LQT2 : amplitude reduction and loss of selectivity in the tail that wags the HERG channel.
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Cation Transport Proteins↗
Biomedical subjects
G A Robertson
Publications and source records attributed to G A Robertson.
Novel mechanism associated with an inherited cardiac arrhythmia: defective protein trafficking by the mutant HERG (G601S) potassium channel.
BACKGROUND: The congenital long-QT syndrome (LQTS) is an inherited disorder characterized by a prolonged cardiac action potential and a QT interval that leads to arrhythmia. Mutations in the human ether-a-go-go-related gene (HERG), which encodes the rapidly activating component of the delayed rectifier current (IKr), cause chromosome 7-linked LQTS (LQT2). Studies of mutant HERG channels in heterologous systems indicate that the mechanisms mediating LQT2 are varied and include mutant subunits that form channels with altered kinetic properties or nonfunctional mutant subunits. We recently reported a novel missense mutation of HERG (G601S) in an LQTS family that we have characterized in the present work. METHODS AND RESULTS: To elucidate the electrophysiological properties of the G601S mutant channels, we expressed these channels in mammalian cells and Xenopus oocytes. The G601S mutant produced less current than wild-type channels but exhibited no change in kinetic properties or dominant-negative suppression when coexpressed with wild-type subunits. To examine the cellular trafficking of mutant HERG channel subunits, enhanced green fluorescent protein tagging and Western blot analyses were performed. These showed deficient protein trafficking of the G601S mutant to the plasma membrane. CONCLUSIONS: Our results from both the Xenopus oocyte and HEK293 cell expression systems and green fluorescent protein tagging and Western blot analyses support the conclusion that the G601S mutant is a hypomorphic mutation, resulting in a reduced current amplitude. Thus, it represents a novel mechanism underlying LQT2.
Animals↗
The eag family of K+ channels in Drosophila and mammals.
Mutations of eag, first identified in Drosophila on the basis of their leg-shaking phenotype, cause repetitive firing and enhanced transmitter release in motor neurons. The encoded EAG polypeptide is related both to voltage-gated K+ channels and to cyclic nucleotide-gated cation channels. Homology screens identified a family of eag-related channel polypeptides, highly conserved from nematodes to humans, comprising three subfamilies: EAG, ELK, and ERG. When expressed in frog oocytes, EAG channels behave as voltage-dependent, outwardly rectifying K(+)-selective channels. Mutations of the human eag-related gene (HERG) result in a form of cardiac arrhythmia that can lead to ventricular fibrillation and sudden death. Electrophysiological and pharmacological studies have provided evidence that HERG channels specify one component of the delayed rectifier, IKr, that contributes to the repolarization phase of cardiac action potentials. An important role for HERG channels in neuronal excitability is also suggested by the expression of these channels in brain tissue. Moreover, mutations of ERG-type channels in the Drosophila sei mutant cause temperature-induced convulsive seizures associated with aberrant bursting activity in the flight motor pathway. The in vivo function of ELK channels has not yet been established, but when these channels are expressed in frog oocytes, they display properties intermediate between those of EAG- and ERG-type channels. Coexpression of the K(+)-channel beta subunit encoded by Hk with EAG in oocytes dramatically increases current amplitude and also affects the gating and modulation of these currents. Biochemical evidence indicates a direct physical interaction between EAG and HK proteins. Overall, these studies highlight the diverse properties of the eag family of K+ channels, which are likely to subserve diverse functions in vivo.
Alternative Splicing↗
Functional analysis of a mouse brain Elk-type K+ channel.
Members of the Ether à go-go (Eag) K+ channel subfamilies Eag, Erg, and Elk are widely expressed in the nervous system, but their neural functions in vivo remain largely unknown. The biophysical properties of channels from the Eag and Erg subfamilies have been described, and based on their characteristic features and expression patterns, Erg channels have been associated with native currents in the heart. Little is known about the properties of channels from the Elk subfamily. We have identified a mouse gene, Melk2, that encodes a predicted polypeptide with 48% amino acid identity to Drosophila Elk but only 40 and 36% identity with mouse Erg (Merg) and Eag (Meag), respectively. Melk2 RNA appears to be expressed at high levels only in brain tissue. Functional expression of Melk2 in Xenopus oocytes reveals large, transient peaks of current at the onset of depolarization. Like Meag currents, Melk2 currents activate relatively quickly, but they lack the nonsuperimposable Cole-Moore shift characteristic of the Eag subfamily. Melk2 currents are insensitive to E-4031, a class III antiarrhythmic compound that blocks the Human Ether-à-go-go-Related Gene (HERG) channel and its counterpart in native tissues, IKr. Melk2 channels exhibit inward rectification because of a fast C-type inactivation mechanism, but the slower rate of inactivation and the faster rate of activation results in less inward rectification than that observed in HERG channels. This characterization of Melk currents should aid in identification of native counterparts to the Elk subfamily of channels in the nervous system.
