scholarly journals Further studies of ion channels in the electroreceptor of the skate through deep sequencing, cloning and cross species comparisons

2018 ◽  
Author(s):  
William T. Clusin ◽  
Ting-Hsuan Wu ◽  
Ling-Fang Shi ◽  
Peter N. Kao
Keyword(s):  
Amino Acids ◽  
Ion Channels ◽  
Potassium Channel ◽  
Calcium Channel ◽  
Beta Subunit ◽  
K Channel ◽  
Rna Seq ◽  
Α Subunit ◽  
Accessory Subunits ◽  
Voltage Dependent

AbstractOur comparative studies seek to understand the structure and function of ion channels in cartilaginous fish that can detect very low voltage gradients in seawater. The principal channels of the electroreceptor include a calcium activated K channel, whose α subunit is Kcnma1, a voltage-dependent calcium channel, Cacna1d, and a relatively uncharacterized K channel which interacts with the calcium channel to produce fast (20 Hz) oscillations. Large conductance calcium-activated K channels (BK) are comprised of four α subunits, encoded by Kcnma1 and modulatory β subunits of the Kcnmb class. We recently cloned and published the skate Kcnma1 gene and most of Kcnmb4 derived from using purified mRNA of homogenized isolated electroreceptors. Bellono et al. have recently performed RNA sequencing (RNA-seq) on purified mRNA from skate electroreceptors and found several ion channels including Kcnma1. We searched the the Bellono et al RNA-seq repository for additional channels and subunits. Our most significant findings are the presence of two Shaker type voltage dependent potassium channel sequences which are grouped together as isoforms in the data repository. The larger of these is a skate ortholog of the voltage dependent fast potassium channel Kv1.1, which is expressed at appreciable levels and seems likely to explain the 20 Hz oscillations believed to occur in vivo. The second was more similar to Kv1.5 than to Kv1.1 but was somewhat atypical. We also found a beta subunit sequence (Kcnab2) which appears not to cause fast inactivation due to specific structural features. The new channels and subunits were verified by RT-PCR and the Kv1.1 sequence was confirmed by cloning. We also searched the RNA-seq repository for accessory subunits of the calcium activated potassium channel, Kcnma1, and found a computer generated assembly that contained a complete sequence of its beta subunit, Kcnmb2. Skate Kcnmb2 has a total of 279 amino acids, with 51 novel amino acids at the N-terminus which may play a specific physiological role. This sequence was confirmed by PCR and cloning. However, skate Kcnmb2 is expressed at low levels in the electroreceptor compared to Kcnma1 and skate Kcnmb1 (beta1) is absent. The evolutionary origin of the newly described channels and subunits was studied by aligning skate sequences with human sequences and those found in related fish: the whale shark (R. typus) an elasmobranch, and ghost shark (C.milii). There is also homology with the lamprey, which has electroreceptors. An evolutionary tree is presented. Further research should include focusing on the subcellular locations of these channels in the receptor cells, their gating behavior, and the effects of accessory subunits on gating.

scholarly journals Chlorella virus ATCV-1 encodes a functional potassium channel of 82 amino acids

2009 ◽  
Vol 420 (2) ◽  
pp. 295-305 ◽  
Author(s):  
Sabrina Gazzarrini ◽  
Ming Kang ◽  
Alessandra Abenavoli ◽  
Giulia Romani ◽  
Claudio Olivari ◽  
...  
Keyword(s):  
Amino Acids ◽  
Potassium Channel ◽  
Xenopus Oocytes ◽  
Single Channel ◽  
K Channel ◽  
Channel Blockers ◽  
Selective Filter ◽  
Chlorella Virus ◽  
Charged Amino Acids ◽  
Voltage Dependent

Chlorella virus PBCV-1 (Paramecium bursaria chlorella virus-1) encodes the smallest protein (94 amino acids, named Kcv) previously known to form a functional K+ channel in heterologous systems. In this paper, we characterize another chlorella virus encoded K+ channel protein (82 amino acids, named ATCV-1 Kcv) that forms a functional channel in Xenopus oocytes and rescues Saccharomyces cerevisiae mutants that lack endogenous K+ uptake systems. Compared with the larger PBCV-1 Kcv, ATCV-1 Kcv lacks a cytoplasmic N-terminus and has a reduced number of charged amino acids in its turret domain. Despite these deficiencies, ATCV-1 Kcv accomplishes all the major features of K+ channels: it assembles into a tetramer, is K+ selective and is inhibited by the canonical K+ channel blockers, barium and caesium. Single channel analyses reveal a stochastic gating behaviour and a voltage-dependent conductance that resembles the macroscopic I/V relationship. One difference between PBCV-1 and ATCV-1 Kcv is that the latter is more permeable to K+ than Rb+. This difference is partially explained by the presence of a tyrosine residue in the selective filter of ATCV-1 Kcv, whereas PBCV-1 Kcv has a phenylalanine. Hence, ATCV-1 Kcv is the smallest protein to form a K+ channel and it will serve as a model for studying structure–function correlations inside the potassium channel pore.

