sensors ◽

Transmembrane Voltage ◽

Wide Range ◽

Activation Gate ◽

Sensor Activation

Large conductance Ca2+-activated K+ channels (BK channels) gate open in response to both membrane voltage and intracellular Ca2+. The channel is formed by a central pore-gate domain (PGD), which spans the membrane, plus transmembrane voltage sensors and a cytoplasmic gating ring that acts as a Ca2+ sensor. How these voltage and Ca2+ sensors influence the common activation gate, and interact with each other, is unclear. A previous study showed that a BK channel core lacking the entire cytoplasmic gating ring (Core-MT) was devoid of Ca2+ activation but retained voltage sensitivity (Budelli et al. 2013. Proc. Natl. Acad. Sci. USA. http://dx.doi.org/10.1073/pnas.1313433110). In this study, we measure voltage sensor activation and pore opening in this Core-MT channel over a wide range of voltages. We record gating currents and find that voltage sensor activation in this truncated channel is similar to WT but that the coupling between voltage sensor activation and gating of the pore is reduced. These results suggest that the gating ring, in addition to being the Ca2+ sensor, enhances the effective coupling between voltage sensors and the PGD. We also find that removal of the gating ring alters modulation of the channels by the BK channel’s β1 and β2 subunits.

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BK channel inhibition by strong extracellular acidification

eLife ◽

10.7554/elife.38060 ◽

2018 ◽

Vol 7 ◽

Cited By ~ 3

Author(s):

Yu Zhou ◽

Xiao-Ming Xia ◽

Christopher J Lingle

Keyword(s):

Bk Channels ◽

Bk Channel ◽

Extracellular Acidification ◽

Channel Inhibition ◽

Wide Range ◽

The Family ◽

Voltage Dependent ◽

Intracellular Organelles ◽

Extracellular Side ◽

Key Residues

Mammalian BK-type voltage- and Ca2+-dependent K+ channels are found in a wide range of cells and intracellular organelles. Among different loci, the composition of the extracellular microenvironment, including pH, may differ substantially. For example, it has been reported that BK channels are expressed in lysosomes with their extracellular side facing the strongly acidified lysosomal lumen (pH ~4.5). Here we show that BK activation is strongly and reversibly inhibited by extracellular H+, with its conductance-voltage relationship shifted by more than +100 mV at pHO 4. Our results reveal that this inhibition is mainly caused by H+ inhibition of BK voltage-sensor (VSD) activation through three acidic residues on the extracellular side of BK VSD. Given that these key residues (D133, D147, D153) are highly conserved among members in the voltage-dependent cation channel superfamily, the mechanism underlying BK inhibition by extracellular acidification might also be applicable to other members in the family.

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Heme Regulates Allosteric Activation of the Slo1 BK Channel

The Journal of General Physiology ◽

10.1085/jgp.200509262 ◽

2005 ◽

Vol 126 (1) ◽

pp. 7-21 ◽

Cited By ~ 66

Author(s):

Frank T. Horrigan ◽

Stefan H. Heinemann ◽

Toshinori Hoshi

Keyword(s):

Divalent Cations ◽

Ionic Currents ◽

Bk Channels ◽

Bk Channel ◽

Voltage Sensor ◽

Gating Currents ◽

Calcium Dependent ◽

Channel Opening ◽

Activation Gate ◽

Voltage Dependent

Large conductance calcium-dependent (Slo1 BK) channels are allosterically activated by membrane depolarization and divalent cations, and possess a rich modulatory repertoire. Recently, intracellular heme has been identified as a potent regulator of Slo1 BK channels (Tang, X.D., R. Xu, M.F. Reynolds, M.L. Garcia, S.H. Heinemann, and T. Hoshi. 2003. Nature. 425:531–535). Here we investigated the mechanism of the regulatory action of heme on heterologously expressed Slo1 BK channels by separating the influences of voltage and divalent cations. In the absence of divalent cations, heme generally decreased ionic currents by shifting the channel's G–V curve toward more depolarized voltages and by rendering the curve less steep. In contrast, gating currents remained largely unaffected by heme. Simulations suggest that a decrease in the strength of allosteric coupling between the voltage sensor and the activation gate and a concomitant stabilization of the open state account for the essential features of the heme action in the absence of divalent ions. At saturating levels of divalent cations, heme remained similarly effective with its influence on the G–V simulated by weakening the coupling of both Ca2+ binding and voltage sensor activation to channel opening. The results thus show that heme dampens the influence of allosteric activators on the activation gate of the Slo1 BK channel. To account for these effects, we consider the possibility that heme binding alters the structure of the RCK gating ring and thereby disrupts both Ca2+- and voltage-dependent gating as well as intrinsic stability of the open state.

