Stallmeyer B.,Heart Genetics
Circulation Research | Year: 2017
RATIONALE:: Familial sinus node and atrioventricular (AV) conduction dysfunction is a rare disorder that leads to paroxysmal dizziness, fatigue and syncope due to a temporarily or permanent reduced heart rate. To date, only a few genes for familial sinus and/or AV conduction dysfunction are known and the majority of cases remain etiologically unresolved. OBJECTIVE:: We aim to identify the disease gene in a large three-generation family (n=25) with autosomal dominant sinus node dysfunction (SND) and AV block (AVB) and to characterise the mutation-related pathomechanisms in familial SND+AVB. METHODS AND RESULTS:: Genome-wide linkage analysis mapped the SND+AVB disease locus to chromosome 7q21.1-q31.1 (2-point LOD score: 4.64; θ=0); in this region targeted exome sequencing identified a novel heterozygous mutation (p.Arg52Leu) in the GNB2 gene that strictly co-segregated with the SND+AVB phenotype. GNB2 encodes the β2-subunit (Gβ2) of the heterotrimeric G-protein complex that is being released from G-protein-coupled receptors (GPCRs) upon vagal stimulation. In two heterologous expression systems (HEK-293T cells and Xenopus laevis oocytes) an enhanced activation of the G-protein activated K channel (GIRK; Kir3.1/Kir3.4) was shown when mutant Gβ2was co-expressed with Gγ2; this was in contrast to co-expression of mutant Gβ2-Gγ2 with other cardiac ion channels (HCN4, HCN2, Cav1.2). Molecular dynamics simulations suggested a reduced binding property of mutant Gβ2 to cardiac GIRK channels when compared to native Gβ2. CONCLUSIONS:: A GNB2 gene mutation is associated with familial SND+AVB and leads to a sustained activation of cardiac GIRK channels which is likely to hyperpolarize the myocellular membrane potential and thus reduces their spontaneous activity. Our findings describe for the first time a role of a mutant G-protein in the non-syndromic pacemaker disease due to GIRK channel activation. © 2017 American Heart Association, Inc.
Vincentz J.W.,Heart Genetics |
vanDusen N.J.,Heart Genetics |
Fleming A.B.,Heart Genetics |
Rubart M.,Heart Genetics |
And 3 more authors.
Journal of Neuroscience | Year: 2012
Neural crest cell specification and differentiation to a sympathetic neuronal fate serves as an important model for neurogenesis and depends upon the function of both bHLH transcription factors, notably Hand2, and homeodomain transcription factors, including Phox2b. Here, we define a 1007 bp cis-regulatory element 5′ of the Hand1 gene sufficient to drive reporter expression within the sympathetic chain of transgenic mice. Comparative genomic analyses uncovered evolutionarily conserved consensus-binding sites within this element, which chromatin immunoprecipitation and electrophoretic mobility shift assays confirm are bound by Hand2 and Phox2b. Mutational analyses revealed that the conserved Phox2 and E-box binding sites are necessary for proper cis-regulatory element activity, and expression analyses on both Hand2 conditionally null and hypomorphic backgrounds demonstrate that Hand2 is required for reporter activation in a gene dosage-dependent manner.Wedemonstrate that Hand2 and Hand1 differentially bind the E-boxes in this cis-regulatory element, establishing molecular differences between these two factors. Finally, we demonstrate that Hand1 is dispensable for normal tyrosine hydroxylase (TH) and dopamine β-hydroxylase (DBH) expression in sympathetic neurons, even when Hand2 gene dosage is concurrently reduced by half. Together, these data define a tissue-specific Hand1 cis-regulatory element controlled by two factors essential for the development of the sympathetic nervous system and provide in vivo regulatory evidence to support previous findings that Hand2, rather than Hand1, is predominantly responsible for regulating TH, DBH, and Hand1 expression in developing sympathetic neurons. © 2012 the authors.
Park S.,Heart Genetics |
Terzic A.,Heart Genetics
Journal of Structural Biology | Year: 2010
Heterodimeric nucleotide binding domains NBD1/NBD2 distinguish the ATP-binding cassette protein SUR2A, a recognized regulatory subunit of cardiac ATP-sensitive K+ (KATP) channels. The tandem function of these core domains ensures metabolism-dependent gating of the Kir6.2 channel pore, yet their structural arrangement has not been resolved. Here, purified monodisperse and interference-free recombinant particles were subjected to synchrotron radiation small-angle X-ray scattering (SAXS) in solution. Intensity function analysis of SAXS profiles resolved NBD1 and NBD2 as octamers. Implemented by ab initio simulated annealing, shape determination prioritized an oblong envelope wrapping NBD1 and NBD2 with respective dimensions of 168 × 80 × 37 Å3 and 175 × 81 × 37 Å3 based on symmetry constraints, validated by atomic force microscopy. Docking crystal structure homology models against SAXS data reconstructed the NBD ensemble surrounding an inner cleft suitable for Kir6.2 insertion. Human heart disease-associated mutations introduced in silico verified the criticality of the mapped protein-protein interface. The resolved quaternary structure delineates thereby a macromolecular arrangement of KATP channel SUR2A regulatory domains. © 2009 Elsevier Inc. All rights reserved.
