Imperial College London

DrLeanneFelkin

Faculty of MedicineNational Heart & Lung Institute

Healthcare Senior Teaching Fellow
 
 
 
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Contact

 

+44 (0)20 7594 8582l.felkin

 
 
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Location

 

G218Guy Scadding BuildingRoyal Brompton Campus

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Summary

 

Publications

Publication Type
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89 results found

Reichart D, Lindberg EL, Maatz H, Miranda AMA, Viveiros A, Shvetsov N, Gaertner A, Nadelmann ER, Lee M, Kanemaru K, Ruiz-Orera J, Strohmenger V, DeLaughter DM, Patone G, Zhang H, Woehler A, Lippert C, Kim Y, Adami E, Gorham JM, Barnett SN, Brown K, Buchan RJ, Chowdhury RA, Constantinou C, Cranley J, Felkin LE, Fox H, Ghauri A, Gummert J, Kanda M, Li R, Mach L, McDonough B, Samari S, Shahriaran F, Yapp C, Stanasiuk C, Theotokis P, Theis FJ, van den Bogaerdt A, Wakimoto H, Ware JS, Worth CL, Barton PJR, Lee Y-A, Teichmann SA, Milting H, Noseda M, Oudit GY, Heinig M, Seidman JG, Hubner N, Seidman CEet al., 2022, Pathogenic variants damage cell composition and single-cell transcription in cardiomyopathies, Science, Vol: 377, Pages: 1-13, ISSN: 0036-8075

INTRODUCTIONHuman heart failure is a highly morbid condition that affects 23 million individuals worldwide. It emerges in the setting of an array of different cardiovascular disorders, which has propelled the notion that diverse stimuli converge on a common final pathway. Consistent with this, initiating etiologies do not direct heart failure treatments, which are often inadequate and necessitate mechanical interventions and cardiac transplantation.The recent application of single-nucleus RNA sequencing (snRNAseq) transcriptional analyses to characterize the cellular composition and molecular states in the healthy adult human heart provides an emerging benchmark by which disease-related changes can be assessed. Moreover, the discovery of human pathogenic variants that cause dilated cardiomyopathy (DCM) and arrhythmogenic cardiomyopathy (ACM), disorders associated with high rates of heart failure, provides direct opportunities to evaluate whether genotype influences heart failure pathways.RATIONALEA systematic identification of shared and distinct molecules and pathways involved in heart failure is lacking, and knowledge of these fundamental data could propel the development of more effective treatments. To enable these discoveries, we performed snRNAseq of explanted ventricular tissues from 18 healthy donors and 61 heart failure patients. By focusing analyses on multiple samples with pathogenic variants in DCM genes (LMNA, RBM20, and TTN), ACM genes (PKP2), or pathogenic variant–negative (PV negative) samples, we characterized genotype-stratified and common heart failure responses.RESULTSFrom 881,081 nuclei isolated from left and right diseased and healthy ventricles, we identified 10 major cell types and 71 distinct transcriptional states. DCM and ACM tissues showed significant depletion of cardiomyocytes and increased endothelial and immune cells. Fibrosis was expanded in disease hearts, but, unexpectedly, fibroblasts were not increased, and instead showed a

Journal article

Clerk A, Meijles DN, Hardyman MA, Fuller SJ, Chothani SP, Cull JJ, Cooper STE, Alharbi HO, Vanezis K, Felkin LE, Markou T, Leonard SJ, Shaw SW, Rackham OJL, Cook SA, Glennon PE, Sheppard MN, Sembrat JC, Rojas M, McTiernan CF, Barton PJ, Sugden PHet al., 2022, Cardiomyocyte BRAF and type 1 RAF inhibitors promote cardiomyocyte and cardiac hypertrophy in mice in vivo, Biochemical Journal, Vol: 479, Pages: 401-424, ISSN: 0264-6021

The extracellular signal-regulated kinase 1/2 (ERK1/2) cascade promotes cardiomyocyte hypertrophy and is cardioprotective, with the three RAF kinases forming a node for signal integration. Our aims were to determine if BRAF is relevant for human heart failure, whether BRAF promotes cardiomyocyte hypertrophy, and if Type 1 RAF inhibitors developed for cancer (that paradoxically activate ERK1/2 at low concentrations: the 'RAF paradox') may have the same effect. BRAF was up-regulated in heart samples from patients with heart failure compared with normal controls. We assessed the effects of activated BRAF in the heart using mice with tamoxifen-activated Cre for cardiomyocyte-specific knock-in of the activating V600E mutation into the endogenous gene. We used echocardiography to measure cardiac dimensions/function. Cardiomyocyte BRAFV600E induced cardiac hypertrophy within 10 d, resulting in increased ejection fraction and fractional shortening over 6 weeks. This was associated with increased cardiomyocyte size without significant fibrosis, consistent with compensated hypertrophy. The experimental Type 1 RAF inhibitor, SB590885, and/or encorafenib (a RAF inhibitor used clinically) increased ERK1/2 phosphorylation in cardiomyocytes, and promoted hypertrophy, consistent with a 'RAF paradox' effect. Both promoted cardiac hypertrophy in mouse hearts in vivo, with increased cardiomyocyte size and no overt fibrosis. In conclusion, BRAF potentially plays an important role in human failing hearts, activation of BRAF is sufficient to induce hypertrophy, and Type 1 RAF inhibitors promote hypertrophy via the 'RAF paradox'. Cardiac hypertrophy resulting from these interventions was not associated with pathological features, suggesting that Type 1 RAF inhibitors may be useful to boost cardiomyocyte function.

