61 results on '"Ulirsch JC"'
Search Results
2. [mu]-Opioid Receptor Gene A118G Polymorphism Predicts Pain Recovery After Sexual Assault.
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Ballina LE, Ulirsch JC, Soward AC, Rossi C, Rotolo S, Linnstaedt SD, Heafner T, Foley KA, Batts J, Collette R, Holbrook D, Zelman S, and McLean SA
- Abstract
Pain is common after sexual assault (SA), but etiology of pain symptoms after SA is unknown. Preclinical studies suggest that the release of endogenous opioids during stress produces delayed-onset hyperalgesia. In human studies, individuals with >=1 G allele at the [mu]-opioid receptor functional single nucleotide polymorphism A118G have been shown to have a reduced response to opioids. We hypothesized that if opioid-mediated hyperalgesia contributes to pain after SA, women SA survivors with 1 or more G alleles at A118G would experience reduced postassault pain. Among 52 European American women SA survivors presenting for care within 48 hours of SA, those with a G allele (12/52, 23%) experienced less severe pain (F[1,39] = 11.55, P = .002) and a reduced extent of pain (F[1,41] = 11.01, P = .002) during the 6 weeks after SA. These associations between the presence of 1 or more G alleles and reduced pain severity and reduced pain extent after SA remained significant in multivariable models controlling for age, income, education, reported pain prior to assault, and pain at the time of initial evaluation. PERSPECTIVE: These results suggest that endogenous opioid-mediated hyperalgesia may contribute to pain symptoms after sexual assault. Further studies examining mechanisms mediating the development of pain after sexual assault, and the potential influence of opioid-mediated hyperalgesia, are needed. [ABSTRACT FROM AUTHOR]
- Published
- 2013
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3. Functional dissection of complex and molecular trait variants at single nucleotide resolution.
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Siraj L, Castro RI, Dewey H, Kales S, Nguyen TTL, Kanai M, Berenzy D, Mouri K, Wang QS, McCaw ZR, Gosai SJ, Aguet F, Cui R, Vockley CM, Lareau CA, Okada Y, Gusev A, Jones TR, Lander ES, Sabeti PC, Finucane HK, Reilly SK, Ulirsch JC, and Tewhey R
- Abstract
Identifying the causal variants and mechanisms that drive complex traits and diseases remains a core problem in human genetics. The majority of these variants have individually weak effects and lie in non-coding gene-regulatory elements where we lack a complete understanding of how single nucleotide alterations modulate transcriptional processes to affect human phenotypes. To address this, we measured the activity of 221,412 trait-associated variants that had been statistically fine-mapped using a Massively Parallel Reporter Assay (MPRA) in 5 diverse cell-types. We show that MPRA is able to discriminate between likely causal variants and controls, identifying 12,025 regulatory variants with high precision. Although the effects of these variants largely agree with orthogonal measures of function, only 69% can plausibly be explained by the disruption of a known transcription factor (TF) binding motif. We dissect the mechanisms of 136 variants using saturation mutagenesis and assign impacted TFs for 91% of variants without a clear canonical mechanism. Finally, we provide evidence that epistasis is prevalent for variants in close proximity and identify multiple functional variants on the same haplotype at a small, but important, subset of trait-associated loci. Overall, our study provides a systematic functional characterization of likely causal common variants underlying complex and molecular human traits, enabling new insights into the regulatory grammar underlying disease risk., Competing Interests: Competing Interests PCS is a co-founder of and consultant to Sherlock Biosciences and Board Member of Danaher Corporation. PCS and RT hold patents related to the application of MPRA. JCU and FA are employees of Illumina. QSW is an employee of Calico Life Sciences LLC. ZRM is an employee of insitro.
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- 2024
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4. Improving fine-mapping by modeling infinitesimal effects.
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Cui R, Elzur RA, Kanai M, Ulirsch JC, Weissbrod O, Daly MJ, Neale BM, Fan Z, and Finucane HK
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- Humans, Bayes Theorem, Multifactorial Inheritance, Algorithms, Genome-Wide Association Study, Polymorphism, Single Nucleotide
- Abstract
Fine-mapping aims to identify causal genetic variants for phenotypes. Bayesian fine-mapping algorithms (for example, SuSiE, FINEMAP, ABF and COJO-ABF) are widely used, but assessing posterior probability calibration remains challenging in real data, where model misspecification probably exists, and true causal variants are unknown. We introduce replication failure rate (RFR), a metric to assess fine-mapping consistency by downsampling. SuSiE, FINEMAP and COJO-ABF show high RFR, indicating potential overconfidence in their output. Simulations reveal that nonsparse genetic architecture can lead to miscalibration, while imputation noise, nonuniform distribution of causal variants and quality control filters have minimal impact. Here we present SuSiE-inf and FINEMAP-inf, fine-mapping methods modeling infinitesimal effects alongside fewer larger causal effects. Our methods show improved calibration, RFR and functional enrichment, competitive recall and computational efficiency. Notably, using our methods' posterior effect sizes substantially increases polygenic risk score accuracy over SuSiE and FINEMAP. Our work improves causal variant identification for complex traits, a fundamental goal of human genetics., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)
- Published
- 2024
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5. Identification of constrained sequence elements across 239 primate genomes.
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Kuderna LFK, Ulirsch JC, Rashid S, Ameen M, Sundaram L, Hickey G, Cox AJ, Gao H, Kumar A, Aguet F, Christmas MJ, Clawson H, Haeussler M, Janiak MC, Kuhlwilm M, Orkin JD, Bataillon T, Manu S, Valenzuela A, Bergman J, Rouselle M, Silva FE, Agueda L, Blanc J, Gut M, de Vries D, Goodhead I, Harris RA, Raveendran M, Jensen A, Chuma IS, Horvath JE, Hvilsom C, Juan D, Frandsen P, Schraiber JG, de Melo FR, Bertuol F, Byrne H, Sampaio I, Farias I, Valsecchi J, Messias M, da Silva MNF, Trivedi M, Rossi R, Hrbek T, Andriaholinirina N, Rabarivola CJ, Zaramody A, Jolly CJ, Phillips-Conroy J, Wilkerson G, Abee C, Simmons JH, Fernandez-Duque E, Kanthaswamy S, Shiferaw F, Wu D, Zhou L, Shao Y, Zhang G, Keyyu JD, Knauf S, Le MD, Lizano E, Merker S, Navarro A, Nadler T, Khor CC, Lee J, Tan P, Lim WK, Kitchener AC, Zinner D, Gut I, Melin AD, Guschanski K, Schierup MH, Beck RMD, Karakikes I, Wang KC, Umapathy G, Roos C, Boubli JP, Siepel A, Kundaje A, Paten B, Lindblad-Toh K, Rogers J, Marques Bonet T, and Farh KK
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- Animals, Female, Humans, Pregnancy, Deoxyribonuclease I metabolism, DNA genetics, DNA metabolism, Mammals classification, Mammals genetics, Placenta, Regulatory Sequences, Nucleic Acid genetics, Reproducibility of Results, Transcription Factors metabolism, Proteins genetics, Gene Expression Regulation genetics, Conserved Sequence genetics, Evolution, Molecular, Genome genetics, Primates classification, Primates genetics
- Abstract
Noncoding DNA is central to our understanding of human gene regulation and complex diseases
1,2 , and measuring the evolutionary sequence constraint can establish the functional relevance of putative regulatory elements in the human genome3-9 . Identifying the genomic elements that have become constrained specifically in primates has been hampered by the faster evolution of noncoding DNA compared to protein-coding DNA10 , the relatively short timescales separating primate species11 , and the previously limited availability of whole-genome sequences12 . Here we construct a whole-genome alignment of 239 species, representing nearly half of all extant species in the primate order. Using this resource, we identified human regulatory elements that are under selective constraint across primates and other mammals at a 5% false discovery rate. We detected 111,318 DNase I hypersensitivity sites and 267,410 transcription factor binding sites that are constrained specifically in primates but not across other placental mammals and validate their cis-regulatory effects on gene expression. These regulatory elements are enriched for human genetic variants that affect gene expression and complex traits and diseases. Our results highlight the important role of recent evolution in regulatory sequence elements differentiating primates, including humans, from other placental mammals., (© 2023. The Author(s).)- Published
- 2024
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6. An encyclopedia of enhancer-gene regulatory interactions in the human genome.
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Gschwind AR, Mualim KS, Karbalayghareh A, Sheth MU, Dey KK, Jagoda E, Nurtdinov RN, Xi W, Tan AS, Jones H, Ma XR, Yao D, Nasser J, Avsec Ž, James BT, Shamim MS, Durand NC, Rao SSP, Mahajan R, Doughty BR, Andreeva K, Ulirsch JC, Fan K, Perez EM, Nguyen TC, Kelley DR, Finucane HK, Moore JE, Weng Z, Kellis M, Bassik MC, Price AL, Beer MA, Guigó R, Stamatoyannopoulos JA, Lieberman Aiden E, Greenleaf WJ, Leslie CS, Steinmetz LM, Kundaje A, and Engreitz JM
- Abstract
Identifying transcriptional enhancers and their target genes is essential for understanding gene regulation and the impact of human genetic variation on disease
1-6 . Here we create and evaluate a resource of >13 million enhancer-gene regulatory interactions across 352 cell types and tissues, by integrating predictive models, measurements of chromatin state and 3D contacts, and largescale genetic perturbations generated by the ENCODE Consortium7 . We first create a systematic benchmarking pipeline to compare predictive models, assembling a dataset of 10,411 elementgene pairs measured in CRISPR perturbation experiments, >30,000 fine-mapped eQTLs, and 569 fine-mapped GWAS variants linked to a likely causal gene. Using this framework, we develop a new predictive model, ENCODE-rE2G, that achieves state-of-the-art performance across multiple prediction tasks, demonstrating a strategy involving iterative perturbations and supervised machine learning to build increasingly accurate predictive models of enhancer regulation. Using the ENCODE-rE2G model, we build an encyclopedia of enhancer-gene regulatory interactions in the human genome, which reveals global properties of enhancer networks, identifies differences in the functions of genes that have more or less complex regulatory landscapes, and improves analyses to link noncoding variants to target genes and cell types for common, complex diseases. By interpreting the model, we find evidence that, beyond enhancer activity and 3D enhancer-promoter contacts, additional features guide enhancerpromoter communication including promoter class and enhancer-enhancer synergy. Altogether, these genome-wide maps of enhancer-gene regulatory interactions, benchmarking software, predictive models, and insights about enhancer function provide a valuable resource for future studies of gene regulation and human genetics., Competing Interests: Conflict of Interest Statement Z.A. is employed by Google DeepMind. J.C.U. is an employee of Illumina, Inc. D.R.K. is employed by Calico Life Sciences LLC. Z.W. co-founded Rgenta Therapeutics, and she serves as a scientific advisor for the company and is a member of its board. W.J.G. is an inventor on IP licensed by 10x Genomics. A.Kundaje is on the scientific advisory board of PatchBio, SerImmune and OpenTargets, was a consultant with Illumina, and owns shares in DeepGenomics, ImmunAI and Freenome. J.M.E. is a consultant and equity holder in Martingale Labs, Inc. and has received materials from 10x Genomics unrelated to this study.- Published
- 2023
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7. RNA polymerase II pausing temporally coordinates cell cycle progression and erythroid differentiation.