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The BK channel gating ring is strongly coupled to the voltage sensor

10.1101/381319 ◽

2018 ◽

Author(s):

Pablo Miranda ◽

Miguel Holmgren ◽

Teresa Giraldez

Keyword(s):

Conformational Changes ◽

Bk Channels ◽

Bk Channel ◽

Voltage Sensor ◽

Voltage Sensitivity ◽

Divalent Ions ◽

Gating Currents ◽

Open Probability ◽

Voltage Dependent ◽

Tightly Coupled

ABSTRACTThe open probability of large conductance voltage- and calcium-dependent potassium (BK) channels is regulated allosterically by changes in the transmembrane voltage and intracellular concentration of divalent ions (Ca2+ and Mg2+). The divalent cation sensors reside within the gating ring formed by eight Regulator of Conductance of Potassium (RCK) domains, two from each of the four channel subunits. Overall, the gating ring contains 12 sites that can bind Ca2+ with different affinities. Using patch-clamp fluorometry, we have shown robust changes in FRET signals within the gating ring in response to divalent ions and voltage, which do not directly track open probability. Only the conformational changes triggered through the RCK1 binding site are voltage-dependent in presence of Ca2+. Because the gating ring is outside the electric field, it must gain voltage sensitivity from coupling to the voltage-dependent channel opening, the voltage sensor or both. Here we demonstrate that alterations of voltage sensor dynamics known to shift gating currents produce a cognate shift in the gating ring voltage dependence, whereas changing BK channels’ relative probability of opening had little effect. These results strongly suggest that the conformational changes of the RCK1 domain of the gating ring are tightly coupled to the voltage sensor function, and this interaction is central to the allosteric modulation of BK channels.

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Calcium-driven regulation of voltage-sensing domains in BK channels

eLife ◽

10.7554/elife.44934 ◽

2019 ◽

Vol 8 ◽

Cited By ~ 3

Author(s):

Yenisleidy Lorenzo-Ceballos ◽

Willy Carrasquel-Ursulaez ◽

Karen Castillo ◽

Osvaldo Alvarez ◽

Ramon Latorre

Keyword(s):

Binding Sites ◽

Channel Gating ◽

Interaction Mechanism ◽

Bk Channels ◽

Bk Channel ◽

Strong Impact ◽

Gating Currents ◽

Voltage Sensing ◽

Α Subunit ◽

C Terminus

Allosteric interactions between the voltage-sensing domain (VSD), the Ca2+-binding sites, and the pore domain govern the mammalian Ca2+- and voltage-activated K+ (BK) channel opening. However, the functional relevance of the crosstalk between the Ca2+- and voltage-sensing mechanisms on BK channel gating is still debated. We examined the energetic interaction between Ca2+ binding and VSD activation by investigating the effects of internal Ca2+ on BK channel gating currents. Our results indicate that Ca2+ sensor occupancy has a strong impact on VSD activation through a coordinated interaction mechanism in which Ca2+ binding to a single α-subunit affects all VSDs equally. Moreover, the two distinct high-affinity Ca2+-binding sites contained in the C-terminus domains, RCK1 and RCK2, contribute equally to decrease the free energy necessary to activate the VSD. We conclude that voltage-dependent gating and pore opening in BK channels is modulated to a great extent by the interaction between Ca2+ sensors and VSDs.