Journal article

Widjaja AA, Dong J, Adami E, Viswanathan S, Ng B, Pakkiri LS, Chothani SP, Singh BK, Lim WW, Zhou J, Shekeran SG, Tan J, Lim SY, Goh J, Wang M, Holgate R, Hearn A, Felkin LE, Yen PM, Dear JW, Drum CL, Schafer S, Cook SAet al., 2021, Redefining IL11 as a regeneration-limiting hepatotoxin and therapeutic target in acetaminophen-induced liver injury, SCIENCE TRANSLATIONAL MEDICINE, Vol: 13, ISSN: 1946-6234

Journal article

Mazzarotto F, Tayal U, Buchan RJ, Midwinter W, Wilk A, Whiffin N, Govind R, Mazaika E, de Marvao A, Dawes T, Felkin LE, Ahmad M, Theotokis PI, Edwards E, Ing AI, Thomson KL, Chan LLH, Sim D, Baksi AJ, Pantazis A, Roberts AM, Watkins H, Funke B, O'Regan D, Olivotto I, Barton PJR, Prasad SK, Cook SA, Ware JS, Walsh Ret al., 2020, Re-evaluating the genetic contribution of monogenic dilated cardiomyopathy, Circulation, Vol: 141, Pages: 387-398, ISSN: 0009-7322

Background: Dilated cardiomyopathy (DCM) is genetically heterogeneous, with >100 purported disease genes tested in clinical laboratories. However, many genes were originally identified based on candidate-gene studies that did not adequately account for background population variation. Here we define the frequency of rare variation in 2538 DCM patients across protein-coding regions of 56 commonly tested genes and compare this to both 912 confirmed healthy controls and a reference population of 60,706 individuals in order to identify clinically interpretable genes robustly associated with dominant monogenic DCM.Methods: We used the TruSight Cardio sequencing panel to evaluate the burden of rare variants in 56 putative DCM genes in 1040 DCM patients and 912 healthy volunteers processed with identical sequencing and bioinformatics pipelines. We further aggregated data from 1498 DCM patients sequenced in diagnostic laboratories and the ExAC database for replication and meta-analysis.Results: Truncating variants in TTN and DSP were associated with DCM in all comparisons. Variants in MYH7, LMNA, BAG3, TNNT2, TNNC1, PLN, ACTC1, NEXN, TPM1 and VCL were significantly enriched in specific patient subsets, with the last 2 genes potentially contributing primarily to early-onset forms of DCM. Overall, rare variants in these 12 genes potentially explained 17% of cases in the outpatient clinic cohort representing a broad range of adult DCM patients and 26% of cases in the diagnostic referral cohort enriched in familial and early-onset DCM. Whilst the absence of a significant excess in other genes cannot preclude a limited role in disease, such genes have limited diagnostic value since novel variants will be uninterpretable and their diagnostic yield is minimal.Conclusion: In the largest sequenced DCM cohort yet described, we observe robust disease association with 12 genes, highlighting their importance in DCM and translating into high interpretability in diagnostic testing. The

Journal article

Walsh R, Mazzarotto F, Whiffin N, Buchan R, Li N, Felkin L, Thomson KL, Watkins H, Barton PJR, Olivotto I, Cook SA, Bezzina CR, Ware JSet al., 2019, Quantitative approaches to variant classification increase the yield and precision of genetic testing in Mendelian diseases, 52nd Conference of the European-Society-of-Human-Genetics (ESHG), Publisher: NATURE PUBLISHING GROUP, Pages: 1720-1720, ISSN: 1018-4813

Conference paper

Chothani S, Schäfer S, Adami E, Viswanathan S, Widjaja AA, Langley SR, Tan J, Wang M, Quaife NM, Pua CJ, D'Agostino G, Shekeran SG, George BL, Lim S, Cao EY, van Heesch S, Witte F, Felkin LE, Christodoulou EG, Dong J, Blachut S, Patone G, Barton PJR, Hubner N, Cook SA, Rackham OJLet al., 2019, Widespread translational control of fibrosis in the human heart by RNA-binding proteins, Circulation, Vol: 140, Pages: 937-951, ISSN: 0009-7322