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Martell DJ, Merens HE, Caulier A, Fiorini C, Ulirsch JC, Ietswaart R, Choquet K, Graziadei G, Brancaleoni V, Cappellini MD, Scott C, Roberts N, Proven M, Roy NBA, Babbs C, Higgs DR, Sankaran VG, and Churchman LS
- Subjects
- Humans, Cell Differentiation, Cell Cycle, Transcription, Genetic, Nuclear Proteins metabolism, Transcriptional Elongation Factors genetics, RNA Polymerase II genetics, RNA Polymerase II metabolism, Gene Expression Regulation
- Abstract
Controlled release of promoter-proximal paused RNA polymerase II (RNA Pol II) is crucial for gene regulation. However, studying RNA Pol II pausing is challenging, as pause-release factors are almost all essential. In this study, we identified heterozygous loss-of-function mutations in SUPT5H, which encodes SPT5, in individuals with β-thalassemia. During erythropoiesis in healthy human cells, cell cycle genes were highly paused as cells transition from progenitors to precursors. When the pathogenic mutations were recapitulated by SUPT5H editing, RNA Pol II pause release was globally disrupted, and as cells began transitioning from progenitors to precursors, differentiation was delayed, accompanied by a transient lag in erythroid-specific gene expression and cell cycle kinetics. Despite this delay, cells terminally differentiate, and cell cycle phase distributions normalize. Therefore, hindering pause release perturbs proliferation and differentiation dynamics at a key transition during erythropoiesis, identifying a role for RNA Pol II pausing in temporally coordinating the cell cycle and erythroid differentiation., Competing Interests: Declaration of interests V.G.S. serves as an advisor to and/or has equity in Branch Biosciences, Ensoma, Novartis, Forma, Sana Biotechnology, and Cellarity, all unrelated to the present work. J.C.U. is an employee of Illumina, Inc., unrelated to the present work. R.I. is a founder, board member, and shareholder of Cellforma, unrelated to the present work., (Copyright © 2023 Elsevier Inc. All rights reserved.)
- Published
- 2023
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8. Leveraging polygenic enrichments of gene features to predict genes underlying complex traits and diseases.
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Weeks EM, Ulirsch JC, Cheng NY, Trippe BL, Fine RS, Miao J, Patwardhan TA, Kanai M, Nasser J, Fulco CP, Tashman KC, Aguet F, Li T, Ordovas-Montanes J, Smillie CS, Biton M, Shalek AK, Ananthakrishnan AN, Xavier RJ, Regev A, Gupta RM, Lage K, Ardlie KG, Hirschhorn JN, Lander ES, Engreitz JM, and Finucane HK
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- Humans, Genome-Wide Association Study methods, Genetic Predisposition to Disease genetics, Phenotype, Polymorphism, Single Nucleotide genetics, Multifactorial Inheritance genetics, Quantitative Trait Loci genetics
- Abstract
Genome-wide association studies (GWASs) are a valuable tool for understanding the biology of complex human traits and diseases, but associated variants rarely point directly to causal genes. In the present study, we introduce a new method, polygenic priority score (PoPS), that learns trait-relevant gene features, such as cell-type-specific expression, to prioritize genes at GWAS loci. Using a large evaluation set of genes with fine-mapped coding variants, we show that PoPS and the closest gene individually outperform other gene prioritization methods, but observe the best overall performance by combining PoPS with orthogonal methods. Using this combined approach, we prioritize 10,642 unique gene-trait pairs across 113 complex traits and diseases with high precision, finding not only well-established gene-trait relationships but nominating new genes at unresolved loci, such as LGR4 for estimated glomerular filtration rate and CCR7 for deep vein thrombosis. Overall, we demonstrate that PoPS provides a powerful addition to the gene prioritization toolbox., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)
- Published
- 2023
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9. Rare penetrant mutations confer severe risk of common diseases.
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Fiziev PP, McRae J, Ulirsch JC, Dron JS, Hamp T, Yang Y, Wainschtein P, Ni Z, Schraiber JG, Gao H, Cable D, Field Y, Aguet F, Fasnacht M, Metwally A, Rogers J, Marques-Bonet T, Rehm HL, O'Donnell-Luria A, Khera AV, and Farh KK
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- Humans, Genome-Wide Association Study, Mutation, Phenotype, Risk Factors, Genetic Predisposition to Disease, Multifactorial Inheritance, Penetrance
- Abstract
We examined 454,712 exomes for genes associated with a wide spectrum of complex traits and common diseases and observed that rare, penetrant mutations in genes implicated by genome-wide association studies confer ~10-fold larger effects than common variants in the same genes. Consequently, an individual at the phenotypic extreme and at the greatest risk for severe, early-onset disease is better identified by a few rare penetrant variants than by the collective action of many common variants with weak effects. By combining rare variants across phenotype-associated genes into a unified genetic risk model, we demonstrate superior portability across diverse global populations compared with common-variant polygenic risk scores, greatly improving the clinical utility of genetic-based risk prediction.
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- 2023
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10. Rare penetrant mutations confer severe risk of common diseases.
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Fiziev P, McRae J, Ulirsch JC, Dron JS, Hamp T, Yang Y, Wainschtein P, Ni Z, Schraiber JG, Gao H, Cable D, Field Y, Aguet F, Fasnacht M, Metwally A, Rogers J, Marques-Bonet T, Rehm HL, O'Donnell-Luria A, Khera AV, and Kai-How Farh K
- Abstract
We examined 454,712 exomes for genes associated with a wide spectrum of complex traits and common diseases and observed that rare, penetrant mutations in genes implicated by genome-wide association studies confer ∼10-fold larger effects than common variants in the same genes. Consequently, an individual at the phenotypic extreme and at the greatest risk for severe, early-onset disease is better identified by a few rare penetrant variants than by the collective action of many common variants with weak effects. By combining rare variants across phenotype-associated genes into a unified genetic risk model, we demonstrate superior portability across diverse global populations compared to common variant polygenic risk scores, greatly improving the clinical utility of genetic-based risk prediction., One Sentence Summary: Rare variant polygenic risk scores identify individuals with outlier phenotypes in common human diseases and complex traits.
- Published
- 2023
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11. Author Correction: FinnGen provides genetic insights from a well-phenotyped isolated population.
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Kurki MI, Karjalainen J, Palta P, Sipilä TP, Kristiansson K, Donner KM, Reeve MP, Laivuori H, Aavikko M, Kaunisto MA, Loukola A, Lahtela E, Mattsson H, Laiho P, Della Briotta Parolo P, Lehisto AA, Kanai M, Mars N, Rämö J, Kiiskinen T, Heyne HO, Veerapen K, Rüeger S, Lemmelä S, Zhou W, Ruotsalainen S, Pärn K, Hiekkalinna T, Koskelainen S, Paajanen T, Llorens V, Gracia-Tabuenca J, Siirtola H, Reis K, Elnahas AG, Sun B, Foley CN, Aalto-Setälä K, Alasoo K, Arvas M, Auro K, Biswas S, Bizaki-Vallaskangas A, Carpen O, Chen CY, Dada OA, Ding Z, Ehm MG, Eklund K, Färkkilä M, Finucane H, Ganna A, Ghazal A, Graham RR, Green EM, Hakanen A, Hautalahti M, Hedman ÅK, Hiltunen M, Hinttala R, Hovatta I, Hu X, Huertas-Vazquez A, Huilaja L, Hunkapiller J, Jacob H, Jensen JN, Joensuu H, John S, Julkunen V, Jung M, Junttila J, Kaarniranta K, Kähönen M, Kajanne R, Kallio L, Kälviäinen R, Kaprio J, Kerimov N, Kettunen J, Kilpeläinen E, Kilpi T, Klinger K, Kosma VM, Kuopio T, Kurra V, Laisk T, Laukkanen J, Lawless N, Liu A, Longerich S, Mägi R, Mäkelä J, Mäkitie A, Malarstig A, Mannermaa A, Maranville J, Matakidou A, Meretoja T, Mozaffari SV, Niemi MEK, Niemi M, Niiranen T, O Donnell CJ, Obeidat ME, Okafo G, Ollila HM, Palomäki A, Palotie T, Partanen J, Paul DS, Pelkonen M, Pendergrass RK, Petrovski S, Pitkäranta A, Platt A, Pulford D, Punkka E, Pussinen P, Raghavan N, Rahimov F, Rajpal D, Renaud NA, Riley-Gillis B, Rodosthenous R, Saarentaus E, Salminen A, Salminen E, Salomaa V, Schleutker J, Serpi R, Shen HY, Siegel R, Silander K, Siltanen S, Soini S, Soininen H, Sul JH, Tachmazidou I, Tasanen K, Tienari P, Toppila-Salmi S, Tukiainen T, Tuomi T, Turunen JA, Ulirsch JC, Vaura F, Virolainen P, Waring J, Waterworth D, Yang R, Nelis M, Reigo A, Metspalu A, Milani L, Esko T, Fox C, Havulinna AS, Perola M, Ripatti S, Jalanko A, Laitinen T, Mäkelä TP, Plenge R, McCarthy M, Runz H, Daly MJ, and Palotie A
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- 2023
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12. FinnGen provides genetic insights from a well-phenotyped isolated population.
