Long QT Syndrome Mutation Research Articles

Inhibition of nonsense-mediated mRNA decay by antisense morpholino oligonucleotides restores functional expression of hERG nonsense and frameshift mutations in long-QT syndrome

Mutations in the human ether-a-go-go-related gene (hERG) cause long-QT syndrome type 2 (LQT2). We previously described a homozygous LQT2 nonsense mutation Q1070X in which the mutant mRNA is degraded by nonsense-mediated mRNA decay (NMD) leading to a severe clinical phenotype. The degradation of the Q1070X transcript precludes the expression of truncated but functional mutant channels. In the present study, we tested the hypothesis that inhibition of NMD can restore functional expression of LQT2 mutations that are targeted by NMD. We showed that inhibition of NMD by RNA interference-mediated knockdown of UPF1 increased Q1070X mutant channel protein expression and hERG current amplitude. More importantly, we found that specific inhibition of downstream intron splicing by antisense morpholino oligonucleotides prevented NMD of the Q1070X mutant mRNA and restored the expression of functional Q1070X mutant channels. The restoration of functional expression by antisense morpholino oligonucleotides was also observed in LQT2 frameshift mutations. Our findings suggest that inhibition of NMD by antisense morpholino oligonucleotides may be a potential therapeutic approach for some LQT2 patients carrying nonsense and frameshift mutations.

Transmural Differences in Myocardial Contraction in Long-QT Syndrome

Background— Long-QT syndrome (LQTS) is characterized by prolonged myocardial action potential duration. The longest action potential duration is reported in the endomyocardium and midmyocardium. Prolonged action potential duration in LQTS may cause prolonged cardiac contraction, which can be assessed by strain echocardiography. We hypothesized that myocardial contraction is most prolonged in subendocardial myofibers in LQTS patients and that inhomogeneous transmural contraction is related to the risk of spontaneous arrhythmia. Methods and Results— We included 101 genotyped LQTS mutation carriers and 35 healthy individuals. A history of cardiac arrhythmias was present in 48 mutations carriers, and 53 were asymptomatic. Myocardial contraction duration was assessed by strain echocardiography as time from the ECG Q wave to peak strain in 16 LV segments. Strain was assessed along the longitudinal axis, predominantly representing subendocardial fibers, and along the circumferential axis, representing midmyocardial fibers. Mean contraction duration was longer in LQTS mutation carriers compared with healthy individuals (445±45 versus 390±40 milliseconds; P <0.001) and longer in symptomatic compared with asymptomatic LQTS mutation carriers (460±40 versus 425±45 milliseconds; P <0.001). Contraction duration by longitudinal strain was longer than by circumferential strain in symptomatic LQTS patients (460±45 versus 445±45 milliseconds; P =0.008) but not in asymptomatic patients and healthy individuals, indicating transmural mechanical dispersion. This time difference was present in a majority of LV segments and was most evident in patients with LQT2 and the Jervell and Lange-Nielsen syndrome. Conclusion— Contraction duration in symptomatic LQTS mutation carriers was longer in the subendocardium than in the midmyocardium, indicating transmural mechanical dispersion, which was not present in asymptomatic and healthy individuals.

Clinical Summaries

Clinical Summaries

Drug-Induced Arrhythmia

Prolongation of the QTc interval has been associated with proarrhythmia resulting from a potentially fatal form of polymorphic ventricular tachycardia called torsades de pointes (TdP). Genetic forms of the long-QT syndrome (LQTS) associated with high arrhythmic risk have been causally related to mutations in ion channels responsible for the cardiac action potential; genetic factors associated with milder degrees of QTc prolongation and arrhythmic risk have also been described. Acquired forms of QTc prolongation and proarrhythmia, particularly related to drug therapy, are frequently related to drug effects on the same ion channels involved in genetic forms of LQTS. As is true for genetic forms of LQTS, there is a wide spectrum of potential drug effects on the QTc interval ranging from trivial to potentially lethal. Drug-induced QTc prolongation is a complicated phenomenon related not just to the properties or dose of a particular drug but also to drug-drug interactions and a variety of patient factors, including age, gender, the presence and severity of underlying heart disease, and genetic predisposition. Additionally, drug effects on other ion channels, including blockade of sodium channels (causing prolongation of the QRS interval rather than the QTc interval), are an important cause of drug-induced proarrhythmia. The purpose of this review is to summarize the available data related to acquired forms of LQTS, with particular focus on drug-related LQTS and proarrhythmia. A brief summary is presented here of genetic LQTS because the relevant genes and proteins form the basis for an understanding of the causes of acquired LQTS, and in any given patient, the QTc is determined by genetic, substrate, and environmental factors. Emphasis is placed on practical strategies for clinicians, with the goal of minimizing the risk of dangerous proarrhythmia related to the use of both cardiac and noncardiac drugs with the potential for QTc prolongation. Genetic …

