Integral membrane voltage-gated potassium ion channel - carries type-A potassium current responsible for the repolarization of the cell - regulates neurotransmitter release at the synapse - regulates sleep - neuromuscular junction
Gene model reviewed during 5.55
Gene model reviewed during 5.45
Low-frequency RNA-Seq exon junction(s) not annotated.
Annotated transcripts do not represent all possible combinations of alternative exons and/or alternative promoters.
Gene model reviewed during 6.02
Unconventional translation start (AUU) postulated; FlyBase analysis.
Gene model reviewed during 6.08
Homotetramer or heterotetramer of potassium channel proteins.
The transmembrane segment S4 functions as voltage-sensor and is characterized by a series of positively charged amino acids at every third position. Channel opening and closing is effected by a conformation change that affects the position and orientation of the voltage-sensor paddle formed by S3 and S4 within the membrane. A transmembrane electric field that is positive inside would push the positively charged S4 segment outwards, thereby opening the pore, while a field that is negative inside would pull the S4 segment inwards and close the pore. Changes in the position and orientation of S4 are then transmitted to the activation gate formed by the inner helix bundle via the S4-S5 linker region.
Click to get a list of regulatory features (enhancers, TFBS, etc.) and gene disruptions (point mutations, indels, etc.) within or overlapping Dmel\Sh using the Feature Mapper tool.
GBrowse - Visual display of RNA-Seq signalsView Dmel\Sh in GBrowse 2
Please Note FlyBase no longer curates genomic clone accessions so this list may not be complete
Please Note This section lists cDNAs and ESTs that fall within the genomic extent of the gene model, which may include cDNAs and ESTs of genes within introns, or of overlapping genes. Please see GBrowse for alignment of the cDNAs and ESTs to the gene model.
For each fully sequenced cDNA the DGRC maintains various forms of the cDNA (e.g tagged or untagged) in several different host vectors for subsequent cloning and expression in Drosophila and Drosophila cell lines.
Sh K+ conductance is important for neural coding precision and as a mechanism for selectively amplifying graded signals in neurons.
The intracellular gate of Sh channels is capable of regulating access even by the small cations Cd2+ and Ag+. It can exclude small neutral or negatively charged molecules, suggesting that the gate operates by steric exclusion rather than electrostatically.
Mutants are hypersensitive to paraquat.
Three permeant ion binding-sites in the pore of Sh channel are studied. Pore-lining resides are identified that appear to contribute to the formation of two deeper sites. Results are consistent with a mechanism of gating that operates by pinching off access of the deep pore to the internal or external solution.
The membrane potential plays a crucial role in the loss of conductance. There is a close connection between the gating and conduction function of the membrane.
Fluorescent labeling allows the examination of voltage-dependent conformational changes in different regions of the Sh channel. The S2 segment may undergo voltage sensitive conformational changes that precede those in the S4 segment. Fluorescence changes in the pore correlate with the voltage dependence and time course of ionic activation and slow inactivation.
Incorporation of Npg (an unnatural amino acid (2-nitrophenyl)glycine) produces peptide backbone cleavage at the site of the novel residue, by analogy with other 2-nitrobenzyl systems.
Recovery from inactivation in Sh K+ channels begins with no delay on repolarisation. Hyperpolarisation hastens only the initial phase of recovery, yet retards the later phase of recovery by increasing the proportion of slow components. The fast and slow components primarily correspond to recovery via the open state and via the closed state, respectively. Sh K+ channel deactivation hinders, rather than facilitates, the unbinding of the inactivating particle and therefore retards recovery from inactivation, whereas external K+ may enhance unbinding of the particle by binding to a site located near the external entrance of the pore.
A series of positions in the S6 transmembrane regions are found to react rapidly with water soluble thiol reagents in the open state but not the closed state. An open-channel blocker can protect several of these Cys residues, showing that they lie in the ion-conduction pore. Results suggest the channels open and close by the movement of an intracellular gate, distinct from the selectivity filter, that regulates access to the pore.
