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In order to quantitatively study the interfered output of the accelerometer under an acoustic injection attack, a mathematical model for fitting and predicting the accelerometer output was proposed. With ADXL103 as an example, an acoustic injection attack experiment with amplitude sweeping and frequency sweeping was performed. In the mathematical model, the R-squared coefficient was R2 = 0.9990 in the acoustic injection attack experiment with amplitude sweeping, and R2 = 0.9888 with frequency sweeping. Based on the mathematical model, the dual frequency acoustic injection attack mode was proposed. The difference frequency signal caused by the nonlinear effect was not filtered by the low-pass filter. At a 115 dB sound pressure level, the maximum acceleration bias of the output was 4.4 m/s2 and the maximum amplitude of fluctuation was 4.97 m/s2. Two kinds of methods of prevention against acoustic injection attack were proposed, including changing the damping ratio of the accelerometer and adding a preposition low-pass filter.

Loss-of-function mutations in K

Calmodulin controls KCNQ1 potassium channel pore opening by competing with PIP2 in the KCNQ1 voltage-sensing domain., Calmodulin (CaM) and phosphatidylinositol 4,5-bisphosphate (PIP2) are potent regulators of the voltage-gated potassium channel KCNQ1 (KV7.1), which conducts the cardiac IKs current. Although cryo–electron microscopy structures revealed intricate interactions between the KCNQ1 voltage-sensing domain (VSD), CaM, and PIP2, the functional consequences of these interactions remain unknown. Here, we show that CaM-VSD interactions act as a state-dependent switch to control KCNQ1 pore opening. Combined electrophysiology and molecular dynamics network analysis suggest that VSD transition into the fully activated state allows PIP2 to compete with CaM for binding to VSD. This leads to conformational changes that alter VSD-pore coupling to stabilize open states. We identify a motif in the KCNQ1 cytosolic domain, which works downstream of CaM-VSD interactions to facilitate the conformational change. Our findings suggest a gating mechanism that integrates PIP2 and CaM in KCNQ1 voltage-dependent activation, yielding insights into how KCNQ1 gains the phenotypes critical for its physiological function.

Large-conductance potassium (BK) channels are transmembrane (TM) proteins that can be synergistically and independently activated by membrane voltage and intracellular Ca2+. The only covalent connection between the cytosolic Ca2+ sensing domain and the TM pore and voltage sensing domains is a 15-residue ‘C-linker’. To determine the linker’s role in human BK activation, we designed a series of linker sequence scrambling mutants to suppress potential complex interplay of specific interactions with the rest of the protein. The results revealed a surprising sensitivity of BK activation to the linker sequence. Combining atomistic simulations and further mutagenesis experiments, we demonstrated that nonspecific interactions of the linker with membrane alone could directly modulate BK activation. The C-linker thus plays more direct roles in mediating allosteric coupling between BK domains than previously assumed. Our results suggest that covalent linkers could directly modulate TM protein function and should be considered an integral component of the sensing apparatus.

Large-conductance Ca(2+) and voltage-activated K(+) (BK) channels control membrane excitability in many cell types. BK channels are tetrameric. Each subunit is composed of a voltage sensor domain (VSD), a central pore-gate domain, and a large cytoplasmic domain (CTD) that contains the Ca(2+) sensors. While it is known that BK channels are activated by voltage and Ca(2+), and that voltage and Ca(2+) activations interact, less is known about the mechanisms involved. We explore here these mechanisms 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 Horrigan, Cui, and Aldrich model indicated a decreased coupling between voltage sensors and pore gate. Proline mutations decreased Ca(2+) activation for both Ca(2+) bowl and RCK1 Ca(2+) sites, suggesting that both high-affinity Ca(2+) sites transduce their effect, at least in part, through the αB helix. Mg(2+) activation 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 wild type, without other notable differences in the CTD, indicating that structural changes from the mutation were confined to the αB helix. These findings indicate that an intact αB helix/VSD interface is required for effective coupling of Ca(2+) binding and voltage depolarization to pore opening and that shared Ca(2+) and voltage transduction pathways involving the αB helix may be involved.