Knock-in gain-of-function sodium channel mutation prolongs atrial action potentials and alters atrial vulnerability

Patients with long QT syndrome (LQTS) are at increased risk not only for ventricular arrhythmias but also for atrial pathology including atrial fibrillation (AF). Some patients with "lone" AF carry Na(+)-channel mutations. The purpose of this study was to determine the mechanisms underlying atrial pathology in LQTS. In mice with a heterozygous knock-in long QT syndrome type 3 (LQT3) mutant of the cardiac Na(+) channel (ΔKPQ-SCN5A) and wild-type (WT) littermates, atrial size, function, and electrophysiologic parameters were measured in intact Langendorff-perfused hearts, and histologic analysis was performed. Atrial action potential duration, effective refractory period, cycle length, and PQ interval were prolonged in ΔKPQ-SCN5A hearts (all P < .05). Flecainide (1 μM) reversed atrial action potential duration prolongation and induced postrepolarization refractoriness (P < .05). Arrhythmias were infrequent during regular rapid atrial rate in both WT and ΔKPQ-SCN5A but were inducible in 15 (38%) of 40 ΔKPQ-SCN5A and 8 (29%) of 28 WT mice upon extrastimulation. Pacing protocols generating rapid alterations in rate provoked atrial extrasystoles and arrhythmias in 6 (66%) of 9 ΔKPQ-SCN5A but in 0 (0%) of 6 WT mice (P < .05). Atrial diameter was increased by nearly 10% in ΔKPQ-SCN5A mice > 5 months old without increase in fibrotic tissue. Murine hearts bearing an LQT3 mutation show abnormalities in atrial electrophysiology and subtle changes in atrial dimension, including an atrial arrhythmogenic phenotype on provocation. These results support clinical data suggesting that LQTS mutations can cause atrial pathology and arrhythmogenesis and indicate that murine sodium channel LQTS models may be useful for exploring underlying mechanisms.

Opioid maintenance patients with QTc prolongation: Congenital long QT syndrome mutation may be a contributing risk factor

Objectives This study investigates opioid maintenance treatment (OMT) patients found to have corrected QT (QTc) interval above 500 ms, with particular focus on past medical history, genetic testing and cardiac investigations. Methods Detailed medical and cardiac history was obtained, with particular focus upon risk factors. Cardiac investigations, including genetic testing for the five most common long QT syndrome (LQTS) mutations, exercise electrocardiography (ECG) and 24-h ECG recordings, were performed. Results Of 200 OMT patients assessed with ECG, seven methadone maintained patients identified with QTc interval above 500 ms participated in this study. Two were identified as heterozygous LQTS mutation carriers. Both had experienced cardiac symptoms prior to and during OMT. No other risk factors for QTc prolongation were detected among the seven patients. Six of the seven patients underwent further cardiac investigations. QTc intervals fluctuated widely over 24 h and during exercise for all patients. Only one of the LQTS mutation carriers switched to buprenorphine and started on a beta-blocker. Despite strong medical advice and information, none of the other patients wanted to switch to buprenorphine or take other cardiac protective measures. Conclusion Findings indicate the importance of recording a thorough past medical history, focusing specifically on previous cardiac symptoms, and on other known risk factors for QTc prolongation, prior to initiating patients on methadone.

A history of stressful life events, prolonged mental stress and arrhythmic events in inherited long QT syndrome

ObjectiveTo examine whether a history of stressful life events and prolonged mental stress are associated with arrhythmic events in inherited long QT syndrome (LQTS).MethodsParticipants who had a molecularly established mutation...