Action Potentials↗
Transfer of rapid inactivation and sensitivity to the class III antiarrhythmic drug E-4031 from HERG to M-eag channels.
The gating behaviour and pharmacological sensitivity of HERG are remarkably different from the corresponding properties of M-eag, a structurally similar member of the Eag family of potassium channels. In contrast to HERG, M-eag exhibits no apparent inactivation and little rectification, and is insensitive to the class III antiarrhythmic drug E-4031. We generated chimeric channels of HERG and M-eag sequences and made point mutations to identify the region necessary for rapid inactivation in HERG. This region includes the P region and half of the S6 putative transmembrane domain, including sites not previously associated with inactivation and rectification in HERG. Transfer of a small segment of the HERG polypeptide to M-eag, consisting largely of the P region and part of the S6 transmembrane domain, is sufficient to confer rapid inactivation and E-4031 sensitivity to M-eag. This region differs from the corresponding region in M-eag by only fifteen residues. Previous hypotheses that rapid inactivation of HERG channels occurs by a C-type inactivation mechanism are supported by the parallel effects on rates of HERG inactivation and Shaker C-type inactivation by a series of mutations at two equivalent sites in the polypeptide sequences. In addition to sites homologous to those previously described for C-type inactivation in Shaker, inactivation in HERG involves a residue in the upstream P region not previously associated with C-type inactivation. Although this site is equivalent to one implicated in P-type inactivation in Kv2.1 channels, our data are most consistent with a single, C-type inactivation mechanism.
Amino Acid Sequence↗
Properties of HERG channels stably expressed in HEK 293 cells studied at physiological temperature.
We have established stably transfected HEK 293 cell lines expressing high levels of functional human ether-a go-go-related gene (HERG) channels. We used these cells to study biochemical characteristics of HERG protein, and to study electrophysiological and pharmacological properties of HERG channel current at 35 degrees C. HERG-transfected cells expressed an mRNA band at 4.0 kb. Western blot analysis showed two protein bands (155 and 135 kDa) slightly larger than the predicted molecular mass (127 kDa). Treatment with N-glycosidase F converted both bands to smaller molecular mass, suggesting that both are glycosylated, but at different levels. HERG current activated at voltages positive to -50 mV, maximum current was reached with depolarizing steps to -10 mV, and the current amplitude declined at more positive voltages, similar to HERG channel current expressed in other heterologous systems. Current density at 35 degrees C, compared with 23 degrees C, was increased by more than twofold to a maximum of 53.4 +/- 6.5 pA/pF. Activation, inactivation, recovery from inactivation, and deactivation kinetics were rapid at 35 degrees C, and more closely resemble values reported for the rapidly activating delayed rectifier K+ current (I(Kr)) at physiological temperatures. HERG channels were highly selective for K+. When we used an action potential clamp technique, HERG current activation began shortly after the upstroke of the action potential waveform. HERG current increased during repolarization to reach a maximum amplitude during phases 2 and 3 of the cardiac action potential. HERG contributed current throughout the return of the membrane to the resting potential, and deactivation of HERG current could participate in phase 4 depolarization. HERG current was blocked by low concentrations of E-4031 (IC50 7.7 nM), a value close to that reported for I(Kr) in native cardiac myocytes. Our data support the postulate that HERG encodes a major constituent of I(Kr) and suggest that at physiological temperatures HERG contributes current throughout most of the action potential and into the postrepolarization period.
Action Potentials↗
Regulation of deactivation by an amino terminal domain in human ether-à-go-go-related gene potassium channels.
Abnormalities in repolarization of the cardiac ventricular action potential can lead to life-threatening arrhythmias associated with long QT syndrome. The repolarization process depends upon the gating properties of potassium channels encoded by the human ether-à-go-go-related gene (HERG), especially those governing the rate of recovery from inactivation and the rate of deactivation. Previous studies have demonstrated that deletion of the NH2 terminus increases the deactivation rate, but the mechanism by which the NH2 terminus regulates deactivation in wild-type channels has not been elucidated. We tested the hypothesis that the HERG NH2 terminus slows deactivation by a mechanism similar to N-type inactivation in Shaker channels, where it binds to the internal mouth of the pore and prevents channel closure. We found that the regulation of deactivation by the HERG NH2 terminus bears similarity to Shaker N-type inactivation in three respects: (a) deletion of the NH2 terminus slows C-type inactivation; (b) the action of the NH2 terminus is sensitive to elevated concentrations of external K+, as if its binding along the permeation pathway is disrupted by K+ influx; and (c) N-ethylmaleimide, covalently linked to an aphenotypic cysteine introduced within the S4-S5 linker, mimics the N deletion phenotype, as if the binding of the NH2 terminus to its receptor site were hindered. In contrast to N-type inactivation in Shaker, however, there was no indication that the NH2 terminus blocks the HERG pore. In addition, we discovered that separate domains within the NH2 terminus mediate the slowing of deactivation and the promotion of C-type inactivation. These results suggest that the NH2 terminus stabilizes the open state and, by a separate mechanism, promotes C-type inactivation.