Large-conductance Ca2+-activated K+ channels (BK channels) are key drug targets due to their association with a wide variety of neurological disorders. Rockman et al. reveal that the smooth muscle relaxant NS11021 activates BK channels by shifting the pore-gate equilibrium toward the open state., Large-conductance Ca2+-activated K+ channels (BK channels) are activated by cytosolic calcium and depolarized membrane potential under physiological conditions. Thus, these channels control electrical excitability in neurons and smooth muscle by gating K+ efflux and hyperpolarizing the membrane in response to Ca2+ signaling. Altered BK channel function has been linked to epilepsy, dyskinesia, and other neurological deficits in humans, making these channels a key target for drug therapies. To gain insight into mechanisms underlying pharmacological modulation of BK channel gating, here we studied mechanisms underlying activation of BK channels by the biarylthiourea derivative, NS11021, which acts as a smooth muscle relaxant. We observe that increasing NS11021 shifts the half-maximal activation voltage for BK channels toward more hyperpolarized voltages, in both the presence and nominal absence of Ca2+, suggesting that NS11021 facilitates BK channel activation primarily by a mechanism that is distinct from Ca2+ activation. 30 µM NS11021 slows the time course of BK channel deactivation at −200 mV by ∼10-fold compared with 0 µM NS11021, while having little effect on the time course of activation. This action is most pronounced at negative voltages, at which the BK channel voltage sensors are at rest. Single-channel kinetic analysis further shows that 30 µM NS11021 increases open probability by 62-fold and increases mean open time from 0.15 to 0.52 ms in the nominal absence of Ca2+ at voltages less than −60 mV, conditions in which BK voltage sensors are largely in the resting state. We could therefore account for the major activating effects of NS11021 by a scheme in which the drug primarily shifts the pore-gate equilibrium toward the open state.

Background The mutations of KCNMA1 BK-type K+ channel have been identified in patients with various movement disorders. The underlying pathophysiology and corresponding therapeutics are lacking. Objectives To report our clinical and biophysical characterizations of a novel de novo KCNMA1 variant, as well as an effective therapy for the patient's dystonia-atonia spells. Methods Combination of phenotypic characterization, therapy, and biophysical characterization of the patient and her mutation. Results The patient had >100 dystonia-atonia spells per day with mild cerebellar atrophy. She also had autism spectrum disorder, intellectual disability, and attention deficit hyperactivity disorder. Whole-exome sequencing identified a heterozygous de novo BK N536H mutation. Our biophysical characterization demonstrates that N536H is a gain-of-function mutation with markedly enhanced voltage-dependent activation. Remarkably, administration of dextroamphetamine completely suppressed the dystonia-atonia spells. Conclusions BK N536H is a gain-of-function that causes dystonia and other neurological symptoms. Our stimulant therapy opens a new avenue to mitigate KCNMA1-linked movement disorders. © 2020 International Parkinson and Movement Disorder Society.

Large-conductance potassium (BK) channels are transmembrane (TM) proteins that can be synergistically and independently activated by membrane voltage and intracellular Ca2+. The only covalent connection between the cytosolic Ca2+ sensing domain and the TM pore and voltage sensing domains is a 15-residue "C-linker". To determine the linker's role in BK activation, we designed a series of linker sequence scrambling mutants to suppress potential complex interplay of specific interactions with the rest of the protein. The results revealed a surprising sensitivity of BK activation to the linker sequence. Combing atomistic simulations and further mutagenesis experiments, we demonstrated that nonspecific interactions of the linker with membrane alone could directly modulate BK activation. The C-linker thus plays more direct roles in mediating allosteric coupling between BK domains than previously assumed. Our results also suggest that covalent linkers could directly modulate TM protein function and should be considered an integral component of the sensing apparatus.

Voltage-gated ion channels feature voltage sensor domains (VSDs) that exist in three distinct conformations during activation: resting, intermediate, and activated. Experimental determination of the structure of a potassium channel VSD in the intermediate state has previously proven elusive. Here, we report and validate the experimental three-dimensional structure of the human KCNQ1 voltage-gated potassium channel VSD in the intermediate state. We also used mutagenesis and electrophysiology in Xenopus laevisoocytes to functionally map the determinants of S4 helix motion during voltage-dependent transition from the intermediate to the activated state. Finally, the physiological relevance of the intermediate state KCNQ1 conductance is demonstrated using voltage-clamp fluorometry. This work illuminates the structure of the VSD intermediate state and demonstrates that intermediate state conductivity contributes to the unusual versatility of KCNQ1, which can function either as the slow delayed rectifier current (IKs) of the cardiac action potential or as a constitutively active epithelial leak current.