Abnormal repolarization dynamics revealed in exercise test in long QT syndrome mutation carriers with normal resting QT interval

The identification of affected family members with long QT syndrome (LQTS) is often difficult due to their normal-or only marginally lengthened-QT interval duration. We examined whether physical exercise test could increase the ability to detect the mutation carrier status in phenotypically normal LQTS family members. Sixty-six subjects were included: 15 were carriers of KCNQ1 (LQT1); 15 of KCNH(2) (LQT2); and 9 of SCN5A (LQT3) gene mutations with no, or borderline, QT lengthening; and 27 were healthy controls. Multiple electrograms over the precordial area were recorded during workload and recovery phases of exercise test. QT intervals and T peak to T end intervals (Tpe intervals) were determined using an automatic algorithm at specified heart rates (HR).The LQT1 mutation carriers had QT interval most prolonged during exercise and recovery, whereas the LQT2 carriers had QT interval longest at low exercise HR. The LQT3 carriers had QT interval longest at rest. The Tpe interval remained nearly unchanged during exercise in LQT1, but shortened in LQT2 and in LQT3 carriers. The Tpe interval was longest in LQT2 carriers at the end of the recovery phase. Tentative dichotomizing values of QT and Tpe intervals improved sensitivity and specificity in distinguishing LQTS subtypes, compared with the QT interval duration alone. LQTS mutation carriers lacking diagnostic QT interval prolongation exhibit abnormal QT and Tpe interval adaptations during physical exercise test. Looking for subtype-specific adaptations might facilitate the identification of LQTS mutation carriers when molecular genetic analysis is not available.

Long QT syndrome with compound mutations is associated with a more severe phenotype: A Japanese multicenter study

Long QT syndrome (LQTS) can be caused by mutations in the cardiac ion channels. Compound mutations occur at a frequency of 4% to 11% among genotyped LQTS cases. The purpose of this study was to determine the clinical characteristics and manner of onset of cardiac events in Japanese patients with LQTS and compound mutations. Six hundred three genotyped LQTS patients (310 probands and 293 family members) were divided into two groups: those with a single mutation (n = 568) and those with two mutations (n = 35). Clinical phenotypes were compared between the two groups. Of 310 genotyped probands, 26 (8.4%) had two mutations in the same or different LQTS-related genes (compound mutations). Among the 603 LQTS patients, compound mutation carriers had significantly longer QTc interval (510 ± 56 ms vs 478± 53 ms, P = .001) and younger age at onset of cardiac events (10 ± 8 years vs 18 ± 16 years, P = .043) than did single mutation carriers. The incidence rate of cardiac events before age 40 years and use of beta-blocker therapy among compound mutation carriers also were different than in single mutation carriers. Subgroup analysis showed more cardiac events in LQTS type 1 (LQT1) and type 2 (LQT2) compound mutations compared to single LQT1 and LQT2 mutations. Compound mutation carriers are associated with a more severe phenotype than single mutation carriers.

Recent progress in congenital long QT syndrome

As genetic testing for long QT syndrome (LQTS) has become readily available, important advances are being made in understanding the exact link between ion channel mutation and observed phenotype. This paper reviews recent findings in the literature. Congenital LQTS is an important cause of sudden cardiac death. To date, 12 genes have been identified as the cause of congenital LQTS. With increasing availability of genetic testing, subtype-specific management of LQTS has become the standard of care. Detailed correlative studies between LQTS mutations and clinical phenotypes are leading the field towards 'mutation-specific' management within LQTS subtypes. A clear link between the distinct functional/biophysical defect in each LQT mutation and disease phenotype is complicated by the variable penetrance and pleiotropic expression of clinical phenotype. This is especially evident with the overlap syndrome now documented for several sodium channel (SCN5A) mutations. The management of LQTS has become subtype-specific due to the availability of genotype information. Review of recent literature suggests that 'mutation-specific' management is possible based upon distinct functional/biophysical characteristics of mutations within each LQT gene. Further research is required to clearly delineate the molecular and cellular mechanisms underlying variable penetrance, and pleiotropic expression of LQTS mutations.