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The Large-Conductance, Calcium-Activated Potassium Channel: A Big Key Regulator of Cell Physiology

Frontiers in Physiology ◽

10.3389/fphys.2021.750615 ◽

2021 ◽

Vol 12 ◽

Author(s):

Maria Sancho ◽

Barry D. Kyle

Keyword(s):

Bk Channels ◽

Bk Channel ◽

Channel Structure ◽

Cell Physiology ◽

Ongoing Research ◽

Post Translational Modifications ◽

Wide Range ◽

And Function ◽

Calcium Activated Potassium Channel ◽

Silver Bullet

Large-conductance Ca2+-activated K+ channels facilitate the efflux of K+ ions from a variety of cells and tissues following channel activation. It is now recognized that BK channels undergo a wide range of pre- and post-translational modifications that can dramatically alter their properties and function. This has downstream consequences in affecting cell and tissue excitability, and therefore, function. While finding the “silver bullet” in terms of clinical therapy has remained elusive, ongoing research is providing an impressive range of viable candidate proteins and mechanisms that associate with and modulate BK channel activity, respectively. Here, we provide the hallmarks of BK channel structure and function generally, and discuss important milestones in the efforts to further elucidate the diverse properties of BK channels in its many forms.

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Coupling of Ca2+ and voltage activation in BK channels through the αB helix/voltage sensor interface

10.1101/2020.01.29.925545 ◽

2020 ◽

Author(s):

Yanyan Geng ◽

Zengqin Deng ◽

Guohui Zhang ◽

Gonzalo Budelli ◽

Alice Butler ◽

...

Keyword(s):

Modular Design ◽

Cell Types ◽

Cytoplasmic Domain ◽

Bk Channels ◽

Bk Channel ◽

Voltage Sensor ◽

Membrane Excitability ◽

Gating Currents ◽

Voltage Sensors ◽

Voltage Activation

AbstractLarge conductance Ca2+ and voltage activated K+ (BK) channels control membrane excitability in many cell types. BK channels are tetrameric. Each subunit is comprised of a voltage sensor domain (VSD), a central pore gate domain, and a large cytoplasmic domain (CTD) that contains the Ca2+ sensors. While it is known that BK channels are activated by voltage and Ca2+, and that voltage and Ca2+ activations interact, less is known about the mechanisms involved. We now explore mechanism by examining the gating contribution of an interface formed between the VSDs and the αB helices located at the top of the CTDs. Proline mutations in the αB helix greatly decreased voltage activation while having negligible effects on gating currents. Analysis with the HCA model indicated a decreased coupling between voltage sensors and pore gate. Proline mutations decreased Ca2+ activation for both Ca2+ bowl and RCK1 Ca2+ sites, suggesting that both high affinity Ca2+ sites transduce their effect, at least in part, through the αB helix. Mg2+ activation was also decreased. The crystal structure of the CTD with proline mutation L390P showed a flattening of the first helical turn in the αB helix compared to WT, without other notable differences in the CTD, indicating structural change from the mutation was confined to the αB helix. These findings indicate that an intact αB helix/VSD interface is required for effective coupling of Ca2+ binding and voltage depolarization to pore opening, and that shared Ca2+ and voltage transduction pathways involving the αB helix may be involved.SignificanceLarge conductance BK (Slo1) K+ channels are activated by voltage, Ca2+, and Mg2+ to modulate membrane excitability in neurons, muscle, and other cells. BK channels are of modular design, with pore-gate and voltage sensors as transmembrane domains and a large cytoplasmic domain CTD containing the Ca2+ sensors. Previous observations suggest that voltage and Ca2+ sensors interact, but less is known about this interaction and its involvement in the gating process. We show that a previously identified structural interface between the CTD and voltage sensors is required for effective activation by both voltage and Ca2+, suggesting that these processes may share common allosteric activation pathways. Such knowledge should help explain disease processes associated with BK channel dysfunction.