BACKGROUND: Fibrosis is a common pathology in many cardiac disorders and is driven by the activation of resident fibroblasts. The global post-transcriptional mechanisms underlying fibroblast-to-myofibroblast conversion in the heart have not been explored. METHODS: Genome-wide changes of RNA transcription and translation during human cardiac fibroblast activation were monitored with RNA sequencing and ribosome profiling. We then used an RNA-binding protein-based analyses to identify translational regulators of fibrogenic genes. The integration with cardiac ribosome occupancy levels of 30 dilated cardiomyopathy patients demonstrates that these post-transcriptional mechanisms are also active in the diseased fibrotic human heart. RESULTS: We generated nucleotide-resolution translatome data during the TGFβ1-driven cellular transition of human cardiac fibroblasts to myofibroblasts. This identified dynamic changes of RNA transcription and translation at several time points during the fibrotic response, revealing transient and early-responder genes. Remarkably, about one-third of all changes in gene expression in activated fibroblasts are subject to translational regulation and dynamic variation in ribosome occupancy affects protein abundance independent of RNA levels. Targets of RNA-binding proteins were strongly enriched in post-transcriptionally regulated genes, suggesting genes such as MBNL2 can act as translational activators or repressors. Ribosome occupancy in the hearts of patients with dilated cardiomyopathy suggested the same post-transcriptional regulatory network was underlying cardiac fibrosis. Key network hubs include RNA-binding proteins such as PUM2 and QKI that work in concert to regulate the translation of target transcripts in human diseased hearts. Furthermore, silencing of both PUM2 and QKI inhibits the transition of fibroblasts toward pro-fibrotic myofibroblasts in response to TGFβ1. CONCLUSIONS: We reveal widespread translational effects of

Journal article

Chen H, Moreno-Moral A, Pesce F, Devapragash N, Mancini M, Heng EL, Rotival M, Srivastava PK, Harmston N, Shkura K, Rackham OJL, Yu W-P, Sun X-M, Tee NGZ, Tan ELS, Barton PJR, Felkin LE, Lara-Pezzi E, Angelini G, Beltrami C, Pravenec M, Schafer S, Bottolo L, Hubner N, Emanueli C, Cook SA, Petretto Eet al., 2019, Author Correction: WWP2 regulates pathological cardiac fibrosis by modulating SMAD2 signaling, Nature Communications, Vol: 10, ISSN: 2041-1723

Journal article

Chen H, Moreno-Moral A, Pesce F, Devapragash N, Mancini M, Heng E, Rotival M, Srivastava P, Harmston N, Shkura K, Rackham O, Yu W-P, Sun X-M, Gui Zhen Tee N, Tan E, Barton P, Felkin L, Lara-Pezzi E, Angelini G, Beltrami C, Pravenec M, Schafer S, Bottolo L, Hubner N, Emanueli C, Cook S, Petretto Eet al., 2019, WWP2 regulates pathological cardiac fibrosis by modulating SMAD2 signaling, Nature Communications, Vol: 10, Pages: 1-19, ISSN: 2041-1723

Cardiac fibrosis is a final common pathology in inherited and acquired heart diseases that causes cardiac electrical and pump failure. Here, we use systems genetics to identify a pro-fibrotic gene network in the diseased heart and show that this network is regulated by the E3 ubiquitin ligase WWP2, specifically by the WWP2-N terminal isoform. Importantly, the WWP2-regulated pro-fibrotic gene network is conserved across different cardiac diseases characterized by fibrosis: human and murine dilated cardiomyopathy and repaired tetralogy of Fallot. Transgenic mice lacking the N-terminal region of the WWP2 protein show improved cardiac function and reduced myocardial fibrosis in response to pressure overload or myocardial infarction. In primary cardiac fibroblasts, WWP2 positively regulates the expression of pro-fibrotic markers and extracellular matrix genes. TGFβ1 stimulation promotes nuclear translocation of the WWP2 isoforms containing the N-terminal region and their interaction with SMAD2. WWP2 mediates the TGFβ1-induced nucleocytoplasmic shuttling and transcriptional activity of SMAD2.

Journal article

van Heesch S, Witte F, Schneider-Lunitz V, Schulz JF, Adami E, Faber AB, Kirchner M, Maatz H, Blachut S, Sandmann C-L, Kanda M, Worth CL, Schafer S, Calviello L, Merriott R, Patone G, Hummel O, Wyler E, Obermayer B, Mücke MB, Lindberg EL, Trnka F, Memczak S, Schilling M, Felkin LE, Barton PJR, Quaife NM, Vanezis K, Diecke S, Mukai M, Mah N, Oh S-J, Kurtz A, Schramm C, Schwinge D, Sebode M, Harakalova M, Asselbergs FW, Vink A, de Weger RA, Viswanathan S, Widjaja AA, Gärtner-Rommel A, Milting H, dos Remedios C, Knosalla C, Mertins P, Landthaler M, Vingron M, Linke WA, Seidman JG, Seidman CE, Rajewsky N, Ohler U, Cook SA, Hubner Net al., 2019, The translational landscape of the human heart, Cell, Vol: 178, Pages: 242-260.e29, ISSN: 0092-8674

Gene expression in human tissue has primarily been studied on the transcriptional level, largely neglecting translational regulation. Here, we analyze the translatomes of 80 human hearts to identify new translation events and quantify the effect of translational regulation. We show extensive translational control of cardiac gene expression, which is orchestrated in a process-specific manner. Translation downstream of predicted disease-causing protein-truncating variants appears to be frequent, suggesting inefficient translation termination. We identify hundreds of previously undetected microproteins, expressed from lncRNAs and circRNAs, for which we validate the protein products in vivo. The translation of microproteins is not restricted to the heart and prominent in the translatomes of human kidney and liver. We associate these microproteins with diverse cellular processes and compartments and find that many locate to the mitochondria. Importantly, dozens of microproteins are translated from lncRNAs with well-characterized noncoding functions, indicating previously unrecognized biology.