Large 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.

Sonogenetics, which uses ultrasound to noninvasively control cells genetically modified with ultrasound-sensitive ion channels, can be a powerful tool for investigating intact brain circuits. Here we show that TRPV1 is an ultrasound-sensitive ion channel that can modify the activity of TRPV1-expressing cells in vitro when exposing to focused ultrasound. We also show that focused ultrasound exposure at the mouse brain in vivo can selectively activate neurons that are genetically modified to express TRPV1. We demonstrate that precise manipulation of neural activity via TRPV1-based sonogenetics can be achieved by spatiotemporal control of ultrasound-induced heating. The focused ultrasound exposure is safe based on our inspection of neuronal integrity, apoptosis, and inflammation markers. This sonogenetic tool enables noninvasive, cell-type specific, spatiotemporally controlled modulation of mammalian brain activity.

Toxoplasma gondii is an obligate protozoan parasite infecting diverse hosts. Studies have demonstrated that different hosts respond differently to Toxoplasma infection. Pigs are among the most susceptible hosts of T. gondii, but the host-pathogen interactions that shape the outcome of infection in pigs are completely unknown. Here, we used dual RNA-seq to profile the transcriptomic changes of porcine alveolar macrophages (PAMs) upon Toxoplasma infection. Our results indicated that PAMs initiated different responses to Toxoplasma infection compared with mouse macrophages. First, although infected PAMs upregulated numerous pro-inflammatory factors, IL-12, which plays critical roles in IL-12~IFN-γ-mediated immunity against Toxoplasma infection in mice, was found unchanged during PAM infection. Second, the gene encoding iNOS that is responsible for nitric oxide (NO) production was also not induced in infected PAMs. Consistently, there was no NO level change in PAMs after infection. Third, it seems like Toxoplasma infection inhibited apoptosis in PAMs. On the parasite side, the most obvious change is the upregulation of genes involved in metabolism and macromolecule synthesis, such as the type II fatty acid synthesis in the apicoplast. Together, these results revealed distinct responses of PAMs to Toxoplasma infection and provide novel insights into Toxoplasma-pig interactions.

KCNQ family K+ channels (KCNQ1-5) in the heart, nerve, epithelium and ear require phosphatidylinositol 4,5-bisphosphate (PIP2) for voltage dependent activation. While membrane lipids are known to regulate voltage sensor domain (VSD) activation and pore opening in voltage dependent gating, PIP2 was found to interact with KCNQ1 and mediate VSD-pore coupling. Here, we show that a compound CP1, identified in silico based on the structures of both KCNQ1 and PIP2, can substitute for PIP2 to mediate VSD-pore coupling. Both PIP2 and CP1 interact with residues amongst a cluster of amino acids critical for VSD-pore coupling. CP1 alters KCNQ channel function due to different interactions with KCNQ compared with PIP2. We also found that CP1 returned drug-induced action potential prolongation in ventricular myocytes to normal durations. These results reveal the structural basis of PIP2 regulation of KCNQ channels and indicate a potential approach for the development of anti-arrhythmic therapy., Yongfeng Liu, Xianjin Xu, Junyuan Gao et al. perform in silico screening targeting compounds that resemble both KCNQ1 and PIP2 structures and identify CP1 as a compound that could restore KCNQ1 currents after PIP2 depletion. Like PIP2, this compound can mediate VSD-pore coupling of the channel, albeit via a different mechanism from PIP2.

Significance The work showed that centipede venom can cause disorders in cardiovascular, respiratory, and nervous systems. The cardiovascular toxicity of the venom comes mostly from a peptide toxin SsTx, which blocks the KCNQ family of potassium channels. Retigabine, a KCNQ channel opener, neutralizes centipede venom toxicity, and thus could be used to treat centipede envenomation., Centipedes can subdue giant prey by using venom, which is metabolically expensive to synthesize and thus used frugally through efficiently disrupting essential physiological systems. Here, we show that a centipede (Scolopendra subspinipes mutilans, ∼3 g) can subdue a mouse (∼45 g) within 30 seconds. We found that this observation is largely due to a peptide toxin in the venom, SsTx, and further established that SsTx blocks KCNQ potassium channels to exert the lethal toxicity. We also demonstrated that a KCNQ opener, retigabine, neutralizes the toxicity of a centipede’s venom. The study indicates that centipedes’ venom has evolved to simultaneously disrupt cardiovascular, respiratory, muscular, and nervous systems by targeting the broadly distributed KCNQ channels, thus providing a therapeutic strategy for centipede envenomation.