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Kurki MI, Karjalainen J, Palta P, Sipilä TP, Kristiansson K, Donner KM, Reeve MP, Laivuori H, Aavikko M, Kaunisto MA, Loukola A, Lahtela E, Mattsson H, Laiho P, Della Briotta Parolo P, Lehisto AA, Kanai M, Mars N, Rämö J, Kiiskinen T, Heyne HO, Veerapen K, Rüeger S, Lemmelä S, Zhou W, Ruotsalainen S, Pärn K, Hiekkalinna T, Koskelainen S, Paajanen T, Llorens V, Gracia-Tabuenca J, Siirtola H, Reis K, Elnahas AG, Sun B, Foley CN, Aalto-Setälä K, Alasoo K, Arvas M, Auro K, Biswas S, Bizaki-Vallaskangas A, Carpen O, Chen CY, Dada OA, Ding Z, Ehm MG, Eklund K, Färkkilä M, Finucane H, Ganna A, Ghazal A, Graham RR, Green EM, Hakanen A, Hautalahti M, Hedman ÅK, Hiltunen M, Hinttala R, Hovatta I, Hu X, Huertas-Vazquez A, Huilaja L, Hunkapiller J, Jacob H, Jensen JN, Joensuu H, John S, Julkunen V, Jung M, Junttila J, Kaarniranta K, Kähönen M, Kajanne R, Kallio L, Kälviäinen R, Kaprio J, Kerimov N, Kettunen J, Kilpeläinen E, Kilpi T, Klinger K, Kosma VM, Kuopio T, Kurra V, Laisk T, Laukkanen J, Lawless N, Liu A, Longerich S, Mägi R, Mäkelä J, Mäkitie A, Malarstig A, Mannermaa A, Maranville J, Matakidou A, Meretoja T, Mozaffari SV, Niemi MEK, Niemi M, Niiranen T, O Donnell CJ, Obeidat ME, Okafo G, Ollila HM, Palomäki A, Palotie T, Partanen J, Paul DS, Pelkonen M, Pendergrass RK, Petrovski S, Pitkäranta A, Platt A, Pulford D, Punkka E, Pussinen P, Raghavan N, Rahimov F, Rajpal D, Renaud NA, Riley-Gillis B, Rodosthenous R, Saarentaus E, Salminen A, Salminen E, Salomaa V, Schleutker J, Serpi R, Shen HY, Siegel R, Silander K, Siltanen S, Soini S, Soininen H, Sul JH, Tachmazidou I, Tasanen K, Tienari P, Toppila-Salmi S, Tukiainen T, Tuomi T, Turunen JA, Ulirsch JC, Vaura F, Virolainen P, Waring J, Waterworth D, Yang R, Nelis M, Reigo A, Metspalu A, Milani L, Esko T, Fox C, Havulinna AS, Perola M, Ripatti S, Jalanko A, Laitinen T, Mäkelä TP, Plenge R, McCarthy M, Runz H, Daly MJ, and Palotie A
- Subjects
- Humans, Middle Aged, Estonia, Finland, Genetic Predisposition to Disease genetics, Genome-Wide Association Study, Meta-Analysis as Topic, United Kingdom, White People genetics, Disease genetics, Gene Frequency genetics, Phenotype
- Abstract
Population isolates such as those in Finland benefit genetic research because deleterious alleles are often concentrated on a small number of low-frequency variants (0.1% ≤ minor allele frequency < 5%). These variants survived the founding bottleneck rather than being distributed over a large number of ultrarare variants. Although this effect is well established in Mendelian genetics, its value in common disease genetics is less explored
1,2 . FinnGen aims to study the genome and national health register data of 500,000 Finnish individuals. Given the relatively high median age of participants (63 years) and the substantial fraction of hospital-based recruitment, FinnGen is enriched for disease end points. Here we analyse data from 224,737 participants from FinnGen and study 15 diseases that have previously been investigated in large genome-wide association studies (GWASs). We also include meta-analyses of biobank data from Estonia and the United Kingdom. We identified 30 new associations, primarily low-frequency variants, enriched in the Finnish population. A GWAS of 1,932 diseases also identified 2,733 genome-wide significant associations (893 phenome-wide significant (PWS), P < 2.6 × 10-11 ) at 2,496 (771 PWS) independent loci with 807 (247 PWS) end points. Among these, fine-mapping implicated 148 (73 PWS) coding variants associated with 83 (42 PWS) end points. Moreover, 91 (47 PWS) had an allele frequency of <5% in non-Finnish European individuals, of which 62 (32 PWS) were enriched by more than twofold in Finland. These findings demonstrate the power of bottlenecked populations to find entry points into the biology of common diseases through low-frequency, high impact variants., (© 2023. The Author(s).)- Published
- 2023
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13. Discovery and systematic characterization of risk variants and genes for coronary artery disease in over a million participants.
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Aragam KG, Jiang T, Goel A, Kanoni S, Wolford BN, Atri DS, Weeks EM, Wang M, Hindy G, Zhou W, Grace C, Roselli C, Marston NA, Kamanu FK, Surakka I, Venegas LM, Sherliker P, Koyama S, Ishigaki K, Åsvold BO, Brown MR, Brumpton B, de Vries PS, Giannakopoulou O, Giardoglou P, Gudbjartsson DF, Güldener U, Haider SMI, Helgadottir A, Ibrahim M, Kastrati A, Kessler T, Kyriakou T, Konopka T, Li L, Ma L, Meitinger T, Mucha S, Munz M, Murgia F, Nielsen JB, Nöthen MM, Pang S, Reinberger T, Schnitzler G, Smedley D, Thorleifsson G, von Scheidt M, Ulirsch JC, Arnar DO, Burtt NP, Costanzo MC, Flannick J, Ito K, Jang DK, Kamatani Y, Khera AV, Komuro I, Kullo IJ, Lotta LA, Nelson CP, Roberts R, Thorgeirsson G, Thorsteinsdottir U, Webb TR, Baras A, Björkegren JLM, Boerwinkle E, Dedoussis G, Holm H, Hveem K, Melander O, Morrison AC, Orho-Melander M, Rallidis LS, Ruusalepp A, Sabatine MS, Stefansson K, Zalloua P, Ellinor PT, Farrall M, Danesh J, Ruff CT, Finucane HK, Hopewell JC, Clarke R, Gupta RM, Erdmann J, Samani NJ, Schunkert H, Watkins H, Willer CJ, Deloukas P, Kathiresan S, and Butterworth AS
- Subjects
- Humans, Genome-Wide Association Study, Coronary Artery Disease genetics
- Abstract
The discovery of genetic loci associated with complex diseases has outpaced the elucidation of mechanisms of disease pathogenesis. Here we conducted a genome-wide association study (GWAS) for coronary artery disease (CAD) comprising 181,522 cases among 1,165,690 participants of predominantly European ancestry. We detected 241 associations, including 30 new loci. Cross-ancestry meta-analysis with a Japanese GWAS yielded 38 additional new loci. We prioritized likely causal variants using functionally informed fine-mapping, yielding 42 associations with less than five variants in the 95% credible set. Similarity-based clustering suggested roles for early developmental processes, cell cycle signaling and vascular cell migration and proliferation in the pathogenesis of CAD. We prioritized 220 candidate causal genes, combining eight complementary approaches, including 123 supported by three or more approaches. Using CRISPR-Cas9, we experimentally validated the effect of an enhancer in MYO9B, which appears to mediate CAD risk by regulating vascular cell motility. Our analysis identifies and systematically characterizes >250 risk loci for CAD to inform experimental interrogation of putative causal mechanisms for CAD., (© 2022. The Author(s).)
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- 2022
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14. Congenital anemia reveals distinct targeting mechanisms for master transcription factor GATA1.
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Ludwig LS, Lareau CA, Bao EL, Liu N, Utsugisawa T, Tseng AM, Myers SA, Verboon JM, Ulirsch JC, Luo W, Muus C, Fiorini C, Olive ME, Vockley CM, Munschauer M, Hunter A, Ogura H, Yamamoto T, Inada H, Nakagawa S, Ohzono S, Subramanian V, Chiarle R, Glader B, Carr SA, Aryee MJ, Kundaje A, Orkin SH, Regev A, McCavit TL, Kanno H, and Sankaran VG
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- Cell Differentiation genetics, Chromatin genetics, Chromatin Immunoprecipitation, Erythropoiesis genetics, Humans, Anemia, GATA1 Transcription Factor genetics, GATA1 Transcription Factor metabolism
- Abstract
Master regulators, such as the hematopoietic transcription factor (TF) GATA1, play an essential role in orchestrating lineage commitment and differentiation. However, the precise mechanisms by which such TFs regulate transcription through interactions with specific cis-regulatory elements remain incompletely understood. Here, we describe a form of congenital hemolytic anemia caused by missense mutations in an intrinsically disordered region of GATA1, with a poorly understood role in transcriptional regulation. Through integrative functional approaches, we demonstrate that these mutations perturb GATA1 transcriptional activity by partially impairing nuclear localization and selectively altering precise chromatin occupancy by GATA1. These alterations in chromatin occupancy and concordant chromatin accessibility changes alter faithful gene expression, with failure to both effectively silence and activate select genes necessary for effective terminal red cell production. We demonstrate how disease-causing mutations can reveal regulatory mechanisms that enable the faithful genomic targeting of master TFs during cellular differentiation., (© 2022 by The American Society of Hematology.)
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- 2022
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15. Author Correction: Direct characterization of cis-regulatory elements and functional dissection of complex genetic associations using HCR-FlowFISH.
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Reilly SK, Gosai SJ, Gutierrez A, Mackay-Smith A, Ulirsch JC, Kanai M, Mouri K, Berenzy D, Kales S, Butler GM, Gladden-Young A, Bhuiyan RM, Stitzel ML, Finucane HK, Sabeti PC, and Tewhey R
- Published
- 2021
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16. Genome-wide functional screen of 3'UTR variants uncovers causal variants for human disease and evolution.
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Griesemer D, Xue JR, Reilly SK, Ulirsch JC, Kukreja K, Davis JR, Kanai M, Yang DK, Butts JC, Guney MH, Luban J, Montgomery SB, Finucane HK, Novina CD, Tewhey R, and Sabeti PC
- Subjects
- Algorithms, Alleles, Gene Expression Regulation, Genes, Reporter, Genetic Variation, Humans, Phenotype, Polymorphism, Single Nucleotide genetics, Polyribosomes metabolism, Quantitative Trait Loci genetics, RNA genetics, 3' Untranslated Regions genetics, Biological Evolution, Disease genetics, Genome-Wide Association Study
- Abstract
3' untranslated region (3'UTR) variants are strongly associated with human traits and diseases, yet few have been causally identified. We developed the massively parallel reporter assay for 3'UTRs (MPRAu) to sensitively assay 12,173 3'UTR variants. We applied MPRAu to six human cell lines, focusing on genetic variants associated with genome-wide association studies (GWAS) and human evolutionary adaptation. MPRAu expands our understanding of 3'UTR function, suggesting that simple sequences predominately explain 3'UTR regulatory activity. We adapt MPRAu to uncover diverse molecular mechanisms at base pair resolution, including an adenylate-uridylate (AU)-rich element of LEPR linked to potential metabolic evolutionary adaptations in East Asians. We nominate hundreds of 3'UTR causal variants with genetically fine-mapped phenotype associations. Using endogenous allelic replacements, we characterize one variant that disrupts a miRNA site regulating the viral defense gene TRIM14 and one that alters PILRB abundance, nominating a causal variant underlying transcriptional changes in age-related macular degeneration., Competing Interests: Declaration of interests P.C.S. is a co-founder of and consultant to Sherlock Biosciences and Board Member of Danaher Corporation. She is a shareholder in both companies., (Copyright © 2021 Elsevier Inc. All rights reserved.)