Journal article

Mazzarotto F, Tayal P, Buchan R, Midwinter W, Wilk A, Whiffin N, Govind R, Mazaika E, De Marvao A, Felkin L, Dawes T, Ahmad M, Edwards E, Ing A, Thomson K, Chan L, Sim D, Baksi J, Pantazis A, Roberts A, Watkins H, Funke B, O'Regan D, Olivotto I, Barton P, Prasad S, Cook S, Ware J, Walsh Ret al., 2019, RE-EVALUATING THE GENETIC CONTRIBUTION OF MONOGENIC DILATED CARDIOMYOPATHY, Annual Conference of the British-Cardiovascular-Society (BCS) - Digital Health Revolution, Publisher: BMJ PUBLISHING GROUP, Pages: A100-A100, ISSN: 1355-6037

Conference paper

Walsh R, Mazzarotto F, Whiffin N, Buchan R, Midwinter W, Wilk A, Li N, Felkin L, Ingold N, Govind R, Ahmad M, Mazaika E, Allouba M, Zhang X, de Marvao A, Day SM, Ashley E, Colan SD, Michels M, Pereira AC, Jacoby D, Ho CY, Thomson KL, Watkins H, Barton PJR, Olivotto I, Cook SA, Ware JSet al., 2019, Quantitative approaches to variant classification increase the yield and precision of genetic testing in Mendelian diseases: The case of hypertrophic cardiomyopathy, Genome Medicine, Vol: 11, ISSN: 1756-994X

BackgroundInternational guidelines for variant interpretation in Mendelian disease set stringent criteria to report a variant as (likely) pathogenic, prioritising control of false-positive rate over test sensitivity and diagnostic yield. Genetic testing is also more likely informative in individuals with well-characterised variants from extensively studied European-ancestry populations. Inherited cardiomyopathies are relatively common Mendelian diseases that allow empirical calibration and assessment of this framework.MethodsWe compared rare variants in large hypertrophic cardiomyopathy (HCM) cohorts (up to 6179 cases) to reference populations to identify variant classes with high prior likelihoods of pathogenicity, as defined by etiological fraction (EF). We analysed the distribution of variants using a bespoke unsupervised clustering algorithm to identify gene regions in which variants are significantly clustered in cases.ResultsAnalysis of variant distribution identified regions in which variants are significantly enriched in cases and variant location was a better discriminator of pathogenicity than generic computational functional prediction algorithms. Non-truncating variant classes with an EF ≥ 0.95 were identified in five established HCM genes. Applying this approach leads to an estimated 14–20% increase in cases with actionable HCM variants, i.e. variants classified as pathogenic/likely pathogenic that might be used for predictive testing in probands’ relatives.ConclusionsWhen found in a patient confirmed to have disease, novel variants in some genes and regions are empirically shown to have a sufficiently high probability of pathogenicity to support a “likely pathogenic” classification, even without additional segregation or functional data. This could increase the yield of high confidence actionable variants, consistent with the framework and recommendations of current guidelines. The techniques outlined offer a consisten

Journal article

Horvat C, Johnson R, Lam L, Munro J, Mazzarotto F, Roberts A, Herman D, Parfenov M, Haghighli A, Macdonough B, DePalma S, Keogh A, Macdonald P, Hayward C, Roberts A, Barton PJR, Felkin L, Giannoulatou E, Cook S, Seidman J, Siedman C, Fatkin Det al., 2019, A gene-centric strategy for identifying disease-causing rare variants in dilated cardiomyopathy, Genetics in Medicine, Vol: 21, Pages: 133-143, ISSN: 1098-3600

PurposeWe evaluated strategies for identifying disease-causing variants in genetic testing for dilated cardiomyopathy (DCM).MethodsCardiomyopathy gene panel testing was performed in 532 DCM patients and 527 healthy control subjects. Rare variants in 41 genes were stratified using variant-level and gene-level characteristics.ResultsA majority of DCM cases and controls carried rare protein-altering cardiomyopathy gene variants. Variant-level characteristics alone had limited discriminative value. Differentiation between groups was substantially improved by addition of gene-level information that incorporated ranking of genes based on literature evidence for disease association. The odds of DCM were increased to nearly 9-fold for truncating variants or high-impact missense variants in the subset of 14 genes that had the strongest biological links to DCM (P <0.0001). For some of these genes, DCM-associated variants appeared to be clustered in key protein functional domains. Multiple rare variants were present in many family probands, however, there was generally only one “driver” pathogenic variant that cosegregated with disease.ConclusionRare variants in cardiomyopathy genes can be effectively stratified by combining variant-level and gene-level information. Prioritization of genes based on their a priori likelihood of disease causation is a key factor in identifying clinically actionable variants in cardiac genetic testing.