Analysis of the interactions between the C-terminal cytoplasmic domains of KCNQ1 and KCNE1 channel subunits

Ion channel subunits encoded by KCNQ1 and KCNE1 produce the slowly activating K+ current (IKs) that plays a central role in myocardial repolarization. The KCNQ1 alpha-subunit and the KCNE1 beta-subunit assemble with their membrane-spanning segments interacting, resulting in transformation of channel activation kinetics. We recently reported a functional interaction involving C-terminal portions of the two subunits with ensuing regulation of channel deactivation. In the present study, we provide evidence characterizing a physical interaction between the KCNQ1-CT (KCNE1 C-terminus) and the KCNE1-CT (KCNE1 C-terminus). When expressed in cultured cells, the KCNE1-CT co-localized with KCNQ1, co-immunoprecipitated with KCNQ1 and perturbed deactivation kinetics of the KCNQ1 currents. Purified KCNQ1-CT and KCNE1-CT physically interacted in pull-down experiments, indicating a direct association. Deletion analysis of KCNQ1-CT indicated that the KCNE1-CT binds to a KCNQ1 region just after the last transmembrane segment, but N-terminal to the tetramerization domain. SPR (surface plasmon resonance) corroborated the pull-down results, showing that the most proximal region (KCNQ1 amino acids 349-438) contributed most to the bimolecular interaction with a dissociation constant of approximately 4 microM. LQT (long QT) mutants of the KCNE1-CT, D76N and W87F, retained binding to the KCNQ1-CT with comparable affinity, indicating that these disease-causing mutations do not alter channel behaviour by disruption of the association. Several LQT mutations involving the KCNQ1-CT, however, showed various effects on KCNQ1/KCNE1 association. Our results indicate that the KCNQ1-CT and the KCNE1-CT comprise an independent interaction domain that may play a role in IKs channel regulation that is potentially affected in some LQTS (LQT syndrome) mutations.

Expression of a Common LQT1 Mutation in Five Apparently Unrelated Families in a Regional Inherited Arrhythmia Clinic

The Inherited Arrhythmia Clinic at the University of Western Ontario services a catchment area of 1.5 million people and follows families with inherited arrhythmia syndromes. Patients referred for evaluation of long-QT Syndrome (LQTS) are evaluated with resting and standing ECGs, and treadmill exercise testing. Patients with findings consistent with LQTS are offered comprehensive genetic testing with screening of all first-degree relatives of genotype-positive patients. Among 31 probands with disease-causing LQTS mutations, 5 probands from apparently unrelated families of Irish descent were found to have an identical disease causing transmembrane mutation in KCNQ1 (Leu266Pro). Systematic screening of 33 first-degree relatives of genotype-positive individuals detected 15 unaffected and 18 asymptomatic affected family members. Symptoms in 6 patients occurred later in life than reported LQT1 populations (61 +/- 18 years, range 44-89). In this cohort, several family members presented with cardiac arrest during acute myocardial ischemia (n = 2), sudden death, unexplained drowning, and torsade de pointes during exercise testing. There was no identifiable common relative for this cohort after pedigree construction of the previous 4-7 generations. Affected patients had mild QT prolongation at rest with dramatic QT prolongation with exercise. Genetic testing in this LQTS population suggests a common KCNQ1 Leu266Pro founder effect, with the descendants clustering in our geographical region even though no common relative has been identified. The observations highlight the utility of genotypic and phenotypic correlation and a specialized clinic.

Utility of Treadmill Testing in Identification and Genotype Prediction in Long-QT Syndrome