- Published
- 2021
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17. Functional characterization of T2D-associated SNP effects on baseline and ER stress-responsive β cell transcriptional activation.
- Author
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Khetan S, Kales S, Kursawe R, Jillette A, Ulirsch JC, Reilly SK, Ucar D, Tewhey R, and Stitzel ML
- Subjects
- Alleles, Animals, Cell Line, Chromatin metabolism, Diabetes Mellitus, Type 2 pathology, Genome-Wide Association Study, Humans, Mice, Quantitative Trait Loci, Short Interspersed Nucleotide Elements genetics, Diabetes Mellitus, Type 2 genetics, Endoplasmic Reticulum Stress genetics, Insulin-Secreting Cells pathology, Polymorphism, Single Nucleotide, Transcriptional Activation genetics
- Abstract
Genome-wide association studies (GWAS) have linked single nucleotide polymorphisms (SNPs) at >250 loci in the human genome to type 2 diabetes (T2D) risk. For each locus, identifying the functional variant(s) among multiple SNPs in high linkage disequilibrium is critical to understand molecular mechanisms underlying T2D genetic risk. Using massively parallel reporter assays (MPRA), we test the cis-regulatory effects of SNPs associated with T2D and altered in vivo islet chromatin accessibility in MIN6 β cells under steady state and pathophysiologic endoplasmic reticulum (ER) stress conditions. We identify 1,982/6,621 (29.9%) SNP-containing elements that activate transcription in MIN6 and 879 SNP alleles that modulate MPRA activity. Multiple T2D-associated SNPs alter the activity of short interspersed nuclear element (SINE)-containing elements that are strongly induced by ER stress. We identify 220 functional variants at 104 T2D association signals, narrowing 54 signals to a single candidate SNP. Together, this study identifies elements driving β cell steady state and ER stress-responsive transcriptional activation, nominates causal T2D SNPs, and uncovers potential roles for repetitive elements in β cell transcriptional stress response and T2D genetics., (© 2021. The Author(s).)
- Published
- 2021
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18. Direct characterization of cis-regulatory elements and functional dissection of complex genetic associations using HCR-FlowFISH.
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Reilly SK, Gosai SJ, Gutierrez A, Mackay-Smith A, Ulirsch JC, Kanai M, Mouri K, Berenzy D, Kales S, Butler GM, Gladden-Young A, Bhuiyan RM, Stitzel ML, Finucane HK, Sabeti PC, and Tewhey R
- Subjects
- Adaptor Proteins, Signal Transducing genetics, Bayes Theorem, Clustered Regularly Interspaced Short Palindromic Repeats, Delta-5 Fatty Acid Desaturase, Deoxyribonuclease I genetics, Deoxyribonuclease I metabolism, Fatty Acid Desaturases genetics, Flow Cytometry, GATA1 Transcription Factor genetics, Humans, K562 Cells, LIM Domain Proteins genetics, Models, Genetic, Polymorphism, Single Nucleotide, Proto-Oncogene Proteins genetics, Quantitative Trait Loci, RNA, Guide, CRISPR-Cas Systems, In Situ Hybridization, Fluorescence methods, Regulatory Sequences, Nucleic Acid
- Abstract
Effective interpretation of genome function and genetic variation requires a shift from epigenetic mapping of cis-regulatory elements (CREs) to characterization of endogenous function. We developed hybridization chain reaction fluorescence in situ hybridization coupled with flow cytometry (HCR-FlowFISH), a broadly applicable approach to characterize CRISPR-perturbed CREs via accurate quantification of native transcripts, alongside CRISPR activity screen analysis (CASA), a hierarchical Bayesian model to quantify CRE activity. Across >325,000 perturbations, we provide evidence that CREs can regulate multiple genes, skip over the nearest gene and display activating and/or silencing effects. At the cholesterol-level-associated FADS locus, we combine endogenous screens with reporter assays to exhaustively characterize multiple genome-wide association signals, functionally nominate causal variants and, importantly, identify their target genes., (© 2021. The Author(s), under exclusive licence to Springer Nature America, Inc.)
- Published
- 2021
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19. Genome-wide enhancer maps link risk variants to disease genes.
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Nasser J, Bergman DT, Fulco CP, Guckelberger P, Doughty BR, Patwardhan TA, Jones TR, Nguyen TH, Ulirsch JC, Lekschas F, Mualim K, Natri HM, Weeks EM, Munson G, Kane M, Kang HY, Cui A, Ray JP, Eisenhaure TM, Collins RL, Dey K, Pfister H, Price AL, Epstein CB, Kundaje A, Xavier RJ, Daly MJ, Huang H, Finucane HK, Hacohen N, Lander ES, and Engreitz JM
- Subjects
- Cell Line, Chromosomes, Human, Pair 10 genetics, Cyclophilins genetics, Dendritic Cells, Female, Humans, Macrophages metabolism, Male, Mitochondria metabolism, Organ Specificity genetics, Phenotype, Enhancer Elements, Genetic genetics, Genetic Predisposition to Disease, Genetic Variation genetics, Genome, Human genetics, Genome-Wide Association Study, Inflammatory Bowel Diseases genetics
- Abstract
Genome-wide association studies (GWAS) have identified thousands of noncoding loci that are associated with human diseases and complex traits, each of which could reveal insights into the mechanisms of disease
1 . Many of the underlying causal variants may affect enhancers2,3 , but we lack accurate maps of enhancers and their target genes to interpret such variants. We recently developed the activity-by-contact (ABC) model to predict which enhancers regulate which genes and validated the model using CRISPR perturbations in several cell types4 . Here we apply this ABC model to create enhancer-gene maps in 131 human cell types and tissues, and use these maps to interpret the functions of GWAS variants. Across 72 diseases and complex traits, ABC links 5,036 GWAS signals to 2,249 unique genes, including a class of 577 genes that appear to influence multiple phenotypes through variants in enhancers that act in different cell types. In inflammatory bowel disease (IBD), causal variants are enriched in predicted enhancers by more than 20-fold in particular cell types such as dendritic cells, and ABC achieves higher precision than other regulatory methods at connecting noncoding variants to target genes. These variant-to-function maps reveal an enhancer that contains an IBD risk variant and that regulates the expression of PPIF to alter the membrane potential of mitochondria in macrophages. Our study reveals principles of genome regulation, identifies genes that affect IBD and provides a resource and generalizable strategy to connect risk variants of common diseases to their molecular and cellular functions.- Published
- 2021
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20. Tractor uses local ancestry to enable the inclusion of admixed individuals in GWAS and to boost power.
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Atkinson EG, Maihofer AX, Kanai M, Martin AR, Karczewski KJ, Santoro ML, Ulirsch JC, Kamatani Y, Okada Y, Finucane HK, Koenen KC, Nievergelt CM, Daly MJ, and Neale BM
- Subjects
- Cholesterol blood, Cholesterol genetics, Guanine Nucleotide Exchange Factors genetics, Haplotypes genetics, Homeodomain Proteins genetics, Humans, Lipids blood, Pedigree, Polymorphism, Single Nucleotide, Transcription Factors genetics, Black or African American genetics, Genome-Wide Association Study statistics & numerical data, Models, Genetic, Software, White People genetics
- Abstract
Admixed populations are routinely excluded from genomic studies due to concerns over population structure. Here, we present a statistical framework and software package, Tractor, to facilitate the inclusion of admixed individuals in association studies by leveraging local ancestry. We test Tractor with simulated and empirical two-way admixed African-European cohorts. Tractor generates accurate ancestry-specific effect-size estimates and P values, can boost genome-wide association study (GWAS) power and improves the resolution of association signals. Using a local ancestry-aware regression model, we replicate known hits for blood lipids, discover novel hits missed by standard GWAS and localize signals closer to putative causal variants.
- Published
- 2021
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21. Prioritizing disease and trait causal variants at the TNFAIP3 locus using functional and genomic features.
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Ray JP, de Boer CG, Fulco CP, Lareau CA, Kanai M, Ulirsch JC, Tewhey R, Ludwig LS, Reilly SK, Bergman DT, Engreitz JM, Issner R, Finucane HK, Lander ES, Regev A, and Hacohen N
- Subjects
- Cell Line, Tumor, Genetic Predisposition to Disease, Genetic Variation immunology, Haplotypes genetics, Haplotypes immunology, Humans, Linkage Disequilibrium, Multifactorial Inheritance immunology, Proof of Concept Study, Autoimmune Diseases genetics, Genetic Loci genetics, Genome-Wide Association Study methods, Multifactorial Inheritance genetics, Tumor Necrosis Factor alpha-Induced Protein 3 genetics
- Abstract
Genome-wide association studies have associated thousands of genetic variants with complex traits and diseases, but pinpointing the causal variant(s) among those in tight linkage disequilibrium with each associated variant remains a major challenge. Here, we use seven experimental assays to characterize all common variants at the multiple disease-associated TNFAIP3 locus in five disease-relevant immune cell lines, based on a set of features related to regulatory potential. Trait/disease-associated variants are enriched among SNPs prioritized based on either: (1) residing within CRISPRi-sensitive regulatory regions, or (2) localizing in a chromatin accessible region while displaying allele-specific reporter activity. Of the 15 trait/disease-associated haplotypes at TNFAIP3, 9 have at least one variant meeting one or both of these criteria, 5 of which are further supported by genetic fine-mapping. Our work provides a comprehensive strategy to characterize genetic variation at important disease-associated loci, and aids in the effort to identify trait causal genetic variants.
- Published
- 2020
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22. Control of human hemoglobin switching by LIN28B-mediated regulation of BCL11A translation.