The spatio-temporal expression of Sh protein in the developing and adult nervous system has been analysed.
Probing the boundary of the electric field with protons indicates that the voltage-sensing residue 365 lies on an internally faced narrow crevice in the resting state, while the sensing charge at position 368 sits in an externally faced crevice in the open state of the channel. Both residues move entirely from the internal to external medium in each stroke of the voltage sensor. The translocation of the two residues accounts for 66% of the total gating charge.
The C-terminal sequences of Fas2 and Sh are both necessary and sufficient for targeting to the subsynaptic muscle membrane at the larval neuromuscular junction, and this localization depends on the product of dlg1.
Opening of a Sh channel is associated with a displacement of 13.6 electron charge units. Mutational analysis reveals that movement of the amino-terminal half but not the carboxy-terminal end of the S4 segment underlies gating charge, and this portion of the S4 segment appears to move across the entire transmembrane voltage difference in association with channel activation.
When applied to Sh channels expressed in mammalian cells, quaternary ammonium blockers produce use-dependent inhibition by promoting an intrinsic conformational change, C-type inactivation, from which recovery is slow.
Subunits from eag and Sh functionally interact in Xenopus oocytes, most likely as heteromultimeric channels. Site directed mutagenesis indicates the eag carboxyl terminus is crucial for the interaction with Sh.
The effects of amino acid replacements on AgTx2 affinity define the eccentricity of amino acids in the pore entryway and imply a different secondary structure for the amino and carboxyl ends of the pore loop.
The putative voltage-sensing charges of S4 actually reside in the membrane and move outward when channels open.
The S4 domain contains the gating charge. Activation consists of the movement of the outer portion of S4 into the extracellular fluid from a position that is buried in the resting state, thus generating the gating current.
Triple mutation can convert the outwardly rectifying Sh channel to an inward rectifier. The conversion does not rely on a difference in sign or direction of charge movement of the voltage sensor, since activation of the Sh outward rectifier is due to a different gate than activation of the mutant inward rectifier.
Studies of the interaction of Agitoxin2 with Sh channels reveals a shallow vestibule formed by the pore loops at the Sh channel entryway. The selectivity filter is located at the center of the vestibule close to (around 5 Angstroms from) the extracellular solution.
Voltage sensing residues have been mapped to the S2 and S4 segments of the Sh K+ channel.
Hk β subunit modulates a wide range of the Sh K+ current properties, inducing its amplitude, activation and inactivation, temperature dependence and drug sensitivity. Modulation is thought to occur via hydrophobic interactions, Hk β subunits modulate Sh channel formation in the cytoplasmic pore region.
The monkey Cos cell line is a reasonable system for transient expression of K+ channels, particularly those with fast inactivation kinetics.
Synthetic inactivating and synthetic noninactivating peptides are used to investigate whether the single amino acid change in the peptide sequence determines alteration in the conformation of the "ball" peptide that might explain the loss of function. The two peptides have a different potential capacity to become structured into a given conformation.
Modulation of Sh channels using serotonin has been studied in a semi-intact preparation of the retina.
Using Ecol\lacZ reporter gene for accurate splicing of variable Sh 3' domains the expression pattern in transgenic animals indicates both temporal and spatial regulation of 3' splice choice. Tissue-specific expression of functionally distinct Sh K+ channels is regulated, in part, at the level of pre-mRNA splicing and implicates sequences in or around the 3' splice sites in regulating the choice of 3' domain.
Mutations conveying both strong and weak suppression of the primary S4 neutralisation mutations K374Q and R377Q have been obtained identifying likely short- and long range electrostatic interactions among transmembrane charged residues.
Immunochemical techniques have identified so called short Sh cDNAs. Genetic criteria determines a good part of the protein variety is generated by alternative splicing. The expression pattern of Sh splice variants changes dramatically throughout development.