Zhou et al. consider the biophysics of large-conductance Ca2+-activated Slo1 channels in the context of Aplysia Slo1 structures., For those interested in the machinery of ion channel gating, the Ca2+ and voltage-activated BK K+ channel provides a compelling topic for investigation, by virtue of its dual allosteric regulation by both voltage and intracellular Ca2+ and because its large-single channel conductance facilitates detailed kinetic analysis. Over the years, biophysical analyses have illuminated details of the allosteric regulation of BK channels and revealed insights into the mechanism of BK gating, e.g., inner cavity size and accessibility and voltage sensor-pore coupling. Now the publication of two structures of an Aplysia californica BK channel—one liganded and one metal free—promises to reinvigorate functional studies and interpretation of biophysical results. The new structures confirm some of the previous functional inferences but also suggest new perspectives regarding cooperativity between Ca2+-binding sites and the relationship between voltage- and Ca2+-dependent gating. Here we consider the extent to which the two structures explain previous functional data on pore-domain properties, voltage-sensor motions, and divalent cation binding and activation of the channel.

Gain-of-function mutations in the pore-forming subunit of IKs channels, KCNQ1, lead to short QT syndrome (SQTS) and lethal arrhythmias. However, how mutant IKs channels cause SQTS and the possibility of IKs-specific pharmacological treatment remain unclear. V141M KCNQ1 is a SQTS associated mutation. We studied its effect on IKs gating properties and changes in the action potentials (AP) of human ventricular myocytes. Xenopus oocytes were used to study the gating mechanisms of expressed V141M KCNQ1/KCNE1 channels. Computational models were used to simulate human APs in endocardial, mid-myocardial, and epicardial ventricular myocytes with and without β-adrenergic stimulation. V141M KCNQ1 caused a gain-of-function in IKs characterized by increased current density, faster activation, and slower deactivation leading to IKs accumulation. V141M KCNQ1 also caused a leftward shift of the conductance-voltage curve compared to wild type (WT) IKs (V1/2 = 33.6 ± 4.0 mV for WT, and 24.0 ± 1.3 mV for heterozygous V141M). A Markov model of heterozygous V141M mutant IKs was developed and incorporated into the O’Hara–Rudy model. Compared to the WT, AP simulations demonstrated marked rate-dependent shortening of AP duration (APD) for V141M, predicting a SQTS phenotype. Transmural electrical heterogeneity was enhanced in heterozygous V141M AP simulations, especially under β-adrenergic stimulation. Computational simulations identified specific IK1 blockade as a beneficial pharmacologic target for reducing the transmural APD heterogeneity associated with V141M KCNQ1 mutation. V141M KCNQ1 mutation shortens ventricular APs and enhances transmural APD heterogeneity under β-adrenergic stimulation. Computational simulations identified IK1 blockers as a potential antiarrhythmic drug of choice for SQTS.

Bingpian is a time‐honored herb in traditional Chinese medicine (TCM). It is an almost pure chemical with a chemical composition of (+)‐borneol and has been historically used as a topical analgesic for millennia. However, the clinical efficacy of topical borneol lacks stringent evidence‐based clinical studies and verifiable scientific mechanism. We examined the analgesic efficacy of topical borneol in a randomized, double‐blind, placebo‐controlled clinical study involving 122 patients with postoperative pain. Topical application of borneol led to significantly greater pain relief than placebo did. Using mouse models of pain, we identified the TRPM8 channel as a molecular target of borneol and showed that topical borneol‐induced analgesia was almost exclusively mediated by TRPM8, and involved a downstream glutamatergic mechanism in the spinal cord. Investigation of the actions of topical borneol and menthol revealed mechanistic differences between borneol‐ and menthol‐induced analgesia and indicated that borneol exhibits advantages over menthol as a topical analgesic. Our work demonstrates that borneol, which is currently approved by the US FDA to be used only as a flavoring substance or adjuvant in food, is an effective topical pain reliever in humans and reveals a key part of the molecular mechanism underlying its analgesic effect.