Journal article

de Marvao A, Biffi C, Walsh R, Doumou G, Dawes T, Shi W, Bai W, Berry A, Buchan R, Pierce I, Tokarczuk P, Statton B, Francis C, Duan J, Quinlan M, Felkin L, Le T-T, Bhuva A, Tang HC, Barton P, Chin CW-L, Rueckert D, Ware J, Prasad S, O'Regan DP, Cook SAet al., 2018, Defining The Effects Of Genetic Variation Using Machine Learning Analysis Of CMRs: A Study In Hypertrophic Cardiomyopathy And In A Healthy Population, Joint Meeting of the British-Society-of-Cardiovascular-Imaging/British-Society-of-Cardiovascular-CT, British-Society-of-Cardiovascular-Magnetic-Resonance and British-Nuclear-Cardiac-Society on British Cardiovascular Imaging, Publisher: BMJ PUBLISHING GROUP, Pages: A7-A8, ISSN: 1355-6037

Conference paper

Walsh R, Buchan R, Wilk A, John S, Felkin L, Thomson KL, Chiaw TH, Chin Woon Loong C, Jian Pua C, Raphael C, Prasad S, Barton P, Funke B, Watkins H, Ware J, Cook SAet al., 2017, Defining the genetic architecture of hypertrophic cardiomyopathy: re-evaluating the role of non-sarcomeric genes, European Heart Journal, Vol: 38, Pages: 3461-3468, ISSN: 1522-9645

Aim: Hypertrophic cardiomyopathy (HCM)exhibits genetic heterogeneity that is dominated by variation in eight sarcomericgenes.Genetic variation in a large number of non-sarcomeric genes has also been implicated in HCM but not formally assessed. Here we used very large case and control cohorts to determine the extent to which variation in non-sarcomeric genes contributes to HCM.Methods and results: We sequenced known and putative HCM genes ina new large prospective HCM cohort (n=804) and analysed data alongside the largest published series of clinically genotyped HCM patients (n=6179), previously published HCM cohorts and reference population samples from the Exome Aggregation Consortium (ExAC, n=60,706) to assess variation in 31 genes implicated in HCM. We foundno significant excess of rare (minor allele frequency < 1:10,000 in ExAC)protein-alteringvariants over controls for most genes tested and conclude that novel variantsin these genes are rarely interpretable, even for genes with previous evidence of co-segregation (e.g. ACTN2). To provide an aid for variant interpretation, weintegratedHCM gene sequencedata with aggregatedpedigreeand functional data and suggest ameans of assessing genepathogenicity in HCMusing this evidence. Conclusions: We show that genetic variation in the majority of non-sarcomeric genes implicated in HCM is not associated with the condition, reinforce the fact that the sarcomeric gene variation is the primary cause of HCM known to date and underscore that the aetiology of HCM is unknown in the majority ofpatients.

Journal article

Heinig M, Adriaens ME, Schafer S, van Deutekom HWM, Lodder EM, Ware JS, Schneider V, Felkin LE, Creemers EE, Meder B, Katus HA, Ruehle F, Stoll M, Cambien F, Villard E, Charron P, Varro A, Bishopric NH, George AL, dos Remedios C, Moreno-Moral A, Pesce F, Bauerfeind A, Rueschendorf F, Rintisch C, Petretto E, Barton PJ, Cook SA, Pinto YM, Bezzina CR, Hubner Net al., 2017, Natural genetic variation of the cardiac transcriptome in non-diseased donors and patients with dilated cardiomyopathy, Genome Biology, Vol: 18, ISSN: 1474-7596

Background: Genetic variation is an important determinant of RNA transcription and splicing, which in turncontributes to variation in human traits, including cardiovascular diseases.Results: Here we report the first in-depth survey of heart transcriptome variation using RNA-sequencing in 97patients with dilated cardiomyopathy and 108 non-diseased controls. We reveal extensive differences of geneexpression and splicing between dilated cardiomyopathy patients and controls, affecting known as well as noveldilated cardiomyopathy genes. Moreover, we show a widespread effect of genetic variation on the regulation oftranscription, isoform usage, and allele-specific expression. Systematic annotation of genome-wide association SNPsidentifies 60 functional candidate genes for heart phenotypes, representing 20% of all published heart genome-wideassociation loci. Focusing on the dilated cardiomyopathy phenotype we found that eQTL variants are also enriched fordilated cardiomyopathy genome-wide association signals in two independent cohorts.Conclusions: RNA transcription, splicing, and allele-specific expression are each important determinants of the dilatedcardiomyopathy phenotype and are controlled by genetic factors. Our results represent a powerful resource for thefield of cardiovascular genetics.