VanDusen N.J.,Heart Genetics |
Firulli A.B.,Heart Genetics
Differentiation | Year: 2012
The heart is a complex organ that is composed of numerous cell types, which must integrate their programs for proper specification, differentiation and cardiac morphogenesis. During cardiogenesis members of the Twist-family of basic helix-loop-helix (bHLH) transcription factors play distinct roles within cardiac lineages such as the endocardium and extra-cardiac lineages such as the cardiac neural crest (cNCC) and epicardium. While the study of these cell populations is often eclipsed by that of cardiomyocytes, the contributions of non-cardiomyocytes to development and disease are increasingly being appreciated as both dynamic and essential. This review summarizes what is known regarding Twist-family bHLH function in extra-cardiac cell populations and the endocardium, with a focus on regulatory mechanisms, downstream targets, and expression profiles. Improving our understanding of the molecular pathways that Twist-family bHLH factors mediate in these lineages will be necessary to ascertain how their dysfunction leads to congenital disease and adult pathologies such as myocardial infarctions and cardiac fibroblast induced fibrosis. Indeed, this knowledge will prove to be critical to clinicians seeking to improve current treatments. © 2012 International Society of Differentiation.
Chung S.,Heart Genetics |
Arrell D.K.,Heart Genetics |
Faustino R.S.,Heart Genetics |
Terzic A.,Heart Genetics |
Dzeja P.P.,Heart Genetics
Journal of Molecular and Cellular Cardiology | Year: 2010
Decoding of the bioenergetic signature underlying embryonic stem cell cardiac differentiation has revealed a mandatory transformation of the metabolic infrastructure with prominent mitochondrial network expansion and a distinctive switch from glycolysis to oxidative phosphorylation. Here, we demonstrate that despite reduction in total glycolytic capacity, stem cell cardiogenesis engages a significant transcriptome, proteome, as well as enzymatic and topological rearrangement in the proximal, medial, and distal modules of the glycolytic pathway. Glycolytic restructuring was manifested by a shift in hexokinase (Hk) isoforms from Hk-2 to cardiac Hk-1, with intracellular and intermyofibrillar localization mapping mitochondrial network arrangement. Moreover, upregulation of cardiac-specific enolase 3, phosphofructokinase, and phosphoglucomutase and a marked increase in glyceraldehyde 3-phosphate dehydrogenase (GAPDH) phosphotransfer activity, along with apparent post-translational modifications of GAPDH and phosphoglycerate kinase, were all distinctive for derived cardiomyocytes compared to the embryonic stem cell source. Lactate dehydrogenase (LDH) isoforms evolved towards LDH-2 and LDH-3, containing higher proportions of heart-specific subunits, and pyruvate dehydrogenase isoforms rearranged between E1α and E1β, transitions favorable for substrate oxidation in mitochondria. Concomitantly, transcript levels of fetal pyruvate kinase isoform M2, aldolase 3, and transketolase, which shunt the glycolytic with pentose phosphate pathways, were reduced. Collectively, changes in glycolytic pathway modules indicate active redeployment, which would facilitate connectivity of the expanding mitochondrial network with ATP utilization sites. Thus, the delineated developmental dynamics of the glycolytic phosphotransfer network is integral to the remodeling of cellular energetic infrastructure underlying stem cell cardiogenesis. © 2009 Elsevier Ltd. All rights reserved.
Dmitrieva R.I.,Institute of Hematology |
Minullina R.,Heart Genetics |
Bilibina A.A.,Heart Genetics |
Tarasova O.V.,Heart Genetics |
And 2 more authors.
Cell Cycle | Year: 2012
Bone marrow (BM) and subcutaneous adipose tissue (Ad) are both considered prospective sources of MSC for therapeutic applications. However, functional properties and therapeutic efficacy of MSC derived from different tissues of the same patient are still poorly investigated. In our study, BM-MSC and F-MSC cultures from 43 adult donors were evaluated in successive passages for immunophenotype; secretion of VEGF, SDF1, MCP1, IL6 and TGFβ1; frequency of colony-forming units (CFU-F); frequency of adipo- and osteoprogenitors (CFU-Ad, CFU-Ost) and for onset of in vitro replicative senescence. We have demonstrated that at early passages (P2-P3 or up to 14-15 in vitro population doublings), BM- and Ad-derived MSC cultures are comparable in such important characteristics as proliferation rate (population-doubling time: 3.4 ± 0.2% in BM-MSC, 3 ± 0.3% in F-MSC), clonogenity (CFU-F frequency: 32 ± 5% in BM-MSC, 31 ± 5% in F-MSC) and differentiation potential (CFU-Ad frequency: 10.4 ± 2% in BM-MSC, 13 ± 3% in F-MSC; CFU-Ost frequency: 18.5 ± 5.5% in BM-MSC, 18 ± 5% in F-MSC) but differ significantly in abundance of CD146 + fraction within the sample (25 ± 5% in BMMSC, 7 ± 3% in F-MSC) and in levels of VEGF, SDF-1, MCP1 and TGFβ1 secretion. We have also demonstrated that BM?MSC enter senescence after P3-4, while most F-MSC did not show senescence features up to P6-8. Together, these data demonstrate that specific properties of MSC from different sources should be always taken into account when developing and optimizing the specific protocols for MSC expansion and evaluation for each particular clinical application. © 2012 Landes Bioscience.