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Basak A, Munschauer M, Lareau CA, Montbleau KE, Ulirsch JC, Hartigan CR, Schenone M, Lian J, Wang Y, Huang Y, Wu X, Gehrke L, Rice CM, An X, Christou HA, Mohandas N, Carr SA, Chen JJ, Orkin SH, Lander ES, and Sankaran VG
- Subjects
- Adult, Animals, Binding Sites, Cells, Cultured, Erythroid Cells metabolism, Erythropoiesis genetics, Gene Expression Regulation, Hemoglobins genetics, Humans, Infant, Newborn, MicroRNAs metabolism, Protein Biosynthesis, RNA, Messenger metabolism, RNA, Ribosomal, 18S metabolism, RNA-Binding Proteins genetics, Repressor Proteins metabolism, Ribosomes genetics, Ribosomes metabolism, Hemoglobins metabolism, RNA-Binding Proteins metabolism, Repressor Proteins genetics
- Abstract
Increased production of fetal hemoglobin (HbF) can ameliorate the severity of sickle cell disease and β-thalassemia
1 . BCL11A represses the genes encoding HbF and regulates human hemoglobin switching through variation in its expression during development2-7 . However, the mechanisms underlying the developmental expression of BCL11A remain mysterious. Here we show that BCL11A is regulated at the level of messenger RNA (mRNA) translation during human hematopoietic development. Despite decreased BCL11A protein synthesis earlier in development, BCL11A mRNA continues to be associated with ribosomes. Through unbiased genomic and proteomic analyses, we demonstrate that the RNA-binding protein LIN28B, which is developmentally expressed in a pattern reciprocal to that of BCL11A, directly interacts with ribosomes and BCL11A mRNA. Furthermore, we show that BCL11A mRNA translation is suppressed by LIN28B through direct interactions, independently of its role in regulating let-7 microRNAs, and that BCL11A is the major target of LIN28B-mediated HbF induction. Our results reveal a previously unappreciated mechanism underlying human hemoglobin switching that illuminates new therapeutic opportunities.- Published
- 2020
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23. X-linked macrocytic dyserythropoietic anemia in females with an ALAS2 mutation.
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Sankaran VG, Ulirsch JC, Tchaikovskii V, Ludwig LS, Wakabayashi A, Kadirvel S, Lindsley RC, Bejar R, Shi J, Lovitch SB, Bishop DF, and Steensma DP
- Published
- 2020
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24. Genetic predisposition to mosaic Y chromosome loss in blood.
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Thompson DJ, Genovese G, Halvardson J, Ulirsch JC, Wright DJ, Terao C, Davidsson OB, Day FR, Sulem P, Jiang Y, Danielsson M, Davies H, Dennis J, Dunlop MG, Easton DF, Fisher VA, Zink F, Houlston RS, Ingelsson M, Kar S, Kerrison ND, Kinnersley B, Kristjansson RP, Law PJ, Li R, Loveday C, Mattisson J, McCarroll SA, Murakami Y, Murray A, Olszewski P, Rychlicka-Buniowska E, Scott RA, Thorsteinsdottir U, Tomlinson I, Moghadam BT, Turnbull C, Wareham NJ, Gudbjartsson DF, Kamatani Y, Hoffmann ER, Jackson SP, Stefansson K, Auton A, Ong KK, Machiela MJ, Loh PR, Dumanski JP, Chanock SJ, Forsberg LA, and Perry JRB
- Subjects
- Adult, Aged, Computational Biology, Databases, Genetic, Female, Genetic Markers genetics, Humans, Male, Middle Aged, Neoplasms genetics, United Kingdom, Chromosome Deletion, Chromosomes, Human, Y genetics, Genetic Predisposition to Disease genetics, Genomic Instability genetics, Leukocytes pathology, Mosaicism
- Abstract
Mosaic loss of chromosome Y (LOY) in circulating white blood cells is the most common form of clonal mosaicism
1-5 , yet our knowledge of the causes and consequences of this is limited. Here, using a computational approach, we estimate that 20% of the male population represented in the UK Biobank study (n = 205,011) has detectable LOY. We identify 156 autosomal genetic determinants of LOY, which we replicate in 757,114 men of European and Japanese ancestry. These loci highlight genes that are involved in cell-cycle regulation and cancer susceptibility, as well as somatic drivers of tumour growth and targets of cancer therapy. We demonstrate that genetic susceptibility to LOY is associated with non-haematological effects on health in both men and women, which supports the hypothesis that clonal haematopoiesis is a biomarker of genomic instability in other tissues. Single-cell RNA sequencing identifies dysregulated expression of autosomal genes in leukocytes with LOY and provides insights into why clonal expansion of these cells may occur. Collectively, these data highlight the value of studying clonal mosaicism to uncover fundamental mechanisms that underlie cancer and other ageing-related diseases.- Published
- 2019
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25. Transcriptional States and Chromatin Accessibility Underlying Human Erythropoiesis.
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Ludwig LS, Lareau CA, Bao EL, Nandakumar SK, Muus C, Ulirsch JC, Chowdhary K, Buenrostro JD, Mohandas N, An X, Aryee MJ, Regev A, and Sankaran VG
- Subjects
- Aged, Cells, Cultured, Chromatin chemistry, Chromatin metabolism, Erythroid Precursor Cells cytology, Erythroid Precursor Cells metabolism, Female, GATA1 Transcription Factor genetics, GATA1 Transcription Factor metabolism, HEK293 Cells, Humans, Kruppel-Like Transcription Factors genetics, Kruppel-Like Transcription Factors metabolism, Membrane Proteins genetics, Membrane Proteins metabolism, Middle Aged, Polymorphism, Single Nucleotide, Chromatin Assembly and Disassembly, Erythropoiesis, Transcriptome
- Abstract
Human erythropoiesis serves as a paradigm of physiologic cellular differentiation. This process is also of considerable interest for better understanding anemias and identifying new therapies. Here, we apply deep transcriptomic and accessible chromatin profiling to characterize a faithful ex vivo human erythroid differentiation system from hematopoietic stem and progenitor cells. We reveal stage-specific transcriptional states and chromatin accessibility during various stages of erythropoiesis, including 14,260 differentially expressed genes and 63,659 variably accessible chromatin peaks. Our analysis suggests differentiation stage-predominant roles for specific master regulators, including GATA1 and KLF1. We integrate chromatin profiles with common and rare genetic variants associated with erythroid cell traits and diseases, finding that variants regulating different erythroid phenotypes likely act at variable points during differentiation. In addition, we identify a regulator of terminal erythropoiesis, TMCC2, more broadly illustrating the value of this comprehensive analysis to improve our understanding of erythropoiesis in health and disease., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)
- Published
- 2019
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26. Gene-centric functional dissection of human genetic variation uncovers regulators of hematopoiesis.
- Author
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Nandakumar SK, McFarland SK, Mateyka LM, Lareau CA, Ulirsch JC, Ludwig LS, Agarwal G, Engreitz JM, Przychodzen B, McConkey M, Cowley GS, Doench JG, Maciejewski JP, Ebert BL, Root DE, and Sankaran VG
- Subjects
- Erythrocytes metabolism, Erythrocytes pathology, Genome-Wide Association Study, Humans, Polymorphism, Single Nucleotide genetics, Genetic Diseases, Inborn, Genetic Predisposition to Disease, Hematopoiesis genetics, Quantitative Trait Loci genetics
- Abstract
Genome-wide association studies (GWAS) have identified thousands of variants associated with human diseases and traits. However, the majority of GWAS-implicated variants are in non-coding regions of the genome and require in depth follow-up to identify target genes and decipher biological mechanisms. Here, rather than focusing on causal variants, we have undertaken a pooled loss-of-function screen in primary hematopoietic cells to interrogate 389 candidate genes contained in 75 loci associated with red blood cell traits. Using this approach, we identify 77 genes at 38 GWAS loci, with most loci harboring 1-2 candidate genes. Importantly, the hit set was strongly enriched for genes validated through orthogonal genetic approaches. Genes identified by this approach are enriched in specific and relevant biological pathways, allowing regulators of human erythropoiesis and modifiers of blood diseases to be defined. More generally, this functional screen provides a paradigm for gene-centric follow up of GWAS for a variety of human diseases and traits., Competing Interests: SN, SM, LM, CL, JU, LL, GA, JE, BP, MM, GC, JD, JM, BE, DR, VS No competing interests declared, (© 2019, Nandakumar et al.)
- Published
- 2019
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27. Impaired human hematopoiesis due to a cryptic intronic GATA1 splicing mutation.
- Author
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Abdulhay NJ, Fiorini C, Verboon JM, Ludwig LS, Ulirsch JC, Zieger B, Lareau CA, Mi X, Roy A, Obeng EA, Erlacher M, Gupta N, Gabriel SB, Ebert BL, Niemeyer CM, Khoriaty RN, Ancliff P, Gazda HT, Wlodarski MW, and Sankaran VG
- Subjects
- Adult, Child, Exons, HEK293 Cells, Hematopoietic Stem Cells metabolism, Humans, Male, Myelodysplastic Syndromes pathology, RNA Splice Sites genetics, Transcription, Genetic genetics, Transfection, Alternative Splicing genetics, Anemia, Dyserythropoietic, Congenital genetics, GATA1 Transcription Factor genetics, Hematopoiesis genetics, Introns genetics, Mutation, Missense, Myelodysplastic Syndromes genetics
- Abstract
Studies of allelic variation underlying genetic blood disorders have provided important insights into human hematopoiesis. Most often, the identified pathogenic mutations result in loss-of-function or missense changes. However, assessing the pathogenicity of noncoding variants can be challenging. Here, we characterize two unrelated patients with a distinct presentation of dyserythropoietic anemia and other impairments in hematopoiesis associated with an intronic mutation in GATA1 that is 24 nucleotides upstream of the canonical splice acceptor site. Functional studies demonstrate that this single-nucleotide alteration leads to reduced canonical splicing and increased use of an alternative splice acceptor site that causes a partial intron retention event. The resultant altered GATA1 contains a five-amino acid insertion at the C-terminus of the C-terminal zinc finger and has no observable activity. Collectively, our results demonstrate how altered splicing of GATA1, which reduces levels of the normal form of this master transcription factor, can result in distinct changes in human hematopoiesis., (© 2019 Abdulhay et al.)
- Published
- 2019
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28. HRI coordinates translation necessary for protein homeostasis and mitochondrial function in erythropoiesis.