The thiazolidine ring is a biologically active chemical structure and is associated with many pharmacological activities. However, the biological molecules that can interact with the thiazolidine ring are not known. We show that thiazolidine causes sustained activation of the TRPA1 channel and chemically reacts with glutathione, and the chemical reactivity of thiazolidine ring is required for TRPA1 activation. Reducing agents reverse thiazolidine-induced TRPA1 activation, and mutagenesis studies show that nucleophilic cysteine residues in TRPA1 are critical, suggesting an activation mechanism involving thioreactive chemical reactions. In vivo studies show that thiazolidine induces acute pain and inflammation in mouse and these responses are specifically dependent on TRPA1. These results indicate that thiazolidine compounds can chemically react with biological molecules containing nucleophilic cysteines, thereby exerting biological activities.

Both cellular depolarization and intracellular Ca2+ can gate open large conductance Ca2+-activated K+ channels. Zhang et al. show that the intracellular gating ring, which forms the Ca2+-sensing machinery of the channel, is also required for activated voltage sensors to effectively gate open the pore., 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.

In voltage-gated potassium (KV) channels, the voltage-sensing domain (VSD) undergoes sequential activation from the resting state to the intermediate state and activated state to trigger pore opening via electro–mechanical (E–M) coupling. However, the spatial and temporal details underlying E–M coupling remain elusive. Here, utilizing KV7.1’s unique two open states, we report a two-stage E–M coupling mechanism in voltage-dependent gating of KV7.1 as triggered by VSD activations to the intermediate and then activated state. When the S4 segment transitions to the intermediate state, the hand-like C-terminus of the VSD-pore linker (S4-S5L) interacts with the pore in the same subunit. When S4 then proceeds to the fully-activated state, the elbow-like hinge between S4 and S4-S5L engages with the pore of the neighboring subunit to activate conductance. This two-stage hand-and-elbow gating mechanism elucidates distinct tissue-specific modulations, pharmacology, and disease pathogenesis of KV7.1, and likely applies to numerous domain-swapped KV channels., In voltage-gated potassium (KV) channels, the voltage-sensing domain (VSD) undergoes activation states to trigger pore opening via electro–mechanical (E–M) coupling. Here authors show that KV7.1 undergoes a two-stage E–M coupling mechanism during voltage-dependent activation.

Upon membrane depolarization, the KCNQ1 potassium channel opens at the intermediate (IO) and activated (AO) states of the stepwise voltage-sensing domain (VSD) activation. In the heart, KCNQ1 associates with KCNE1 subunits to form IKs channels that regulate heart rhythm. KCNE1 suppresses the IO state so that the IKs channel opens only to the AO state. Here, we tested modulations of human KCNQ1 channels by an activator ML277 in Xenopus oocytes. It exclusively changes the pore opening properties of the AO state without altering the IO state, but does not affect VSD activation. These observations support a distinctive mechanism responsible for the VSD-pore coupling at the AO state that is sensitive to ML277 modulation. ML277 provides insights and a tool to investigate the gating mechanism of KCNQ1 channels, and our study reveals a new strategy for treating long QT syndrome by specifically enhancing the AO state of native IKs currents.

In response to membrane depolarization, the KCNQ1 potassium channel opens at the intermediate (IO) and activated (AO) states that correspond to the stepwise activation of the voltage sensing domain (VSD) to the intermediate (I) and activated (A) states. In the heart, KCNQ1 associates with the auxiliary subunit KCNE1 to form the IKs channel that regulates heart rhythm. More than 300 of loss-of-function KCNQ1 mutations cause long QT syndrome (LQTS). KCNE1 suppresses the IO state so that the IKs channel opens only to the AO state. Thus, enhancing AO state presents a potential therapy for anti-LQTS. Here, we systematically tested modulations of KCNQ1 channels by a KCNQ1 activator, ML277. It enhances the current amplitude, slows down activation, deactivation and inactivation kinetics, shifts the voltage dependence of activation to more positive voltages, decreases the Rb+/K+ permeability ratio, and selectively increases currents of mutant KCNQ1 channels that open only to the AO state. All these observations are consistent with the mechanism that ML277 specifically potentiates the AO state. On the other hand, ML277 does not affect the VSD activation, suggesting that it potentiates the AO state by enhancing the electromechanical (E-M) coupling when the VSD moves to the activated state. Our results suggest that ML277 provides a unique tool to investigate the gating mechanism of KCNQ1 and IKs channels. The specificity of ML277 to increase the AO state of native IKs currents also suggests a new strategy for anti-LQTS therapy.