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BK channel inhibition by strong extracellular acidification

10.1101/317339 ◽

2018 ◽

Author(s):

Yu Zhou ◽

Xiao-Ming Xia ◽

Christopher J. Lingle

Keyword(s):

Bk Channels ◽

Bk Channel ◽

Extracellular Acidification ◽

Channel Inhibition ◽

Wide Range ◽

The Family ◽

Voltage Dependent ◽

Intracellular Organelles ◽

Extracellular Side ◽

Key Residues

AbstractBK-type voltage- and Ca2+-dependent K+ channels are found in a wide range of cells and intracellular organelles. Among different loci, the composition of the extracellular microenvironment, including pH, may differ substantially. For example, it has been reported that BK channels are expressed in lysosomes with their extracellular side facing the strongly acidified lysosomal lumen (pH ~ 4.5). Here we show that BK activation is strongly and reversibly inhibited by extracellular H+, with its conductance-voltage relationship shifted by more than +100 mV at pHO 4. Our results reveal that this inhibition is mainly caused by H+ inhibition of BK voltage-sensor (VSD) activation through three acidic residues on the extracellular side of BK VSD. Given that these key residues (D133, D147, D153) are highly conserved among members in the voltage-dependent cation channel superfamily, the mechanism underlying BK inhibition by extracellular acidification might also be applicable to other members in the family.

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Calcium-driven regulation of voltage-sensing domains in BK channels

10.1101/520429 ◽

2019 ◽

Author(s):

Yenisleidy Lorenzo-Ceballos ◽

Willy Carrasquel-Ursulaez ◽

Karen Castillo ◽

Osvaldo Alvarez ◽

Ramon Latorre

Keyword(s):

Binding Sites ◽

Interaction Mechanism ◽

Bk Channels ◽

Bk Channel ◽

Strong Impact ◽

Gating Currents ◽

Voltage Sensing ◽

Α Subunit ◽

C Terminus ◽

Voltage Dependent

AbstractAllosteric interplays between voltage-sensing domains (VSD), Ca2+-binding sites, and the pore domain govern the Ca2+- and voltage-activated K+ (BK) channel opening. However, the functional relevance of the Ca2+- and voltage-sensing mechanisms crosstalk on BK channel gating is still debated. We examined the energetic interaction between Ca2+ binding and VSD activation measuring and analyzing the effects of internal Ca2+ on BK channels gating currents. Our results indicate that the Ca2+ sensors occupancy has a strong impact on the VSD activation through a coordinated interaction mechanism in which Ca2+ binding to a single α-subunit affects all VSDs equally. Moreover, the two distinct high-affinity Ca2+-binding sites contained in the C-terminus domains, RCK1 and RCK2, appear to contribute equally to decrease the free energy necessary to activate the VSD. We conclude that voltage-dependent gating and pore opening in BK channels is modulated to a great extent by the interaction between Ca2+ sensors and VSDs.

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The Yin and Yang of BK Channels in Epilepsy

CNS & Neurological Disorders - Drug Targets ◽

10.2174/1871527317666180213142403 ◽

2018 ◽

Vol 17 (4) ◽

pp. 272-279 ◽

Cited By ~ 12

Author(s):

Yudan Zhu ◽

Shuzhang Zhang ◽

Yijun Feng ◽

Qian Xiao ◽

Jiwei Cheng ◽

...

Keyword(s):

Central Nervous System ◽

Nervous System ◽

Bk Channels ◽

Bk Channel ◽

Potential Shape ◽

Yin And Yang ◽

The Central Nervous System ◽

Action Potential Shape ◽

Excitability Of Neurons

Background & Objective: The large conductance calcium-activated potassium (BK) channel, extensively distributed in the central nervous system (CNS), is considered as a vital player in the pathogenesis of epilepsy, with evidence implicating derangement of K+ as well as regulating action potential shape and duration. However, unlike other channels implicated in epilepsy whose function in neurons could clearly be labeled “excitatory” or “inhibitory”, the unique physiological behavior of the BK channel allows it to both augment and decrease the excitability of neurons. Thus, the role of BK in epilepsy is controversial so far, and a growing area of intense investigation. Conclusion: Here, this review aims to highlight recent discoveries on the dichotomous role of BK channels in epilepsy, focusing on relevant BK-dependent pro- as well as antiepileptic pathways, and discuss the potential of BK specific modulators for the treatment of epilepsy.

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