Coimmunoprecipitation and yeast two-hybrid system studies demonstrate the association of the hydrophilic N-terminal domains of the genes encoding channel proteins plays an important role in determining the specificity of α subunit association to form heteromultimeric potassium channels.
Abnormal function of Sh K+ channels at motor nerves specifically abolishes post-tetanic potentiation (PTP) in the larval neuromuscular junction.
The time course of N-type inactivation of Sh K+ channels is prolonged upon exposure of the cytoplasmic face to phosphatases and this effect is completely reversed by subsequent application of the purified catalytic subunit of the cAMP-dependent protein kinase (PKA) and ATP.
The functional consequences of introducing point mutations into the signature sequence of Sh channel is studied.
Physical dimensions and biochemical characteristics of Sh channels expressed in Sf9 cells studied using electron microscopy demonstrates they have a tetrameric subunit composition. Negative staining reveals a four-fold symmetric tetramer with a large, central vestibule that presumably constitutes part of the pathway for ions.
4-AP binds to an internally accessible site of Sh and alters channel gating. The binding is state-dependent and when bound 4-AP prevents channels from opening and blocks distinct conformational rearrangements.
Ion permeation of Sh channels appears to have many of the properties consistent with multi-ion pores.
Expression of Sh using a mammalian transient transfection system allows N-ethlymaleamide-labelled charybdotoxin (NEM-CTX) quantification and characterisation of assembled tetrameric channels in both isolated membrane fragments and detergent extracts. The channels produced can be functionally reconstituted into model membranes accessible to direct electrical recording.
Ion channel mutants alter synaptic activity at the embryonic neuromuscular junction (NMJ). GluRIIA expression in the postsynaptic membrane is reduced by changes in presynaptic electrical activity. The size of the synaptic domain depends on the level of neural activity during embryonic synaptogenesis.
The conduction properties of the cloned Sh channel are characteristic of those traditionally found in other K+ channels.
Electrophysiological measurements and numerical simulation studies of channels expressed in transfected cells reveals that the slow properties of the encoded channel help to determine the voltage trajectories in the synthetic neurons. Slow inactivation of transient K+ currents could play a role in the encoding properties of real neurons.
In eag,Sh hyperexcitable double mutants nerve/muscle synaptic ultrastructure is dramatically altered. Two types of synaptic vesicle are depleted and a third is altered in appearance, and there are changes in number and appearance of synaptic densities, and multivesicular bodies.
The role of conserved L370 residue is investigated. Substitutions result in alteration in the relative stabilities of open, closed and inactivated conformational states, corresponding to the size and hydrophobicity of the substituted residue. Data suggests that nonconservative substitutions of L370 influence the ability of Sh subunits to assemble into functional channel complexes. All observations are consistent with the idea that L370 and other residues in the region undergo protein interactions that are important determinants in the formational structure of the channel.
In mutant channels that have completely abolished ion conduction the channel can still undergo the closed-open conformation in response to voltage changes.
The effect of mutations that decrease the steepness of the conductance-voltage relationship is to alter the kinetics and equilibria of charge-moving transitions.
A 20 amino acid synthetic peptide corresponding to the amino terminal of the B splice variant of the Sh K+ channel, and responsible for its fast inactivation, can block large conductance Ca2+-dependent K+ channels from rat brain and muscle, in two kinetically distinct types of blocking, "long" and "short". Pharmacological experiments indicate the peptide induces short block by binding in the pore of Ca2+-dependent K+ channels. This short block and the inactivation of Sh exhibit similar characteristics, therefore the binding region for the peptide in the pore regions is conserved in these very different K+ channels.
Effects of potassium channel blocking drugs on the presynaptic action potential repolarization after electrotonic stimulation was studied. At least four K+ currents contribute to repolarization of the nerve terminal.