Journal article

Gómez-Salinero JM, López-Olañeta MM, Ortiz-Sánchez P, Larrasa-Alonso J, Gatto A, Felkin LE, Barton PJ, Navarro-Lérida I, Ángel Del Pozo M, García-Pavía P, Sundararaman B, Giovinazo G, Yeo GW, Lara-Pezzi Eet al., 2016, The Calcineurin Variant CnAβ1 controls mouse embryonic stem cell differentiation by directing mTORC2 membrane localization and activation, Cell Chemical Biology, Vol: 23, Pages: 1372-1382, ISSN: 2451-9456

Embryonic stem cells (ESC) have the potential to generate all the cell lineages that form the body. However, the molecular mechanisms underlying ESC differentiation and especially the role of alternative splicing in this process remain poorly understood. Here, we show that the alternative splicing regulator MBNL1 promotes generation of the atypical calcineurin Aβ variant CnAβ1 in mouse ESCs (mESC). CnAβ1 has a unique C-terminal domain that drives its localization mainly to the Golgi apparatus by interacting with Cog8. CnAβ1 regulates the intracellular localization and activation of the mTORC2 complex. CnAβ1 knockdown results in delocalization of mTORC2 from the membrane to the cytoplasm, inactivation of the AKT/GSK3β/β-catenin signaling pathway, and defective mesoderm specification. In summary, here we unveil the structural basis for the mechanism of action of CnAβ1 and its role in the differentiation of mESCs to the mesodermal lineage.

Journal article

Felkin LE, Walsh R, Ware JS, Yacoub MH, Birks EJ, Barton PJR, Cook SAet al., 2016, Recovery of cardiac function in cardiomyopathy due to titin truncation, JAMA Cardiology, Vol: 1, Pages: 234-235, ISSN: 2380-6583

Dilated cardiomyopathy (DCM) is a frequent cause of heart failure and a common indication for heart transplantation. Dilated cardiomyopathy has a strong genetic basis, and the most common disease-causing mutations are variants that truncate the sarcomeric protein titin (TTN-truncating variants [TTNtvs] are prevalent in 25%1 of familial DCM cases and 13%2 of idiopathic DCM cases). The prognosis of DCM is poor, but functional recovery from end-stage failure has been reported following both optimal medical therapy3 and left ventricular assist device (LVAD) support,4,5 although the determinants of successful recovery are unknown. It has been proposed that recovery from genetic cardiomyopathy may not be expected because the underlying cause is irreversible, whereas recovery may be more likely when DCM is caused by reversible, nongenetic factors (eg, myocarditis).6 To address this directly, we sequenced TTN in patients with end-stage DCM who either recovered or did not recover following LVAD support.

Journal article

Fatkin D, Lam L, Herman DS, Benson CC, Felkin LE, Barton PJR, Walsh R, Candan S, Ware JS, Roberts AM, Chung WK, Smoot L, Bornaun H, Keogh AM, Macdonald PS, Hayward CS, Seidman JG, Roberts AE, Cook SA, Seidman CEet al., 2016, Titin truncating mutations: a rare cause of dilated cardiomyopathy in the young, Progress in Pediatric Cardiology, Vol: 40, Pages: 41-45, ISSN: 1058-9813

Truncating mutations in the TTN gene are the most common genetic cause of dilated cardiomyopathy in adults but their role in young patients is unknown. We studied 82 young dilated cardiomyopathy subjects and found that the prevalence of truncating TTN mutations in adolescents was similar to adults, but surprisingly few truncating TTN mutations were identified in affected children, including one confirmed de novo variant. In several cases, truncating TTN mutations in children with dilated cardiomyopathy had evidence of additional clinical or genetic risk factors. These findings have implications for genetic testing and suggest that single truncating TTN mutations are insufficient alone to cause pediatric-onset dilated cardiomyopathy.

Journal article

Buyandelger B, Mansfield C, Kostin S, Choi O, Roberts AM, Ware JS, Mazzarotto F, Pesce F, Buchan R, Isaacson RL, Vouffo J, Gunkel S, Knöll G, McSweeney SJ, Wei H, Perrot A, Pfeiffer C, Toliat MR, Ilieva K, Krysztofinska E, López-Olañeta MM, Gómez-Salinero JM, Schmidt A, Ng KE, Teucher N, Chen J, Teichmann M, Eilers M, Haverkamp W, Regitz-Zagrosek V, Hasenfuss G, Braun T, Pennell DJ, Gould I, Barton PJ, Lara-Pezzi E, Schafer S, Hübner N, Felkin LE, O'Regan DP, Petretto E, Brand T, Milting H, Nürnberg P, Schneider MD, Prasad S, Knöll Ret al., 2015, ZBTB17 (MIZ1) Is Important for the Cardiac Stress Response and a Novel Candidate Gene for Cardiomyopathy and Heart Failure., Circulation. Cardiovascular Genetics, Vol: 8, Pages: 643-652, ISSN: 1942-3268

BACKGROUND: -Mutations in sarcomeric and cytoskeletal proteins are a major cause of hereditary cardiomyopathies, but our knowledge remains incomplete as to how the genetic defects execute their effects. METHODS AND RESULTS: -We used cysteine and glycine-rich protein 3 (CSRP3), a known cardiomyopathy gene, in a yeast two-hybrid screen and identified zinc finger and BTB domain containing protein 17 (ZBTB17) as a novel interacting partner. ZBTB17 is a transcription factor that contains the peak association signal (rs10927875) at the replicated 1p36 cardiomyopathy locus. ZBTB17 expression protected cardiac myocytes from apoptosis in vitro and in a mouse model with cardiac myocyte-specific deletion of Zbtb17, which develops cardiomyopathy and fibrosis after biomechanical stress. ZBTB17 also regulated cardiac myocyte hypertrophy in vitro and in vivo in a calcineurin-dependent manner. CONCLUSIONS: -We revealed new functions for ZBTB17 in the heart, a transcription factor which may play a role as a novel cardiomyopathy gene.