scholarly journals K+ channel blockade in the NTS alters efficacy of two cardiorespiratory reflexes in vivo

1998 ◽  
Vol 274 (3) ◽  
pp. R677-R685 ◽  
Author(s):  
James W. Butcher ◽  
Julian F. R. Paton
Keyword(s):  
Potassium Channel ◽  
Neuronal Excitability ◽  
Baroreceptor Reflex ◽  
Right Atrial ◽  
K Channel ◽  
Reflex Bradycardia ◽  
Potassium Conductances ◽  
Voltage Dependent

We investigated the role of potassium conductances in the nucleus of the solitary tract (NTS) in determining the efficacy of the baroreceptor and cardiopulmonary reflexes in anesthetized rats. The baroreceptor reflex was elicited with an intravenous injection of phenylephrine to evoke a reflex bradycardia, and the cardiopulmonary reflex was evoked with a right atrial injection of phenylbiguanide. Microinjection of two Ca-dependent potassium channel antagonists (apamin and charybdotoxin) into the NTS potentiated the baroreceptor reflex bradycardia. This may reflect the increased neuronal excitability observed previously in vitro with these blockers. In contrast, the Ca-dependent potassium channel antagonists attenuated the cardiopulmonary reflex, whereas voltage-dependent potassium channel antagonists (4-aminopyridine and dendrotoxin) attenuated both the baro- and cardiopulmonary reflexes when microinjected into the NTS. The possibility that the reflex attenuation observed indicates a predominant distribution of certain potassium channels on γ-aminobutyric acid interneurons is discussed.

Differential distribution of potassium channels in acutely demyelinated, primary-auditory neurons in vitro

1996 ◽  
Vol 76 (1) ◽  
pp. 438-447 ◽  
Author(s):  
R. L. Davis
Keyword(s):  
Potassium Channels ◽  
Potassium Channel ◽  
Myelin Sheath ◽  
Lucifer Yellow ◽  
The Other ◽  
K Channel ◽  
Channel Activity ◽  
Auditory Neurons ◽  
Axonal Membrane ◽  
Voltage Dependent

1. Single-channel recordings of potassium channel activity were made from two populations of primary-auditory neurons maintained in tissue culture. The saccular nerve, which is the auditory component of the eighth cranial nerve in goldfish, was separated into two branches according to its peripheral innervation pattern. Neurons which innervated the rostral saccular macula corresponded to a class of cells that showed spike frequency adaptation; whereas, neurons which innervated the caudal macula were consistent with another type of cell that demonstrated bursting spontaneous firing patterns in vivo. Both somatic and internodal axonal membranes from each of these neuronal classes were studied after acute removal of the myelin sheath by microdissection. 2. Dye injections were used to discriminate neuronal from myelin membrane. After successful removal of the myelin, patch electrodes containing Lucifer yellow were used to fill a neuron and reveal its morphology within the myelin sheath. Patches on myelin led to filling of Schwann cells that surrounded the neuron. 3. Four kinds of potassium channels were observed and characterized according to unitary conductance, inactivation, and sensitivity to internal calcium. Three voltage-dependent K+ channel types were found on the somatic and axonal membrane of the two neuronal populations. Two channel types showed voltage-dependent inactivation and had average conductances of 32 and 19 pS, each with distinctive subconductance states. The third type of channel activity had an estimated conductance of 12 pS and was noninactivating. 4. The fourth type of channel was the Ca2(+)-activated K+ channel (k(Ca)), which was classified by the dependence of its activity on the calcium concentration at its cytoplasmic surface. Unlike the other three potassium channel types, this kind of channel was found exclusively on neurons that innervated the caudal sensory epithelium. As with the other kinds of potassium channels, it was found on both somatic and axonal internodal membranes.

scholarly journals Uncharged S4 Residues and Cooperativity in Voltage-dependent Potassium Channel Activation

1998 ◽  
Vol 111 (3) ◽  
pp. 421-439 ◽  
Author(s):  
Catherine J. Smith-Maxwell ◽  
Jennifer L. Ledwell ◽  
Richard W. Aldrich
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Potassium Channel ◽  
Voltage Dependence ◽  
Channel Gating ◽  
Ionic Currents ◽  
Channel Activation ◽  
Functional Changes ◽  
Activation Pathway ◽  
Voltage Dependent