Animals↗
Therapeutic fasciectomy for eosinophilic fasciitis.
Eosinophilic fasciitis is an uncommon and potentially debilitating sclerodermalike connective tissue disorder. A patient is presented for whom medical and pharmacological management had failed to prevent the progression of forearm pain, fibrosis, and flexor contractures. Definitive management with forearm fasciectomy resolved the pain and contractures of the hand. Therapeutic fasciectomy may be a consideration earlier in the course of this disease to ameliorate its debilitating factors. The literature is reviewed.
Adult↗
Two isoforms of the mouse ether-a-go-go-related gene coassemble to form channels with properties similar to the rapidly activating component of the cardiac delayed rectifier K+ current.
HERG, the human ether-a-go-go-related gene, encodes a K(+)-selective channel with properties similar to the rapidly activating component of the delayed rectifier K+ current (IKr). Mutations of HERG cause the autosomal-dominant long-QT syndrome (LQTS), presumably by disrupting the normal function of IKr. The current produced by HERG is not identical to IKr, however, and the mechanism by which HERG mutations cause LQTS remains uncertain. To better define the role of Erg in the heart, we cloned Merg1 from mouse genomic and cardiac cDNA libraries. Merg1 has 16 exons and maps to mouse chromosome 5 in an area syntenic to human chromosome 7q, the map locus of HERG. We isolated three cardiac isoforms of Merg1: Merg1a is homologous to HERG and is expressed in heart, brain, and testes, Merg1a' lacks the first 59 amino acids of Merg1a and is not expressed abundantly, and Merg1b has a markedly shorter divergent N-terminal cytoplasmic domain and is expressed specifically in the heart. The Merg1 isoforms, like HERG, produce inwardly rectifying E-4031-sensitive currents when heterologously expressed in Xenopus oocytes. Merg1a and HERG produce currents with slow deactivation kinetics, whereas Merg1a' and Merg1b currents deactivate more rapidly. Merg1b coassembles with Merg1a to form channels with deactivation kinetics that are more rapid than those of Merg1a or HERG and nearly identical to IKr. In addition, a homologue of Merg1b is present in human cardiac and smooth muscle. Thus, we have identified a novel N-terminal Erg isoform that is expressed specifically in the heart, has rapid deactivation kinetics, and coassembles with the longer isoform in Xenopus oocytes. This N-terminal Erg isoform may determine the properties of IKr and contribute to the pathogenesis of LQTS.
Amino Acid Sequence↗
HERG sequence correction.
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Base Sequence↗
Potassium currents expressed from Drosophila and mouse eag cDNAs in Xenopus oocytes.
The ether-a-go-go (eag) gene family encodes a set of related ion channel polypeptides expressed in the excitable cells of organisms ranging from invertebrates to mammals. Earlier studies demonstrated that eag mutations in Drosophila cause an increase in membrane excitability in the nervous system. Mutations in the human eag-related gene (HERG) have been implicated in cardiac arrhythmia, and recent studies show that HERG subunits contribute to the channels mediating IKr and the terminal repolarization of the cardiac action potential. A physiological role for M-EAG, the mouse counterpart to Drosophila eag, has not been determined. Here, we describe basic properties of Eag and M-EAG channels expressed in frog oocytes, using two-electrode voltage clamp and patch clamp techniques. Both Eag and M-EAG channels are voltage-dependent, outwardly rectifying and highly selective for K+ over Na+ over Na+ ions. In contrast to previous reports, we found no evidence for Ca2+ flux through Eag channels. The most notable difference between these closely related channels is that Eag currents exhibit partial inactivation, whereas M-EAG currents are sustained for the duration of an activating voltage command. In addition, Eag currents run down more rapidly than do M-EAG currents in excised macropatches. Rundown is reversible by inserting the patch into the interior of the oocyte, indicating that a cytosolic factor regulates channel activity or stability. These studies should facilitate identification of currents mediated by Eag and M-EAG channels in vivo.
Animals↗
HERG, a human inward rectifier in the voltage-gated potassium channel family.