Journal article

Roberts AM, Ware J, Herman D, Schafer S, Mazzarotto F, Baksi J, Buchan R, Walsh R, John S, Wilkinson S, Felkin L, Bick A, Radke M, Gotthardt M, Barton P, Hubner N, Seidman J, Seidman C, Cook Set al., 2015, INTEGRATED ALLELIC, TRANSCRIPTIONAL, AND PHENOTYPIC DISSECTION OF THE CARDIAC EFFECTS OF TITIN VARIATION IN HEALTH AND DISEASER, British-Cardiac-Society (BCS) Annual Conference on Hearts and Genes, Publisher: BMJ PUBLISHING GROUP, Pages: A126-A126, ISSN: 1355-6037

Conference paper

Roberts A, Ware J, Herman D, Schafer S, Mazzarotto F, Baksi J, Buchan R, Walsh R, John S, Wilkinson S, Felkin L, Bick A, Radke M, Gotthardt M, Barton P, Hubner N, Seidman J, Seidman C, Cook Set al., 2015, INTEGRATED ALLELIC, TRANSCRIPTIONAL, AND PHENOTYPIC DISSECTION OF THE CARDIAC EFFECTS OF TITIN VARIATION IN HEALTH AND DISEASE, British-Cardiac-Society (BCS) Annual Conference on Hearts and Genes, Publisher: BMJ PUBLISHING GROUP, Pages: A93-A93, ISSN: 1355-6037

Conference paper

Roberts AM, Ware JS, Herman DS, Schafer S, Baksi J, Bick AG, Buchan RJ, Walsh R, John S, Wilkinson S, Mazzarotto F, Felkin LE, Gong S, L MacArthur JA, Cunningham F, Flannick J, Gabriel SB, Altshuler DM, Macdonald PS, Heinig M, Keogh AM, Hayward CS, Banner NR, Pennell DJ, O'Regan DP, San TR, de Marvao A, W Dawes TJ, Gulati A, Birks EJ, Yacoub MH, Radke M, Gotthardt M, Wilson JG, O'Donnell CJ, Prasad SK, Barton PJ, Fatkin D, Hubner N, Seidman JG, Seidman CE, Cook SAet al., 2015, Integrated allelic, transcriptional, and phenomic dissection of the cardiac effects of titin truncations in health and disease, Science Translational Medicine, Vol: 7, Pages: 270ra6-270ra6, ISSN: 1946-6234

The recent discovery of heterozygous human mutations that truncate full-length titin (TTN, an abundant structural, sensory, and signaling filament in muscle) as a common cause of end-stage dilated cardiomyopathy (DCM) promises new prospects for improving heart failure management. However, realization of this opportunity has been hindered by the burden of TTN-truncating variants (TTNtv) in the general population and uncertainty about their consequences in health or disease. To elucidate the effects of TTNtv, we coupled TTN gene sequencing with cardiac phenotyping in 5267 individuals across the spectrum of cardiac physiology and integrated these data with RNA and protein analyses of human heart tissues. We report diversity of TTN isoform expression in the heart, define the relative inclusion of TTN exons in different isoforms (using the TTN transcript annotations available at http://cardiodb.org/titin), and demonstrate that these data, coupled with the position of the TTNtv, provide a robust strategy to discriminate pathogenic from benign TTNtv. We show that TTNtv is the most common genetic cause of DCM in ambulant patients in the community, identify clinically important manifestations of TTNtv-positive DCM, and define the penetrance and outcomes of TTNtv in the general population. By integrating genetic, transcriptome, and protein analyses, we provide evidence for a length-dependent mechanism of disease. These data inform diagnostic criteria and management strategies for TTNtv-positive DCM patients and for TTNtv that are identified as incidental findings.

Journal article

Roberts AM, Ware JS, Herman DS, Schafer S, Baksi J, Bick AG, Buchan RJ, Walsh R, John S, Wilkinson S, Mazzarotto F, Felkin LE, Gong S, MacArthur JAL, Cunningham F, Flannick J, Gabriel SB, Altshuler DM, Macdonald PS, Heinig M, Keogh AM, Hayward CS, Banner NR, Pennell DJ, O'Regan DP, San TR, De Marvao A, Dawes TJW, Gulati A, Birks EJ, Yacoub MH, Radke M, Gotthardt M, Wilson JG, O'Donnell CJ, Prasad SK, Barton PJR, Fatkin D, Hubner N, Seidman JG, Seidman CE, Cook SAet al., 2015, What Happens When Titins Are Trimmed?, SCIENCE TRANSLATIONAL MEDICINE, Vol: 7, ISSN: 1946-6234

Journal article

Lee R, Xu B, Rame JE, Felkin LE, Barton P, Dries DLet al., 2014, Regulated Inositol‐Requiring Protein 1‐Dependent Decay as a Mechanism of Corin RNA and Protein Deficiency in Advanced Human Systolic Heart Failure, Journal of the American Heart Association, Vol: 3, ISSN: 2047-9980