The clinical diagnosis of long-QT syndrome (LQTS) remains challenging when ECG abnormalities are borderline or intermittent. Despite issues with access, cost, and heterogeneity of LQTS mutations, genetic testing remains the diagnostic gold standard for diagnosis of LQTS. We sought to develop a provocative testing strategy to unmask the LQTS phenotype and relate this to the results of genetic testing. From 1995 to 2008, 159 consecutive patients with suspected LQTS underwent provocative testing that consisted of a modified Bruce protocol treadmill exercise test, with ECGs recorded supine at rest, immediately on standing, and at 1-minute intervals during exercise, at peak exercise, and at 1-minute intervals during the recovery phase. Similar testing was carried out on a stationary bike in a gradual and burst exercise fashion. LQTS was confirmed with genotyping in all 95 affected LQTS patients and excluded with negative family screening in 64 control subjects. Patients were studied before and after initiation of beta-blockers. Of 159 patients, 50 had an LQT1 mutation and 45 had an LQT2 mutation. In the LQTS group, 44.3% of patients had a normal-to-borderline resting QTc interval. LQTS patients exhibited a greater prolongation in QTc with postural change than unaffected patients (LQT1: 40 ms [IQR, 42]; LQT2: 35 ms [IQR, 46]; and LQTS-negative: 21 ms [IQR, 37]; P=0.029). During exercise, LQT1 patients had marked QTc prolongation compared with LQT2 and LQTS-negative patients (LQT1: 65 ms [60], LQT2: 3 ms [46], LQTS negative: 5 ms [41]; P<0.0001). QT hysteresis was more pronounced in patients with LQT2 mutations compared with LQT1 and LQT-negative patients (LQT2: 40 ms [10], LQT1: 15 ms [40]; LQTS-negative: 20 ms [20]; P<0.001). beta-Blockade normalized the QTc changes seen with standing and QT hysteresis. The presence and genotype of LQTS can be predicted by a combination of postural and exercise changes in the QT/RR relationship. beta-Blockade normalized these changes. Routine exercise testing is useful in predicting and directing genetic testing in LQTS.

Posthumous diagnosis of long QT syndrome from neonatal screening cards

Molecular autopsy in sudden unexplained death in the young (SUDY) victims cannot usually be performed if tissue suitable for DNA extraction is not retained at autopsy. The purpose of this study was to assess the feasibility and clinical value of posthumous genetic testing for long QT syndrome (LQTS) using residual material from the neonatal screening (Guthrie) card in SUDY victims. Twenty-one cases were investigated up to 13 years after death. Deaths occurred at <1 year in one, 1-18 years in 18, and 19-35 years in two patients. Guthrie cards were 3-39 years old. DNA was extracted, and amplicons corresponding to the coding regions of the LQTS genes 1, 2, 3, 5, and 6 underwent either denaturing high-performance liquid chromatography screening or direct DNA sequencing. Adequate DNA was extracted in every case, although repeated purification and amplification was often required. Rare variants were detected in six of 19 cases undergoing diagnostic screening. Four (21%) are considered to be pathological and have been used for family screening: R243C and H455Y in KCNQ1 in 12-year-old and 13-year-old boys, respectively, and Q81H and S621R in KCNH2 in 21-month and 28-year-old females, respectively. Variants of uncertain significance were R1047L in KCNH2 in a 2-year-old girl and S38G in KCNE1 in a 19-month-old boy. Point mutation tests for previously identified familial LQTS mutations revealed a positive result in both cases: E146K in KCNQ1 and exon 6-4del in KCNH2. Residual material from Guthrie cards collected for newborn metabolic screening can be used as a reliable source of DNA for the posthumous diagnosis of LQTS decades after SUDY, although purification and amplification of DNA is time intensive.

Two automatic QT algorithms compared with manual measurement in identification of long QT syndrome

Background Long QT syndrome (LQTS) is an inherited disorder that increases the risk of syncope and malignant ventricular arrhythmias, which may result in sudden death. Methods We compared manual measurement by 4 observers (QT manual) and 3 computerized measurements for QT interval accuracy in the diagnosis of LQTS: 1. QT measured from the vector magnitude calculated from the 3 averaged orthogonal leads X, Y, and Z (QT VCG) and classified using the same predefined QTc cut-points for classification of QT prolongation as in manual measurements; 2. QT measured by a 12-lead electrocardiogram (ECG) program (QT ECG) and subsequently classified using the same cut-points as in (1) above; 3. The same QT value as in (2) above, automatically classified by a 12-lead ECG program with thresholds for QT prolongation adjusted for age and sex (QT interpret). The population consisted of 94 genetically confirmed carriers of KCNQ1 (LQT1) and KCNH2 (LQT2) mutations and a combined control group of 28 genetically confirmed noncarriers and 66 unrelated healthy volunteers. Results QT VCG provided the best combination of sensitivity (89%) and specificity (90%) in diagnosing LQTS, with 0.948 as the area under the receiver operating characteristic curve. The evaluation of QT measurement by the 4 observers revealed a high interreader variability, and only 1 of 4 observers showed acceptable level of agreement in LQTS mutation carrier identification ( κ coefficient >0.75). Conclusion Automatic QT measurement by the Mida1000/CoroNet system (Ortivus AB, Danderyd, Sweden) is an accurate, efficient, and easily applied method for initial screening for LQTS.