- Author
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Zhang S, Macias-Garcia A, Ulirsch JC, Velazquez J, Butty VL, Levine SS, Sankaran VG, and Chen JJ
- Subjects
- Activating Transcription Factor 4 genetics, Anemia, Iron-Deficiency, Animals, Cell Differentiation, Erythroblasts, Eukaryotic Initiation Factor-2 metabolism, GRB10 Adaptor Protein genetics, GRB10 Adaptor Protein metabolism, Mice, Mice, Knockout, Oxygen metabolism, Phosphorylation, Protein Biosynthesis, Ribosomal Proteins, Unfolded Protein Response, eIF-2 Kinase genetics, Erythropoiesis physiology, Heme metabolism, Iron metabolism, Mitochondria metabolism, Proteostasis physiology, eIF-2 Kinase metabolism
- Abstract
Iron and heme play central roles in the production of red blood cells, but the underlying mechanisms remain incompletely understood. Heme-regulated eIF2α kinase (HRI) controls translation by phosphorylating eIF2α. Here, we investigate the global impact of iron, heme, and HRI on protein translation in vivo in murine primary erythroblasts using ribosome profiling. We validate the known role of HRI-mediated translational stimulation of integratedstressresponse mRNAs during iron deficiency in vivo. Moreover, we find that the translation of mRNAs encoding cytosolic and mitochondrial ribosomal proteins is substantially repressed by HRI during iron deficiency, causing a decrease in cytosolic and mitochondrial protein synthesis. The absence of HRI during iron deficiency elicits a prominent cytoplasmic unfolded protein response and impairs mitochondrial respiration. Importantly, ATF4 target genes are activated during iron deficiency to maintain mitochondrial function and to enable erythroid differentiation. We further identify GRB10 as a previously unappreciated regulator of terminal erythropoiesis., Competing Interests: SZ, AM, JU, JV, VB, SL, VS, JC No competing interests declared, (© 2019, Zhang et al.)
- Published
- 2019
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29. Interrogation of human hematopoiesis at single-cell and single-variant resolution.
- Author
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Ulirsch JC, Lareau CA, Bao EL, Ludwig LS, Guo MH, Benner C, Satpathy AT, Kartha VK, Salem RM, Hirschhorn JN, Finucane HK, Aryee MJ, Buenrostro JD, and Sankaran VG
- Subjects
- Cell Lineage genetics, Chromatin genetics, Chromosome Mapping methods, Epigenomics methods, Genome-Wide Association Study methods, Humans, Linkage Disequilibrium genetics, Phenotype, Quantitative Trait Loci genetics, Regulatory Sequences, Nucleic Acid genetics, Hematopoiesis genetics, Polymorphism, Single Nucleotide genetics
- Abstract
Widespread linkage disequilibrium and incomplete annotation of cell-to-cell state variation represent substantial challenges to elucidating mechanisms of trait-associated genetic variation. Here we perform genetic fine-mapping for blood cell traits in the UK Biobank to identify putative causal variants. These variants are enriched in genes encoding proteins in trait-relevant biological pathways and in accessible chromatin of hematopoietic progenitors. For regulatory variants, we explore patterns of developmental enhancer activity, predict molecular mechanisms, and identify likely target genes. In several instances, we localize multiple independent variants to the same regulatory element or gene. We further observe that variants with pleiotropic effects preferentially act in common progenitor populations to direct the production of distinct lineages. Finally, we leverage fine-mapped variants in conjunction with continuous epigenomic annotations to identify trait-cell type enrichments within closely related populations and in single cells. Our study provides a comprehensive framework for single-variant and single-cell analyses of genetic associations.
- Published
- 2019
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30. Lineage Tracing in Humans Enabled by Mitochondrial Mutations and Single-Cell Genomics.
- Author
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Ludwig LS, Lareau CA, Ulirsch JC, Christian E, Muus C, Li LH, Pelka K, Ge W, Oren Y, Brack A, Law T, Rodman C, Chen JH, Boland GM, Hacohen N, Rozenblatt-Rosen O, Aryee MJ, Buenrostro JD, Regev A, and Sankaran VG
- Subjects
- Base Sequence, Cell Lineage, Chromatin, Colorectal Neoplasms genetics, Colorectal Neoplasms pathology, Genomics methods, HEK293 Cells, Hematopoietic Stem Cells physiology, High-Throughput Nucleotide Sequencing methods, Humans, Mutation, Single-Cell Analysis, Transposases, DNA, Mitochondrial genetics, Mitochondria genetics
- Abstract
Lineage tracing provides key insights into the fate of individual cells in complex organisms. Although effective genetic labeling approaches are available in model systems, in humans, most approaches require detection of nuclear somatic mutations, which have high error rates, limited scale, and do not capture cell state information. Here, we show that somatic mutations in mtDNA can be tracked by single-cell RNA or assay for transposase accessible chromatin (ATAC) sequencing. We leverage somatic mtDNA mutations as natural genetic barcodes and demonstrate their utility as highly accurate clonal markers to infer cellular relationships. We track native human cells both in vitro and in vivo and relate clonal dynamics to gene expression and chromatin accessibility. Our approach should allow clonal tracking at a 1,000-fold greater scale than with nuclear genome sequencing, with simultaneous information on cell state, opening the way to chart cellular dynamics in human health and disease., (Copyright © 2019 Elsevier Inc. All rights reserved.)
- Published
- 2019
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31. The Genetic Landscape of Diamond-Blackfan Anemia.
- Author
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Ulirsch JC, Verboon JM, Kazerounian S, Guo MH, Yuan D, Ludwig LS, Handsaker RE, Abdulhay NJ, Fiorini C, Genovese G, Lim ET, Cheng A, Cummings BB, Chao KR, Beggs AH, Genetti CA, Sieff CA, Newburger PE, Niewiadomska E, Matysiak M, Vlachos A, Lipton JM, Atsidaftos E, Glader B, Narla A, Gleizes PE, O'Donohue MF, Montel-Lehry N, Amor DJ, McCarroll SA, O'Donnell-Luria AH, Gupta N, Gabriel SB, MacArthur DG, Lander ES, Lek M, Da Costa L, Nathan DG, Korostelev AA, Do R, Sankaran VG, and Gazda HT
- Published
- 2019
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32. The Genetic Landscape of Diamond-Blackfan Anemia.
- Author
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Ulirsch JC, Verboon JM, Kazerounian S, Guo MH, Yuan D, Ludwig LS, Handsaker RE, Abdulhay NJ, Fiorini C, Genovese G, Lim ET, Cheng A, Cummings BB, Chao KR, Beggs AH, Genetti CA, Sieff CA, Newburger PE, Niewiadomska E, Matysiak M, Vlachos A, Lipton JM, Atsidaftos E, Glader B, Narla A, Gleizes PE, O'Donohue MF, Montel-Lehry N, Amor DJ, McCarroll SA, O'Donnell-Luria AH, Gupta N, Gabriel SB, MacArthur DG, Lander ES, Lek M, Da Costa L, Nathan DG, Korostelev AA, Do R, Sankaran VG, and Gazda HT
- Subjects
- Adolescent, Child, Child, Preschool, Cohort Studies, Exome genetics, Exons genetics, Female, Gene Deletion, Genetic Association Studies methods, Humans, Intercellular Signaling Peptides and Proteins genetics, Male, Mutation genetics, Phenotype, Ribosomal Proteins genetics, Ribosomes genetics, Sequence Analysis, RNA methods, Exome Sequencing methods, Anemia, Diamond-Blackfan genetics
- Abstract
Diamond-Blackfan anemia (DBA) is a rare bone marrow failure disorder that affects 7 out of 1,000,000 live births and has been associated with mutations in components of the ribosome. In order to characterize the genetic landscape of this heterogeneous disorder, we recruited a cohort of 472 individuals with a clinical diagnosis of DBA and performed whole-exome sequencing (WES). We identified relevant rare and predicted damaging mutations for 78% of individuals. The majority of mutations were singletons, absent from population databases, predicted to cause loss of function, and located in 1 of 19 previously reported ribosomal protein (RP)-encoding genes. Using exon coverage estimates, we identified and validated 31 deletions in RP genes. We also observed an enrichment for extended splice site mutations and validated their diverse effects using RNA sequencing in cell lines obtained from individuals with DBA. Leveraging the size of our cohort, we observed robust genotype-phenotype associations with congenital abnormalities and treatment outcomes. We further identified rare mutations in seven previously unreported RP genes that may cause DBA, as well as several distinct disorders that appear to phenocopy DBA, including nine individuals with biallelic CECR1 mutations that result in deficiency of ADA2. However, no new genes were identified at exome-wide significance, suggesting that there are no unidentified genes containing mutations readily identified by WES that explain >5% of DBA-affected case subjects. Overall, this report should inform not only clinical practice for DBA-affected individuals, but also the design and analysis of rare variant studies for heterogeneous Mendelian disorders., (Copyright © 2018 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)
- Published
- 2018
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33. Publisher Correction: The NORAD lncRNA assembles a topoisomerase complex critical for genome stability.
- Author
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Munschauer M, Nguyen CT, Sirokman K, Hartigan CR, Hogstrom L, Engreitz JM, Ulirsch JC, Fulco CP, Subramanian V, Chen J, Schenone M, Guttman M, Carr SA, and Lander ES
- Abstract
A typo in the 'Reviewer information' section of this Letter was corrected online.
- Published
- 2018
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34. Detecting genome-wide directional effects of transcription factor binding on polygenic disease risk.
- Author
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Reshef YA, Finucane HK, Kelley DR, Gusev A, Kotliar D, Ulirsch JC, Hormozdiari F, Nasser J, O'Connor L, van de Geijn B, Loh PR, Grossman SR, Bhatia G, Gazal S, Palamara PF, Pinello L, Patterson N, Adams RP, and Price AL
- Subjects
- Binding Sites genetics, Blood Cells metabolism, Blood Cells pathology, Blood Chemical Analysis, Gene Expression Regulation, Genetic Predisposition to Disease, Humans, Linkage Disequilibrium, Phenotype, Polymorphism, Single Nucleotide, Protein Binding, Risk Factors, Disease genetics, Genome-Wide Association Study, Multifactorial Inheritance genetics, Quantitative Trait Loci, Transcription Factors metabolism
- Abstract
Biological interpretation of genome-wide association study data frequently involves assessing whether SNPs linked to a biological process, for example, binding of a transcription factor, show unsigned enrichment for disease signal. However, signed annotations quantifying whether each SNP allele promotes or hinders the biological process can enable stronger statements about disease mechanism. We introduce a method, signed linkage disequilibrium profile regression, for detecting genome-wide directional effects of signed functional annotations on disease risk. We validate the method via simulations and application to molecular quantitative trait loci in blood, recovering known transcriptional regulators. We apply the method to expression quantitative trait loci in 48 Genotype-Tissue Expression tissues, identifying 651 transcription factor-tissue associations including 30 with robust evidence of tissue specificity. We apply the method to 46 diseases and complex traits (average n = 290 K), identifying 77 annotation-trait associations representing 12 independent transcription factor-trait associations, and characterize the underlying transcriptional programs using gene-set enrichment analyses. Our results implicate new causal disease genes and new disease mechanisms.