Animal toxins that are used to subdue prey and deter predators act as the key drivers in natural food chains and ecosystems. However, the predators of venomous animals may exploit feeding adaptation strategies to overcome toxins their prey produce. Much remains unknown about the genetic and molecular game process in the toxin-dominant food chain model. Here, we show an evolutionary strategy in different trophic levels of scorpion-eating amphibians, scorpions and insects, representing each predation relationship in habitats dominated by the paralytic toxins of scorpions. For scorpions preying on insects, we found that the scorpion α-toxins irreversibly activate the skeletal muscle sodium channel of their prey (insect, BgNaV1) through a membrane delivery mechanism and an efficient binding with the Asp/Lys-Tyr motif of BgNaV1. However, in the predatory game between frogs and scorpions, with a single point mutation (Lys to Glu) in this motif of the frog's skeletal muscle sodium channel (fNaV1.4), fNaV1.4 breaks this interaction and diminishes muscular toxicity to the frog; thus, frogs can regularly prey on scorpions without showing paralysis. Interestingly, this molecular strategy also has been employed by some other scorpion-eating amphibians, especially anurans. In contrast to these amphibians, the Asp/Lys-Tyr motifs are structurally and functionally conserved in other animals that do not prey on scorpions. Together, our findings elucidate the protein-protein interacting mechanism of a toxin-dominant predator-prey system, implying the evolutionary game theory at a molecular level.

Iodine antiseptics exhibit superior antimicrobial efficacy and do not cause acquired microbial resistance. However, they are underused in comparison with antibiotics in infection treatments, partly because of their adverse effects such as pain and allergy. The cause of these noxious effects is not fully understood, and no specific molecular targets or mechanisms have been discovered. In this study, we show that iodine antiseptics cause pain and promote allergic contact dermatitis in mouse models, and iodine stimulates a subset of sensory neurons that express TRPA1 and TRPV1 channels. In vivo pharmacological inhibition or genetic ablation of these channels indicates that TRPA1 plays a major role in iodine antiseptics‐induced pain and the adjuvant effect of iodine antiseptics on allergic contact dermatitis and that TRPV1 is also involved. We further demonstrate that iodine activates TRPA1 through a redox mechanism but has no direct effects on TRPV1. Our study improves the understanding of the adverse effects of iodine antiseptics and suggests a means to minimize their side effects through local inhibition of TRPA1 and TRPV1 channels.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

To overcome the adverse effects of conventional chemotherapy for cancers, various nanoparticles based drug delivery systems have been developed. However, nanoparticles delivering drugs directly to kill tumor cells still faced with challenges, because tumors possessed adopt complex mechanism to resist damages, which compromised the therapeutic efficacy. TMEM16A/CaCCs (Calcium activates chloride channels) has been identified to be overexpressed in lung adenocarcinoma which can serve as a novel tumor specific drug target in our previous work. Here, we developed a novel dual-targeted antitumor strategy via designing a novel nano-assembled, pH-sensitive drug-delivery system loading with specific inhibitors of TMEM16A against lung adenocarcinoma. For validation, we assayed the novel dual-targeting therapy on xenograft mouse model which exhibited significant antitumor activity and not affect mouse body weight. The dual targeting therapy accomplished in this study will shed light on the development of advanced antitumor strategy.

The gating mechanism of transmembrane ion channels is crucial for understanding how these proteins control ion flow across membranes in various physiological processes. Big potassium (BK) channels are particularly interesting with large single-channel conductance and dual regulation by membrane voltage and intracellular Ca2+. Recent atomistic structures of BK channels failed to identify structural features that could physically block the ion flow in the closed state. Here, we show that gating of BK channels does not seem to require a physical gate. Instead, changes in the pore shape and surface hydrophobicity in the Ca2+-free state allow the channel to readily undergo hydrophobic dewetting transitions, giving rise to a large free energy barrier for K+ permeation. Importantly, the dry pore remains physically open and is readily accessible to quaternary ammonium channel blockers. The hydrophobic gating mechanism is also consistent with scanning mutagenesis studies showing that modulation of pore hydrophobicity is correlated with activation properties., BK channels are regulated by membrane voltage and intracellular Ca2+ but the structural features that block the ion flow in the closed state remain unknown. Here authors use molecular dynamics simulation and show that a physical gate is not required; instead ion flow is regulated by hydrophobic dewetting due to changes in pore shape and surface hydrophobicity.