Abnormal left ventricular relaxation in patients with long QT syndrome

We read with great interest the recent report by Haugaa et al. 1 and the accompanying editorial by De Ferrari and Schwartz2 on the association between abnormal ventricular repolarization and mechanical dysfunction (prolonged contraction and/or impaired diastolic function) in patients with long QT syndrome (LQTS). We were particularly interested by the statement of Haugaa et al. 1 that 'these findings imply an impairment of diastolic function in a number of symptomatic LQTS mutation carriers'. In keeping with this view, Moss et al. 3 also linked the prolonged ventricular repolarization in LQT3 patients ( SCN5A- ΔKPQ mutation) with slowed left ventricular (LV) relaxation.3 In these patients, the …

LQTS mutation N1325S in cardiac sodium channel gene SCN5A causes cardiomyocyte apoptosis, cardiac fibrosis and contractile dysfunction in mice

Mutations in the cardiac sodium channel gene SCN5A cause long QT syndrome (LQTS). We previously generated an LQTS mouse model (TG-NS) that overexpresses the LQTS mutation N1325S in SCN5A. The TG-NS mice manifested the clinical features of LQTS including spontaneous VT, syncope and sudden death. However, the long-term prognosis of LQTS on the structure of the heart has not been investigated in this or any other LQTS models and human patients. Impaired systolic function and reduced left ventricular fractional shortening were detected by echocardiography, morphological and histological examination in two lines of adult mutant transgenic mice. Histological and TUNEL analyses of heart sections revealed fibrosis lesions and increased apoptosis in an age-dependent manner. Cardiomyocyte apoptosis was associated with the increased activation of caspases 3 and 9 in TG-NS hearts. Western blot analysis showed a significantly increased expression of the key Ca(2+) handling proteins L-type Ca(2+) channel, RYR2 and NCX in TG-NS hearts. Increased apoptosis and an altered expression of Ca(2+) handling proteins could be detected as early as 3months of age when echocardiography showed little or no alterations in TG-NS mice. Our findings revealed for the first time that the LQTS mutation N1325S in SCN5A causes cardiac fibrosis and contractile dysfunction in mice, possibly through cellular mechanisms involving aberrant cardiomyocyte apoptosis. Therefore, we provide the experimental evidence supporting the notion that some LQTS patients have an increased risk of structural and functional cardiac damage in a prolonged disease course.

Trafficking-deficient long QT syndrome mutation KCNQ1-T587M confers severe clinical phenotype by impairment of KCNH2 membrane localization: Evidence for clinically significant I Kr-I Ks α-subunit interaction

KCNQ1-T587M is a trafficking-deficient long QT syndrome (LQTS) missense mutation. Affected patients exhibit severe clinical phenotypes that are not explained by the mutant's effects on I(Ks). Previous work showed a KCNH2 and KCNQ1 alpha-subunit interaction that increases KCNH2 membrane localization and function. We hypothesized that failure of trafficking-deficient KCNQ1-T587M to enhance KCNH2 membrane expression could reduce KCNH2 current versus wild-type KCNQ1 (KCNQ1-WT), contributing to the LQTS phenotype of KCNQ1-T587M carriers. Patch-clamp, protein biochemical studies, confocal imaging, and in vivo transfection of guinea pig cardiomyocytes were performed. KCNQ1-T587M failed to generate functional current when coexpressed with KCNE1 and caused haploinsufficiency when coexpressed with KCNQ1-WT/KCNE1. Coexpression of KCNQ1-WT with KCNH2 increased I(KCNH2) versus KCNH2 alone (P <.05). Immunoblots and confocal microscopy indicated increased plasma membrane localization of KCNH2 alpha-subunits in cells cotransfected with KCNQ1-WT plasmid, while total KCNH2 protein synthesis and KCNH2 glycosylation remained unaffected, which suggests a chaperone effect of KCNQ1-WT to enhance the membrane localization of KCNH2. KCNH2 also coimmunoprecipitated with KCNQ1-WT. Although KCNQ1-T587M coprecipitated with KCNH2, the mutant was retained intracellularly and failed to increase KCNH2 membrane localization, abolishing the KCNQ1-WT chaperone function and reducing I(KCNH2) upon coexpression substantially compared with coexpression with KCNQ1-WT (P <.05). In vivo transfection of KCNQ1-T587M in guinea pigs suppressed I(Kr) in isolated cardiomyocytes. The trafficking-deficient LQTS mutation KCNQ1-T587M fails to show the chaperoning function that enhances KCNH2 membrane localization with KCNQ1-WT. This novel mechanism results in reduced I(KCNH2), which would be expected to decrease repolarization reserve and synergize with reduced I(KCNQ1) caused directly by the mutation, potentially explaining the malignant clinical phenotype in affected patients.