- Published
- 2018
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35. The NORAD lncRNA assembles a topoisomerase complex critical for genome stability.
- Author
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Munschauer M, Nguyen CT, Sirokman K, Hartigan CR, Hogstrom L, Engreitz JM, Ulirsch JC, Fulco CP, Subramanian V, Chen J, Schenone M, Guttman M, Carr SA, and Lander ES
- Subjects
- Binding Sites, Cell Cycle, Cell Cycle Proteins metabolism, Cell Nucleus metabolism, Cell Survival, Chromosome Segregation, DNA Damage, DNA Repair, DNA Repair Enzymes metabolism, DNA Replication, Heterogeneous-Nuclear Ribonucleoproteins metabolism, Humans, Mass Spectrometry, Nuclear Proteins metabolism, Protein Binding, RNA Splicing Factors metabolism, RNA, Long Noncoding genetics, Ribonucleoproteins metabolism, Transcription Factors metabolism, DNA Topoisomerases, Type I metabolism, Genomic Instability, Multiprotein Complexes chemistry, Multiprotein Complexes metabolism, RNA, Long Noncoding metabolism, RNA-Binding Proteins metabolism
- Abstract
The human genome contains thousands of long non-coding RNAs
1 , but specific biological functions and biochemical mechanisms have been discovered for only about a dozen2-7 . A specific long non-coding RNA-non-coding RNA activated by DNA damage (NORAD)-has recently been shown to be required for maintaining genomic stability8 , but its molecular mechanism is unknown. Here we combine RNA antisense purification and quantitative mass spectrometry to identify proteins that directly interact with NORAD in living cells. We show that NORAD interacts with proteins involved in DNA replication and repair in steady-state cells and localizes to the nucleus upon stimulation with replication stress or DNA damage. In particular, NORAD interacts with RBMX, a component of the DNA-damage response, and contains the strongest RBMX-binding site in the transcriptome. We demonstrate that NORAD controls the ability of RBMX to assemble a ribonucleoprotein complex-which we term NORAD-activated ribonucleoprotein complex 1 (NARC1)-that contains the known suppressors of genomic instability topoisomerase I (TOP1), ALYREF and the PRPF19-CDC5L complex. Cells depleted for NORAD or RBMX display an increased frequency of chromosome segregation defects, reduced replication-fork velocity and altered cell-cycle progression-which represent phenotypes that are mechanistically linked to TOP1 and PRPF19-CDC5L function. Expression of NORAD in trans can rescue defects caused by NORAD depletion, but rescue is significantly impaired when the RBMX-binding site in NORAD is deleted. Our results demonstrate that the interaction between NORAD and RBMX is important for NORAD function, and that NORAD is required for the assembly of the previously unknown topoisomerase complex NARC1, which contributes to maintaining genomic stability. In addition, we uncover a previously unknown function for long non-coding RNAs in modulating the ability of an RNA-binding protein to assemble a higher-order ribonucleoprotein complex.- Published
- 2018
- Full Text
- View/download PDF
36. Common α-globin variants modify hematologic and other clinical phenotypes in sickle cell trait and disease.
- Author
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Raffield LM, Ulirsch JC, Naik RP, Lessard S, Handsaker RE, Jain D, Kang HM, Pankratz N, Auer PL, Bao EL, Smith JD, Lange LA, Lange EM, Li Y, Thornton TA, Young BA, Abecasis GR, Laurie CC, Nickerson DA, McCarroll SA, Correa A, Wilson JG, Lettre G, Sankaran VG, and Reiner AP
- Subjects
- Adult, Black or African American, Anemia, Sickle Cell blood, Anemia, Sickle Cell physiopathology, Cohort Studies, DNA Copy Number Variations, Erythrocytes, Abnormal, Glomerular Filtration Rate, Glycated Hemoglobin metabolism, Humans, Phenotype, Young Adult, alpha-Thalassemia genetics, Anemia, Sickle Cell genetics, Hemoglobin, Sickle genetics, Sickle Cell Trait, alpha-Globins genetics
- Abstract
Co-inheritance of α-thalassemia has a significant protective effect on the severity of complications of sickle cell disease (SCD), including stroke. However, little information exists on the association and interactions for the common African ancestral α-thalassemia mutation (-α3.7 deletion) and β-globin traits (HbS trait [SCT] and HbC trait) on important clinical phenotypes such as red blood cell parameters, anemia, and chronic kidney disease (CKD). In a community-based cohort of 2,916 African Americans from the Jackson Heart Study, we confirmed the expected associations between SCT, HbC trait, and the -α3.7 deletion with lower mean corpuscular volume/mean corpuscular hemoglobin and higher red blood cell count and red cell distribution width. In addition to the recently recognized association of SCT with lower estimated glomerular filtration rate and glycated hemoglobin (HbA1c), we observed a novel association of the -α3.7 deletion with higher HbA1c levels. Co-inheritance of each additional copy of the -α3.7 deletion significantly lowered the risk of anemia and chronic kidney disease among individuals with SCT (P-interaction = 0.031 and 0.019, respectively). Furthermore, co-inheritance of a novel α-globin regulatory variant was associated with normalization of red cell parameters in individuals with the -α3.7 deletion and significantly negated the protective effect of α-thalassemia on stroke in 1,139 patients with sickle cell anemia from the Cooperative Study of Sickle Cell Disease (CSSCD) (P-interaction = 0.0049). Functional assays determined that rs11865131, located in the major alpha-globin enhancer MCS-R2, was the most likely causal variant. These findings suggest that common α- and β-globin variants interact to influence hematologic and clinical phenotypes in African Americans, with potential implications for risk-stratification and counseling of individuals with SCD and SCT.
- Published
- 2018
- Full Text
- View/download PDF
37. Ribosome Levels Selectively Regulate Translation and Lineage Commitment in Human Hematopoiesis.
- Author
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Khajuria RK, Munschauer M, Ulirsch JC, Fiorini C, Ludwig LS, McFarland SK, Abdulhay NJ, Specht H, Keshishian H, Mani DR, Jovanovic M, Ellis SR, Fulco CP, Engreitz JM, Schütz S, Lian J, Gripp KW, Weinberg OK, Pinkus GS, Gehrke L, Regev A, Lander ES, Gazda HT, Lee WY, Panse VG, Carr SA, and Sankaran VG
- Subjects
- 5' Untranslated Regions, Anemia, Diamond-Blackfan genetics, Apoptosis Regulatory Proteins antagonists & inhibitors, Apoptosis Regulatory Proteins genetics, Apoptosis Regulatory Proteins metabolism, Bone Marrow Cells metabolism, Cells, Cultured, Female, GATA1 Transcription Factor genetics, GATA1 Transcription Factor metabolism, Hematopoietic Stem Cells cytology, Hematopoietic Stem Cells metabolism, Humans, Male, Mutation, Missense, RNA Interference, RNA, Small Interfering metabolism, Ribosomal Proteins antagonists & inhibitors, Ribosomal Proteins genetics, Ribosomal Proteins metabolism, Ribosomes genetics, Transcription Factors genetics, Transcription Factors metabolism, Anemia, Diamond-Blackfan pathology, Ribosomes metabolism
- Abstract
Blood cell formation is classically thought to occur through a hierarchical differentiation process, although recent studies have shown that lineage commitment may occur earlier in hematopoietic stem and progenitor cells (HSPCs). The relevance to human blood diseases and the underlying regulation of these refined models remain poorly understood. By studying a genetic blood disorder, Diamond-Blackfan anemia (DBA), where the majority of mutations affect ribosomal proteins and the erythroid lineage is selectively perturbed, we are able to gain mechanistic insight into how lineage commitment is programmed normally and disrupted in disease. We show that in DBA, the pool of available ribosomes is limited, while ribosome composition remains constant. Surprisingly, this global reduction in ribosome levels more profoundly alters translation of a select subset of transcripts. We show how the reduced translation of select transcripts in HSPCs can impair erythroid lineage commitment, illuminating a regulatory role for ribosome levels in cellular differentiation., (Copyright © 2018 Elsevier Inc. All rights reserved.)
- Published
- 2018
- Full Text
- View/download PDF
38. Whole-exome sequencing identifies an α-globin cluster triplication resulting in increased clinical severity of β-thalassemia.
- Author
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Steinberg-Shemer O, Ulirsch JC, Noy-Lotan S, Krasnov T, Attias D, Dgany O, Laor R, Sankaran VG, and Tamary H
- Subjects
- Exome genetics, Genotype, Humans, Male, Mutation genetics, Exome Sequencing methods, Young Adult, alpha-Globins metabolism, beta-Globins genetics, DNA Copy Number Variations genetics, alpha-Globins genetics, beta-Thalassemia genetics
- Abstract
Whole-exome sequencing (WES) has been increasingly useful for the diagnosis of patients with rare causes of anemia, particularly when there is an atypical clinical presentation or targeted genotyping approaches are inconclusive. Here, we describe a 20-yr-old man with a lifelong moderate-to-severe anemia with accompanying splenomegaly who lacked a definitive diagnosis. After a thorough clinical workup and targeted genetic sequencing, we identified a paternally inherited β-globin mutation ( HBB :c.93-21G>A, IVS-I-110:G>A), a known cause of β-thalassemia minor. As this mutation alone was inconsistent with the severity of the anemia, we performed WES. Although we could not identify any relevant pathogenic single-nucleotide variants (SNVs) or small indels, copy-number variant (CNV) analyses revealed a likely triplication of the entire α-globin cluster, which was subsequently confirmed by multiplex ligation-dependent probe amplification. Treatment and follow-up was redefined according to the diagnosis of β-thalassemia intermedia resulting from a single β-thalassemia mutation in combination with an α-globin cluster triplication. Thus, we describe a case where the typical WES-based analysis of SNVs and small indels was unrevealing, but WES-based CNV analysis resulted in a definitive diagnosis that informed clinical decision-making. More generally, this case illustrates the value of performing CNV analysis when WES is otherwise unable to elucidate a clear genetic diagnosis., (© 2017 Steinberg-Shemer et al.; Published by Cold Spring Harbor Laboratory Press.)