Trichinellosis caused by parasitic nematodes of the genus Trichinella may result in human morbidity and mortality worldwide. Deciphering processes that drive species diversity and adaptation are key to understanding parasitism and developing effective control strategies. Our goal was to identify genes that are under positive selection and possible mechanisms of adaptive evolution of Trichinella spiralis genes using a comparative genomic analysis with the genomes of Brugia malayi, Trichuris suis, Ancylostoma ceylanicum, and Caenorhabditis elegans. The CODEML program derived from the PAML package was used to deduce the most probable dN/dS ratio, a measurement to detect genes/proteins undergoing adaptation. For each pair of sequences, those with a dN/dS ratio > 1 were considered positively selected genes (PSGs). Altogether, 986 genes were positively selected (p-value < 0.01). Genes involved in metabolic pathways, signaling pathways, and cytosolic DNA-sensing pathways were significantly enriched among the PSGs. Several PSGs are associated with exploitation of the host: modification of the host's metabolism, creation of new parasite-specific morphological structures between T. spiralis and the host interface, xenobiotic metabolism to combat low oxygen concentrations and host toxicity, muscle cell transformation, cell cycle arrest, DNA repair processes during nurse cell formation, antiapoptotic factors, immunomodulation, and regulation of epigenetic processes. Some of the T. spiralis PSGs have C. elegans orthologs that confer severe or lethal RNAi phenotypes. Fifty-seven PSGs in T. spiralis were analyzed to encode differentially expressed proteins. The present study utilized an overall comparative genomic analysis to discover PSGs within T. spiralis and their relationships with biological function and organism fitness. This analysis adds to our understanding of the possible mechanism that contributes to T. spiralis parasitism and biological adaptation within the host, and thus these identified genes may be potential targets for drug and vaccine development.

The Xenopus oocyte expression system is ideal for electrophysiological characterization of voltage-dependent and ligand-dependent ion channels because of its relatively low background of endogenous channels and the large size of the cell. Here, we present a protocol to study voltage- and ligand-dependent activation of ion channels expressed in Xenopus oocytes using patch-clamp techniques designed to control both the membrane voltage and the intracellular solution. In this protocol, the large conductance voltage- and Ca2+-activated K+ (BK) channel is studied as an example. After injection of BK channel mRNA, oocytes are incubated for 2–7 d at 18°C. Inside-out membrane patches containing single or multiple BK channels are excised with perfusion of different solutions during recording. The protocol can be used to study structure–function relations for ion channels and neurotransmitter receptors.

A new self-compensation method is proposed in this paper. The impedance of a quartz tuning fork changes along with the temperature. A digital driving circuit is designed so that the frequency can be adjusted by controlling the phase. The relationship between the driving amplitude and the temperature is captured in a digital circuit, which can reflect the temperature information. As the relationship is nonlinear, after third-order fitting, the temperature has a precision of 2.356 °C. Finally, the bias stability is optimized and the drift decreases to 1.7 %.

BK channel β subunits (β1-β4) modulate the function of channels formed by slo1 subunits to produce tissue-specific phenotypes. The molecular mechanism of how the homologous β subunits differentially alter BK channel functions and the role of different BK channel functions in various physiologic processes remain unclear. By studying channels expressed in Xenopus laevis oocytes, we show a novel disulfide-cross-linked dimer conopeptide, Vt3.1 that preferentially inhibits BK channels containing the β4 subunit, which is most abundantly expressed in brain and important for neuronal functions. Vt3.1 inhibits the currents by a maximum of 71%, shifts the G-V relation by 45 mV approximately half-saturation concentrations, and alters both open and closed time of single channel activities, indicating that the toxin alters voltage dependence of the channel. Vt3.1 contains basic residues and inhibits voltage-dependent activation by electrostatic interactions with acidic residues in the extracellular loops of the slo1 and β4 subunits. These results suggest a large interaction surface between the slo1 subunit of BK channels and the β4 subunit, providing structural insight into the molecular interactions between slo1 and β4 subunits. The results also suggest that Vt3.1 is an excellent tool for studying β subunit modulation of BK channels and for understanding the physiological roles of BK channels in neurophysiology.