In contrast to other members of the Eag family of voltage-gated, outwardly rectifying potassium channels, the human eag-related gene (HERG) has now been shown to encode an inwardly rectifying potassium channel. The properties of HERG channels are consistent with the gating properties of Eag-related and other outwardly rectifying, S4-containing potassium channels, but with the addition of an inactivation mechanism that attenuates potassium efflux during depolarization. Because mutations in HERG cause a form of long-QT syndrome, these properties of HERG channel function may be critical to the maintenance of normal cardiac rhythmicity.
Animals↗
Intra- and interexaminer reliability of the chiropractic biophysics lateral lumbar radiographic mensuration procedure.
OBJECTIVE: To determine the intra- and interexaminer reliability of a specific method of mensuration commonly used to evaluate the positional configuration of the lumbopelvic spine viewed on lateral lumbar radiographs. DESIGN: A blind, repeated-measures design was used. Lateral lumbopelvic radiographs were presented to each of three examiners in random order. Each film was marked and measurements were recorded. The films were cleaned of all markings and randomized again for a second run by each examiner. Each examiner's measurements were unavailable to the other examiners. SETTING: Private, primary-care chiropractic clinic. MAIN OUTCOME MEASURES: Anterior/posterior thoracic translation in millimeters, Ferguson's sacral-plane angle to horizontal, arcuate line angle to horizontal, L1 to L5 absolute rotation angle and four relative rotation angles for L1-L2, L2-L3, L3-L4 and L4-L5. Intra- and interrelibility of the three radiographic examiners were analyzed. RESULTS: Intraexaminer reliability for (a) L1-L5 absolute rotation angle was .98, with confidence intervals included in the range of 0.95-0.99, (b) anterior/posterior thorax translation [+/- Sz] was .97-.99, with confidence intervals included in the range of 0.94-1.00, (c) arcuate angle (AA) .40-.81, with confidence intervals included in the range of 0.07-0.90, (d) Ferguson's angle (FA) was .91-.97, with confidence intervals included in the range of 0.82-0.98, (e) relative rotation angle reliability ranges were L1-L2, .84-.94; L2-L3, .80-.85; L3-L4, .78-.89; L4-L5, .87-.92. Interexaminer reliabilities for the three examiners ranged from .66-.98. CONCLUSION: With the exception of the arcuate angle measurement, the reliabilities for all other measurements were at least .78. Those measurements with reliabilities approaching .80 or better would be considered accurate enough for use in future clinical studies. The arcuate angle measurement may have been least reliable because of the subjective nature of the method of affixing a best-fit line to a radiographic landmark that often takes on the appearance of a mild curvature. Establishing reliability is an important first step toward evaluating these and other similar radiographic measurements that have yet to be examined for their validity.
Analysis of Variance↗
Entrapment neuropathy of the palmar cutaneous nerve within its tunnel.
A case of isolated entrapment of the palmar cutaneous branch of the median nerve is presented. At operation, a ganglion compressing the nerve within its tunnel was found. Symptoms were relieved with no recurrence at 2 years after decompression and excision of the ganglion.
Carpal Tunnel Syndrome↗
Pseudo-anterior interosseous nerve syndrome: a case report.
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Adult↗
An anatomical study of the deep transverse metacarpal ligament.
The anatomy of the deep transverse metacarpal ligament was examined in cadaveric dissections. Previously unreported findings were transverse muscular slips from abductor digiti minimi that inserted into the medial border of the ligament and, in 1 case, an extension from the medial half of the tendon of extensor carpi ulnaris that inserted into the medial aspect of the ligament. Applied anatomy in relation to the dynamics of the transverse metacarpal arch is reviewed.
Finger Joint↗
A biomechanical analysis of a new interlock suture technique for flexor tendon repair.
Using a computerised tensometer, both the gap-producing and breaking forces of a new interlocking suture for flexor tendon repair were compared to the modified Kessler and the Strickland techniques. 30 porcine deep flexor tendons were used in each group and all repairs were performed with 3/0 polypropylene sutures. The interlock technique withstood gap-producing and breaking forces significantly better than the modified Kessler and Strickland techniques. Also, the gap-producing force was closer to the breaking force with the interlock technique than with the other two techniques.
Animals↗
A component of calcium-activated potassium channels encoded by the Drosophila slo locus.
Calcium-activated potassium channels mediate many biologically important functions in electrically excitable cells. Despite recent progress in the molecular analysis of voltage-activated K+ channels, Ca(2+)-activated K+ channels have not been similarly characterized. The Drosophila slowpoke (slo) locus, mutations of which specifically abolish a Ca(2+)-activated K+ current in muscles and neurons, provides an opportunity for molecular characterization of these channels. Genomic and complementary DNA clones from the slo locus were isolated and sequenced. The polypeptide predicted by slo is similar to voltage-activated K+ channel polypeptides in discrete domains known to be essential for function. Thus, these results indicate that slo encodes a structural component of Ca(2+)-activated K+ channels.
Amino Acid Sequence↗