The effects of mutations where the pore sequences mimic those of the cyclic nucleotide gated channels were tested in the Xenopus oocyte expression system. These channels behave as cyclic nucleotide gated channels, demonstrating that the two physiologically distinct types of channel are actually closely related.
Channel gating is investigated.
Cell-free protein translation, microsomal membrane processing of nascent channel protein, and reconstitution of newly synthesised ion channels into planar lipid bilayers generated glycosylated, active, functional Shaker potassium channels.
Shaker splice variant B 20 amino acid inactivating peptide (known as "ball peptide" BP) interacts with Ca2+-activated K+ channels in porcine coronary smooth muscle from cytoplasmic side only, producing inhibition of channel activity. Effect is reversible and dose- and voltage-dependent. BP binds KCa channels in a bimolecular reaction, and results suggest that Ca-dependent K+ channels and the Sh channels have the same receptor for "BP".
Subunit assembly of Sh does not depend on the leucine heptad repeat. Substitutions of the Leu residues in the repeat produce large effects on the observed voltage dependence of conductance voltage and prepulse inactivation curves. Results suggest the Leu residues mediate interactions that play an important role in the transduction of charge movement into channel opening and closing.
Site directed mutagenesis of the S4 sequence of the Sh potassium channel and electrophysiological analysis suggest that voltage-dependent activation involves the S4 sequence but it is not solely due to electrostatic interactions.
An amino acid residue that specifically affects the affinity for the intracellular tetraethylammonium (TEA) has been identified and is in the middle of a conserved stretch of 18 amino acids. Results suggest this conserved region is intimately involved in the formation of the ion conduction pore of the channel.
Molecular region of Sh has been identified that influences ionic selectivity. H5 (fifth hydrophobic region) is likely to line th pore of the potassium channel.
Variations in amino acid sequence in a small region of Sh near the amino terminus can cause changes in channel inactivation rates.
The effect of Ca2+ removal causes a nonselective leak of expressed K+ channels due to a massive functional alteration of the channel.
Although Sh, Shal, Shab and Shaw proteins share a conserved structral organisation, their potassium channel currents (expressed in Xenopus oocytes) differ greatly in individual kinetic properties and voltage sensitivity.
The Sh locus can be dissected by means of aneuploids into three regions: maternal effect (ME), viable (V) and haplolethal (HL). Mutations of the ME region of the Sh locus affect oogenesis and the differentiation of the hypoderm and/or the physiology of the CGF.
Expression of mutated Sh potassium channels in Xenopus oocytes has identified a region near the amino terminus that has an important role in inactivation of the channel.
Cloning and characterisation of Sh has identified structural elements involved in potassium channel gating. The amino and carboxy terminus are specialised for, and appear to interact in, inactivation gating.
Analysis of Sh polypeptides expressed in Xenopus oocytes suggests that they assemble to form multimeric channels.
Both the 5' and 3' variable domains of the Sh gene product influence the kinetics of macroscopic inactivation. The amino domain dictates a general range of inactivation properties. Chimaeric Sh cDNAs exhibit variable time constants for inactivation, variable incomplete inactivation and variable recovery from inactivation.
Specific amino acid residues have been identified that affect channel blockade by an external source tetraethylammonium (tetraethylammonium (TEA)), as well as conductance of ions through the pore.
The distribution of Sh proteins in the brain of the adult fly has been determined.
Sh, Shal, Shab and Shaw encode voltage gated potassium channels with widely varying kinetics (rate of macroscopic current activation and inactivation) and voltage sensitivity of steady state inactivation.
Potassium channel diversity could result from an extended gene family as well as from alternate splicing of the Sh primary transcript.
The Glu residue at position 422 is near or in the externally facing mouth of the potassium conduction pathway. The positively charged toxin charybdotoxin (CTX) is electrostatically focused toward its blocking site by the negative potential set up by Glu-422.
P-element mediated germline transformation has been used to express ShB channel in Sh mutants. The transformant A-current inactivates rapidly and recovers from inactivation similarly to ShB channels expressed in Xenopus oocytes. Unlike channels in oocytes the transformant A-current is insensitive to charybdotoxin (CTX).