BACKGROUND: The compensatory actions of the endogenous natriuretic peptide system require adequate processing of natriuretic peptide pro‐hormones into biologically active, carboxyl‐terminal fragments. Natriuretic peptide pro‐peptide processing is accomplished by corin, a transmembrane serine protease expressed by cardiomyocytes. Brain natriuretic peptide (BNP) processing is inadequate in advanced heart failure and is independently associated with adverse outcomes; however, the molecular mechanisms causing impaired BNP processing are not understood. We hypothesized that the development of endoplasmic reticulum stress in cardiomyocytes in advanced heart failure triggers inositol‐requiring protein 1 (IRE1)‐dependent corin mRNA decay, which would favor a molecular substrate favoring impaired natriuretic peptide pro‐peptide processing. METHODS AND RESULTS: Two independent samples of hearts obtained from patients with advanced heart failure at transplant demonstrated that corin RNA was reduced as Atrial natriuretic peptide (ANP)/BNP RNA increased. Increases in spliced X‐box protein 1, a marker for IRE1‐endoribonuclease activity, were associated with decreased corin RNA. Moreover, ≈50% of the hearts demonstrated significant reductions in corin RNA and protein as compared to the nonfailing control sample. In vitro experiments demonstrated that induction of endoplasmic reticulum stress in cultured cardiomyocytes with thapsigargin activated IRE1's endoribonuclease activity and time‐dependent reductions in corin mRNA. In HL‐1 cells, overexpression of IRE1 activated IRE1 endoribonuclease activity and caused corin mRNA decay, whereas IRE1‐RNA interference with shRNA attenuated corin mRNA decay after induction of endoplasmic reticulum stress with thapsigargin. Pre‐treatment of cells with Actinomycin D to inhibit transcription did not alter the magnitude or time course of thapsigargin‐induced corin mRNA decline, supporting the hypothesis that this was the result of IRE1‐medi

Journal article

Seemampillai B, Germack R, Felkin LE, McCormack A, Rose MLet al., 2014, Heat Shock Protein-27 Delays Acute Rejection After Cardiac Transplantation: An Experimental Model, TRANSPLANTATION, Vol: 98, Pages: 29-38, ISSN: 0041-1337

Journal article

Dias P, Navaratnarajah M, Alayoubi S, Kane C, Felkin LE, Cartledge JE, Jayaratne N, Latif N, Yacoub MH, Terracciano CMet al., 2014, Ivabradine Reduces α-Smooth Muscle Actin Expression, Proliferation and Collagen Production in Human Cardiac Fibroblasts, Publisher: Elsevier, Pages: 759a-759a

Conference paper

Panse KD, Felkin LE, Lopez-Olaneta MM, Gomez-Salinero J, Villalba M, Munoz L, Nakamura K, Shimano M, Walsh K, Barton PJR, Rosenthal N, Lara-Pezzi Eet al., 2012, Follistatin-Like 3 Mediates Paracrine Fibroblast Activation by Cardiomyocytes, JOURNAL OF CARDIOVASCULAR TRANSLATIONAL RESEARCH, Vol: 5, Pages: 814-826, ISSN: 1937-5387

Journal article

Buyandelger B, Linke W, Zou P, Kostin S, Ku C, Felkin L, Birks E, Barton P, Sattler M, Knoell Ret al., 2012, Telethonin, Z-disc and myocardial performance, 2nd Congress of the European-Society-of-Cardiology Council on Basic Cardiovascular Science - Frontiers in Cardiovascular Biology, Publisher: OXFORD UNIV PRESS, Pages: S100-S100, ISSN: 0008-6363

Conference paper

McDermott-Roe C, Ye J, Ahmed R, Sun XM, Serafin A, Ware J, Bottolo L, Mucket P, Canas X, Zhang J, Rowe GC, Buchan R, Lu H, Braithwaite A, Mancici M, Hauton D, Marti R, García-Arumí E, Hubner N, Jacob H, Serikawa T, Zidek P, Papousek F, Kolar F, Cardona M, Ruiz-Meana M, García-Dorado D, Comella JX, Felkin LE, Barton PJ, Arany Z, Pravenec M, Sanchis D, Cook SAet al., 2011, Endonuclease G is a novel determinant of cardiac hypertrophy and mitochondrial function, Nature, Vol: 478, Pages: 114-118

Journal article

Knoell R, Linke WA, Zou P, Miocic S, Kostin S, Buyandelger B, Ku C-H, Neef S, Bug M, Schaefer K, Knoell G, Felkin LE, Wessels J, Toischer K, Hagn F, Kessler H, Didie M, Quentin T, Maier LS, Teucher N, Unsoeld B, Schmidt A, Birks EJ, Gunkel S, Lang P, Granzier H, Zimmermann W-H, Field LJ, Faulkner G, Dobbelstein M, Barton PJR, Sattler M, Wilmanns M, Chien KRet al., 2011, Telethonin Deficiency Is Associated With Maladaptation to Biomechanical Stress in the Mammalian Heart, CIRCULATION RESEARCH, Vol: 109, Pages: 758-U153, ISSN: 0009-7330

Journal article

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