Arrell D.K.,Marriott Heart Disease Research Program |
Arrell D.K.,Molecular Therapeutics |
Arrell D.K.,Heart Genetics |
Terzic A.,Marriott Heart Disease Research Program |
And 2 more authors.
Clinical Pharmacology and Therapeutics | Year: 2010
Systems biology provides a platform for integrating multiple components and interactions underlying cell, organ, and organism processes in health and disease. Beyond traditional approaches focused on individual molecules or pathways, bioinformatic network analysis of high-throughput data sets offers an opportunity for integration of biological complexity and multilevel connectivity. Emerging applications in rational drug discovery range from targeting and modeling disease-corrupted networks to screening chemical or ligand libraries to identification/validation of drug-target interactions for improved efficacy and safety. © 2010 American Society for Clinical Pharmacology and Therapeutics.
Vincentz J.W.,Heart Genetics |
Barnes R.M.,Heart Genetics |
Firulli A.B.,Heart Genetics
Birth Defects Research Part A - Clinical and Molecular Teratology | Year: 2011
Almost 15 years of careful study have established the related basic Helix-Loop-Helix (bHLH) transcription factors Hand1 and Hand2 as critical for heart development across evolution. Hand factors make broad contributions, revealed through animal models, to the development of multiple cellular lineages that ultimately contribute to the heart. They perform critical roles in ventricular cardiomyocyte growth, differentiation, morphogenesis, and conduction. They are also important for the proper development of the cardiac outflow tract, epicardium, and endocardium. Molecularly, they function both through DNA binding and through protein-protein interactions, which are regulated transcriptionally, posttranscriptionally by microRNAs, and posttranslationally through phosphoregulation. Although direct Hand factor transcriptional targets are progressively being identified, confirmed direct targets of Hand factor transcriptional activity in the heart are limited. Identification of these targets will be critical to model the mechanisms by which Hand factor bHLH interactions affect developmental pathways. Improved understanding of Hand factor-mediated transcriptional cascades will be necessary to determine how Hand factor dysregulation translates to human disease phenotypes. This review summarizes the insight that animal models have provided into the regulation and function of these factors during heart development, in addition to the recent findings that suggest roles for HAND1 and HAND2 in human congenital heart disease. © 2011 Wiley-Liss, Inc.
Pereira T.V.,University of Ioannina |
Pereira T.V.,Heart Genetics |
Ioannidis J.P.A.,University of Ioannina |
Ioannidis J.P.A.,Stanford University |
Ioannidis J.P.A.,Tufts University
Journal of Clinical Epidemiology | Year: 2011
Objective: To assess whether nominally statistically significant effects in meta-analyses of clinical trials are true and whether their magnitude is inflated. Study Design and Setting: Data from the Cochrane Database of Systematic Reviews 2005 (issue 4) and 2010 (issue 1) were used. We considered meta-analyses with binary outcomes and four or more trials in 2005 with P < 0.05 for the random-effects odds ratio (OR). We examined whether any of these meta-analyses had updated counterparts in 2010. We estimated the credibility (true-positive probability) under different prior assumptions and inflation in OR estimates in 2005. Results: Four hundred sixty-one meta-analyses in 2005 were eligible, and 80 had additional trials included by 2010. The effect sizes (ORs) were smaller in the updating data (2005-2010) than in the respective meta-analyses in 2005 (median 0.85-fold, interquartile range [IQR]: 0.66-1.06), even more prominently for meta-analyses with less than 300 events in 2005 (median 0.67-fold, IQR: 0.54-0.96). Mean credibility of the 461 meta-analyses in 2005 was 63-84% depending on the assumptions made. Credibility estimates changed >20% in 19-31 (24-39%) of the 80 updated meta-analyses. Conclusions: Most meta-analyses with nominally significant results pertain to truly nonnull effects, but exceptions are not uncommon. The magnitude of observed effects, especially in meta-analyses with limited evidence, is often inflated. © 2011 Elsevier Inc. All rights reserved.