A recombinant N-terminal domain fully restores deactivation gating in N-truncated and long QT syndrome mutant hERG potassium channels

Human ether á go-go related gene (hERG) potassium channels play a central role in cardiac repolarization where channel closing (deactivation) regulates current density during action potentials. Consequently, mutations in hERG that perturb deactivation are linked to long QT syndrome (LQTS), a catastrophic cardiac arrhythmia. Interactions between an N-terminal domain and the pore-forming "core" of the channel were proposed to regulate deactivation, however, despite its central importance the mechanistic basis for deactivation is unclear. Here, to more directly examine the mechanism for regulation of deactivation, we genetically fused N-terminal domains to fluorescent proteins and tested channel function with electrophysiology and protein interactions with Förster resonance energy transfer (FRET) spectroscopy. Truncation of hERG N-terminal regions markedly sped deactivation, and here we report that reapplication of gene fragments encoding N-terminal residues 1-135 (the "eag domain") was sufficient to restore regulation of deactivation. We show that fluorophore-tagged eag domains and N-truncated channels were in close proximity at the plasma membrane as determined with FRET. The eag domains with Y43A or R56Q (a LQTS locus) mutations showed less regulation of deactivation and less FRET, whereas eag domains restored regulation of deactivation gating to full-length Y43A or R56Q channels and showed FRET. This study demonstrates that direct, noncovalent interactions between the eag domain and the channel core were sufficient to regulate deactivation gating, that an LQTS mutation perturbed physical interactions between the eag domain and the channel, and that small molecules such as the eag domain represent a novel method for restoring function to channels with disease-causing mutations.

State of Postmortem Genetic Testing Known as the Cardiac Channel Molecular Autopsy in the Forensic Evaluation of Unexplained Sudden Cardiac Death in the Young

Thousands of infants, children, adolescents, and young adults die sudden and unexpectedly each year in the United States. A significant proportion are autopsy negative and are classified as autopsy negative sudden unexplained death (SUD) after the first year of life and as sudden infant death syndrome (SIDS) if prior to their first birthday. Postmortem genetic testing known as the cardiac channel molecular autopsy is capable of identifying the subset of channelopathic SUD/SIDS. Review of the literature and analysis of the state of such postmortem genetic testing in the evaluation of SUD/SIDS. Although still confined to anecdotal reports, relatively small case series of coroner/medical examiner-referred cases of SUD/SIDS, and one population-based cohort of SIDS, it is estimated that approximately 25-35% of autopsy-negative SUD and approximately 10% of SIDS may stem from mutations in either long QT syndrome (LQTS)- or catecholaminergic polymorphic ventricular tachycardia (CPVT)-susceptibility genes. Whether the cardiac channel molecular autopsy should become the standard of care in the postmortem evaluation of autopsy negative SUD or SIDS will require further scrutiny. Cost effectiveness analyses of a more intense postmortem focus on the decedent compared to the current battery of tests recommended for the deceased SUD victim's first degree relatives should be performed. If deemed justified to upgrade such postmortem genetic testing from "investigational" to clinically indicated, uniform "standard operating procedures" to ensure that tissue is acquired and archived in a manner that is "DNA friendly" and insurance coverage that extends beyond one's final breath will be needed.