The molecular transition rates leading to the first opening of ShB and ShD channels are voltage-dependent. All further transitions are independent of voltage. The difference in macroscopic current between the channels is due to the quantitative difference in transition rates. Voltage dependence of macroscopic currents is determined by the voltage dependency on the time to first opening.
Transcripts synthesised in vitro from Sh cDNA express A-type K+ currents when injected into Xenopus oocytes.
Two basic forms of conceptual proteins are encoded by Sh cDNA: the more common form containing seven potential membrane spanning domains and the other form containing 3 potential membrane spanning domains. The cDNAs contain variable 5' and 3' ends joined to a constant central region. The different cDNAs encode proteins with distinct structural features.
At least four probable components of potassium channels are encoded at the Sh locus by a family of alternatively spliced transcripts.
Four different Sh mRNAs have been tested and found to produce A currents in Xenopus oocytes. The four currents differ in kinetics of inactivation indicating that the Sh products may contribute to kinetic diversity in A-channels. Sequences in both the amino- and carboxy-terminal regions are important for inactivation.
The Sh genomic region has been cloned and the transcription pattern of this region has been analysed.
Some Sh phenotypes are suppressed by nap and para alleles; i.e., in double mutant combinations, abnormal leg-shaking, repetitive firing of larval action potentials, and transmitter release at larval neuromuscular junction are nearly normal; the interactions are not allele-specific (Ganetzky and Wu, 1982a; Ganetzky and Wu, 1982b). Some Sh phenotypes are enhanced by eag; i.e., in double mutant combinations, abnormal leg-shaking, repetitive firing of larval action potentials and transmitter release are more extreme; also, adults have down-turned wings and dented-in thoraces at the sites of the dorsal longitudinal muscle insertions; the interactions are not allele-specific (Ganetzky and Wu, 1983). Some Sh phenotypes are enhanced by dnc; i.e., in double mutant combinations, abnormal leg-shaking is more extreme; abnormal spontaneous activity is seen in the giant fiber (Ferrus and Tanouye). The breakpoint of T(1;2)B27, induced in a Sh14 background, causes an alteration in the pattern of leg-shaking (Tanouye and Ferrus).
Under moderate ether anesthesia, legs shake abnormally, antennae twitch, abdomen pulsates; wings scissor in some alleles; very little effect in deeply etherized flies; unetherized mutants twitch and shudder occasionally; severed legs shake (Kaplan and Trout, 1969; Trout and Kaplan, 1973; Tanouye, Ferrus and Fujita, 1981; Ganetzky and Wu, 1982a; Tanouye and Ferrus, 1985). Structural gene for several types of potassium channel (Iverson et al., 1988; Timpe et al. Jan, 1988). Abnormal action potential repolarization of adult giant fiber; repetitive firing of action potentials in larval nerves; prolonged transmitter release at larval neuromuscular junction (Jan, Jan and Dennis, 1977; Tanouye, Ferrus and Fujita, 1981; Ganetzky and Wu, 1982b; Tanouye and Ferrus, 1985). Abnormal in one class of potassium channel (A channel) present in embryonic myocytes, larval and pupal muscle (Salkoff and Wyman, 1981; Salkoff, 1983; Wu and Haugland, 1985; Timpe and Jan, 1987; Haugland and Wu, 1990). Sh mutations do not affect four other distinct potassium-channel types (KD, K1, A2, Calcium-gated) (Salkoff and Wyman, 1981; Salkoff, 1983; Wu et al., 1983; Solc et al., 1987; Solc and Aldrich, 1988). Males carrying hemizygous deletions of Sh are viable (Tanouye, Ferrus and Fujita, 1981). Abnormal associative learning in some paradigms (Tully); activity patterns high, but show normal circadian rhythmicity (Konopka).