Substitution of the S4 of Shaw into Shaker alters cooperativity in channel activation by slowing a cooperative transition late in the activation pathway. To determine the amino acids responsible for the functional changes in Shaw S4, we created several mutants by substituting amino acids from Shaw S4 into Shaker. The S4 amino acid sequences of Shaker and Shaw S4 differ at 11 positions. Simultaneous substitution of just three noncharged residues from Shaw S4 into Shaker (V369I, I372L, S376T; ILT) reproduces the kinetic and voltage-dependent properties of Shaw S4 channel activation. These substitutions cause very small changes in the structural and chemical properties of the amino acid side chains. In contrast, substituting the positively charged basic residues in the S4 of Shaker with neutral or negative residues from the S4 of Shaw S4 does not reproduce the shallow voltage dependence or other properties of Shaw S4 opening. Macroscopic ionic currents for ILT could be fit by modifying a single set of transitions in a model for Shaker channel gating (Zagotta, W.N., T. Hoshi, and R.W. Aldrich. 1994. J. Gen. Physiol. 103:321–362). Changing the rate and voltage dependence of a final cooperative step in activation successfully reproduces the kinetic, steady state, and voltage-dependent properties of ILT ionic currents. Consistent with the model, ILT gating currents activate at negative voltages where the channel does not open and, at more positive voltages, they precede the ionic currents, confirming the existence of voltage-dependent transitions between closed states in the activation pathway. Of the three substitutions in ILT, the I372L substitution is primarily responsible for the changes in cooperativity and voltage dependence. These results suggest that noncharged residues in the S4 play a crucial role in Shaker potassium channel gating and that small steric changes in these residues can lead to large changes in cooperativity within the channel protein.

scholarly journals Voltage-dependent gating and gating charge measurements in the Kv1.2 potassium channel

2015 ◽  
Vol 145 (4) ◽  
pp. 345-358 ◽  
Author(s):  
Itzel G. Ishida ◽  
Gisela E. Rangel-Yescas ◽  
Julia Carrasco-Zanini ◽  
León D. Islas
Keyword(s):  
Ion Channels ◽  
Potassium Channel ◽  
Structural Data ◽  
Activation Mechanism ◽  
Voltage Sensitivity ◽  
Structural Interpretation ◽  
Kv Channel ◽  
Gating Charge ◽  
Voltage Dependent ◽  
Electrophysiological Findings

Much has been learned about the voltage sensors of ion channels since the x-ray structure of the mammalian voltage-gated potassium channel Kv1.2 was published in 2005. High resolution structural data of a Kv channel enabled the structural interpretation of numerous electrophysiological findings collected in various ion channels, most notably Shaker, and permitted the development of meticulous computational simulations of the activation mechanism. The fundamental premise for the structural interpretation of functional measurements from Shaker is that this channel and Kv1.2 have the same characteristics, such that correlation of data from both channels would be a trivial task. We tested these assumptions by measuring Kv1.2 voltage-dependent gating and charge per channel. We found that the Kv1.2 gating charge is near 10 elementary charges (eo), ∼25% less than the well-established 13–14 eo in Shaker. Next, we neutralized positive residues in the Kv1.2 S4 transmembrane segment to investigate the cause of the reduction of the gating charge and found that, whereas replacing R1 with glutamine decreased voltage sensitivity to ∼50% of the wild-type channel value, mutation of the subsequent arginines had a much smaller effect. These data are in marked contrast to the effects of charge neutralization in Shaker, where removal of the first four basic residues reduces the gating charge by roughly the same amount. In light of these differences, we propose that the voltage-sensing domains (VSDs) of Kv1.2 and Shaker might undergo the same physical movement, but the septum that separates the aqueous crevices in the VSD of Kv1.2 might be thicker than Shaker’s, accounting for the smaller Kv1.2 gating charge.

The Voltage-Dependent K+ Channel Family

2019 ◽  
Author(s):  
Hanne B. Rasmussen ◽  
James S. Trimmer
Keyword(s):  
Ion Channels ◽  
Neuronal Excitability ◽  
Expression Patterns ◽  
Basic Knowledge ◽  
K Channel ◽  
Dependent Manner ◽  
Kv Channels ◽  
Voltage Dependent ◽  
The Individual ◽  
Α Subunits

Voltage-dependent K+ (potassium; Kv) channels are ion channels that critically impact neuronal excitability and function. Four principal α subunits assemble to create a membrane-spanning pore that opens in a voltage-dependent manner to allow the selective passage of K+ ions across the cell membrane. Forty human genes encoding Kv channel α subunits have been identified, and most of them are expressed in the nervous system. The individual Kv subunits display unique cellular and subcellular expression patterns and co-assemble into distinct homo- and hetero-tetrameric channels that differ in their electrophysiological and pharmacological properties, and their sensitivity to dynamic modulation, by cellular signaling pathways. The resulting diversity allows Kv channels to impact all steps in electrical information processing, as well as numerous other aspects of neuronal functions, including those in which they appear to play a non-conducting role. This chapter reviews the current basic knowledge about this large and important family of ion channels.

Charged amino acids near the H5-region and channel functions in voltage-dependent potassium channel, Kv1.5

1994 ◽  
Vol 19 ◽  
pp. S11
Author(s):  
Tetsushi Furukawa ◽  
Yoshifumi Katayama ◽  
Tei-Ichi Yamane ◽  
Masayasu Hiraoka
Keyword(s):  
Amino Acids ◽  
Potassium Channel ◽  
Charged Amino Acids ◽  
Voltage Dependent
Sign in / Sign up

Export Citation Format

Share Document