- Published
- 2017
- Full Text
- View/download PDF
39. Developmentally-faithful and effective human erythropoiesis in immunodeficient and Kit mutant mice.
- Author
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Fiorini C, Abdulhay NJ, McFarland SK, Munschauer M, Ulirsch JC, Chiarle R, and Sankaran VG
- Subjects
- Animals, Heterografts, Humans, Mice, Mice, Inbred NOD, Mice, SCID, Proto-Oncogene Proteins c-kit genetics, Erythropoiesis, Hematopoietic Stem Cell Transplantation, Mutation, Proto-Oncogene Proteins c-kit metabolism
- Abstract
Immunodeficient mouse models have been valuable for studies of human hematopoiesis, but high-fidelity recapitulation of erythropoiesis in most xenograft recipients remains elusive. Recently developed immunodeficient and Kit mutant mice, however, have provided a suitable background to achieve higher-level human erythropoiesis after long-term hematopoietic engraftment. While there has been some characterization of human erythropoiesis in these models, a comprehensive analysis from various human developmental stages has not yet been reported. Here, we have utilized cell surface phenotypes, morphologic analyses, and molecular studies to fully characterize human erythropoiesis from multiple developmental stages in immunodeficient and Kit mutant mouse models following long-term hematopoietic stem and progenitor cell engraftment. We show that human erythropoiesis in such models demonstrates complete maturation and enucleation, as well as developmentally appropriate globin gene expression. These results provide a framework for future studies to utilize this model system for interrogating disorders affecting human erythropoiesis and for developing improved therapeutic approaches., (© 2017 Wiley Periodicals, Inc.)
- Published
- 2017
- Full Text
- View/download PDF
40. Confounding in ex vivo models of Diamond-Blackfan anemia.
- Author
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Ulirsch JC, Lareau C, Ludwig LS, Mohandas N, Nathan DG, and Sankaran VG
- Subjects
- Humans, Ribosomal Proteins, Anemia, Diamond-Blackfan, Fanconi Anemia
- Published
- 2017
- Full Text
- View/download PDF
41. Functional Selectivity in Cytokine Signaling Revealed Through a Pathogenic EPO Mutation.
- Author
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Kim AR, Ulirsch JC, Wilmes S, Unal E, Moraga I, Karakukcu M, Yuan D, Kazerounian S, Abdulhay NJ, King DS, Gupta N, Gabriel SB, Lander ES, Patiroglu T, Ozcan A, Ozdemir MA, Garcia KC, Piehler J, Gazda HT, Klein DE, and Sankaran VG
- Subjects
- Anemia, Diamond-Blackfan therapy, Child, Consanguinity, Enzyme Activation, Erythropoiesis, Erythropoietin chemistry, Female, Humans, Janus Kinase 2 metabolism, Kinetics, Male, Receptors, Erythropoietin chemistry, Receptors, Erythropoietin genetics, Receptors, Erythropoietin metabolism, Anemia, Diamond-Blackfan genetics, Anemia, Diamond-Blackfan pathology, Erythropoietin genetics, Mutation, Missense, Signal Transduction
- Abstract
Cytokines are classically thought to stimulate downstream signaling pathways through monotonic activation of receptors. We describe a severe anemia resulting from a homozygous mutation (R150Q) in the cytokine erythropoietin (EPO). Surprisingly, the EPO R150Q mutant shows only a mild reduction in affinity for its receptor but has altered binding kinetics. The EPO mutant is less effective at stimulating erythroid cell proliferation and differentiation, even at maximally potent concentrations. While the EPO mutant can stimulate effectors such as STAT5 to a similar extent as the wild-type ligand, there is reduced JAK2-mediated phosphorylation of select downstream targets. This impairment in downstream signaling mechanistically arises from altered receptor dimerization dynamics due to extracellular binding changes. These results demonstrate how variation in a single cytokine can lead to biased downstream signaling and can thereby cause human disease. Moreover, we have defined a distinct treatable form of anemia through mutation identification and functional studies., (Copyright © 2017 Elsevier Inc. All rights reserved.)
- Published
- 2017
- Full Text
- View/download PDF
42. Comprehensive population-based genome sequencing provides insight into hematopoietic regulatory mechanisms.
- Author
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Guo MH, Nandakumar SK, Ulirsch JC, Zekavat SM, Buenrostro JD, Natarajan P, Salem RM, Chiarle R, Mitt M, Kals M, Pärn K, Fischer K, Milani L, Mägi R, Palta P, Gabriel SB, Metspalu A, Lander ES, Kathiresan S, Hirschhorn JN, Esko T, and Sankaran VG
- Subjects
- Base Sequence, Basophils cytology, Cell Differentiation genetics, Cell Lineage genetics, Chromosome Mapping, Databases, Nucleic Acid, Enhancer Elements, Genetic, Epigenesis, Genetic, Estonia, Female, GATA2 Transcription Factor genetics, Gene Expression Regulation, Developmental, Genome-Wide Association Study, Humans, Leukocyte Count, Male, Polymorphism, Single Nucleotide, Whole Genome Sequencing, CCAAT-Enhancer-Binding Proteins genetics, Hematopoiesis genetics
- Abstract
Genetic variants affecting hematopoiesis can influence commonly measured blood cell traits. To identify factors that affect hematopoiesis, we performed association studies for blood cell traits in the population-based Estonian Biobank using high-coverage whole-genome sequencing (WGS) in 2,284 samples and SNP genotyping in an additional 14,904 samples. Using up to 7,134 samples with available phenotype data, our analyses identified 17 associations across 14 blood cell traits. Integration of WGS-based fine-mapping and complementary epigenomic datasets provided evidence for causal mechanisms at several loci, including at a previously undiscovered basophil count-associated locus near the master hematopoietic transcription factor CEBPA The fine-mapped variant at this basophil count association near CEBPA overlapped an enhancer active in common myeloid progenitors and influenced its activity. In situ perturbation of this enhancer by CRISPR/Cas9 mutagenesis in hematopoietic stem and progenitor cells demonstrated that it is necessary for and specifically regulates CEBPA expression during basophil differentiation. We additionally identified basophil count-associated variation at another more pleiotropic myeloid enhancer near GATA2, highlighting regulatory mechanisms for ordered expression of master hematopoietic regulators during lineage specification. Our study illustrates how population-based genetic studies can provide key insights into poorly understood cell differentiation processes of considerable physiologic relevance., Competing Interests: The authors declare no conflict of interest.
- Published
- 2017
- Full Text
- View/download PDF
43. A novel pathogenic mutation in RPL11 identified in a patient diagnosed with diamond Blackfan anemia as a young adult.
- Author
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Narla A, Yuan D, Kazerounian S, LaVasseur C, Ulirsch JC, Narla J, Glader B, Sankaran VG, and Gazda H
- Subjects
- Adult, Anemia, Diamond-Blackfan blood, Humans, Male, Ribosomal Proteins metabolism, Young Adult, Anemia, Diamond-Blackfan diagnosis, Anemia, Diamond-Blackfan genetics, Ribosomal Proteins genetics
- Published
- 2016
- Full Text
- View/download PDF
44. Exome sequencing results in successful diagnosis and treatment of a severe congenital anemia.
- Author
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Lacy JN, Ulirsch JC, Grace RF, Towne MC, Hale J, Mohandas N, Lux SE 4th, Agrawal PB, and Sankaran VG
- Abstract
Whole-exome sequencing is increasingly used for diagnosis and identification of appropriate therapies in patients. Here, we present the case of a 3-yr-old male with a lifelong and severe transfusion-dependent anemia of unclear etiology, despite an extensive clinical workup. Given the difficulty of making the diagnosis and the potential side effects from performing interventions in patients with a congenital anemia of unknown etiology, we opted to perform whole-exome sequencing on the patient and his parents. This resulted in the identification of homozygous loss-of-function mutations in the EPB41 gene, encoding erythrocyte protein band 4.1, which therefore causes a rare and severe form of hereditary elliptocytosis in the patient. Based on prior clinical experience in similar patients, a surgical splenectomy was performed that resulted in subsequent transfusion independence in the patient. This case illustrates how whole-exome sequencing can lead to accurate diagnoses (and exclusion of diagnoses where interventions, such as splenectomy, would be contraindicated), thereby resulting in appropriate and successful therapeutic intervention-a major goal of precision medicine.
- Published
- 2016
- Full Text
- View/download PDF
45. Systematic Functional Dissection of Common Genetic Variation Affecting Red Blood Cell Traits.
- Author
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Ulirsch JC, Nandakumar SK, Wang L, Giani FC, Zhang X, Rogov P, Melnikov A, McDonel P, Do R, Mikkelsen TS, and Sankaran VG
- Subjects
- Alternative Splicing, Cell Line, Cell Lineage genetics, Erythropoiesis genetics, Gene Library, Genes, Reporter, Humans, Regulatory Sequences, Nucleic Acid, Transcription, Genetic, Erythrocytes, Genetic Techniques, Genetic Variation
- Abstract
Genome-wide association studies (GWAS) have successfully identified thousands of associations between common genetic variants and human disease phenotypes, but the majority of these variants are non-coding, often requiring genetic fine-mapping, epigenomic profiling, and individual reporter assays to delineate potential causal variants. We employ a massively parallel reporter assay (MPRA) to simultaneously screen 2,756 variants in strong linkage disequilibrium with 75 sentinel variants associated with red blood cell traits. We show that this assay identifies elements with endogenous erythroid regulatory activity. Across 23 sentinel variants, we conservatively identified 32 MPRA functional variants (MFVs). We used targeted genome editing to demonstrate endogenous enhancer activity across 3 MFVs that predominantly affect the transcription of SMIM1, RBM38, and CD164. Functional follow-up of RBM38 delineates a key role for this gene in the alternative splicing program occurring during terminal erythropoiesis. Finally, we provide evidence for how common GWAS-nominated variants can disrupt cell-type-specific transcriptional regulatory pathways., (Copyright © 2016 Elsevier Inc. All rights reserved.)
- Published
- 2016
- Full Text
- View/download PDF
46. Insight into GATA1 transcriptional activity through interrogation of cis elements disrupted in human erythroid disorders.
- Author
-
Wakabayashi A, Ulirsch JC, Ludwig LS, Fiorini C, Yasuda M, Choudhuri A, McDonel P, Zon LI, and Sankaran VG
- Subjects
- Anemia, Diamond-Blackfan genetics, Basic Helix-Loop-Helix Transcription Factors genetics, Basic Helix-Loop-Helix Transcription Factors metabolism, CRISPR-Cas Systems, GATA1 Transcription Factor genetics, Humans, K562 Cells, Nucleotide Motifs, Proto-Oncogene Proteins genetics, Proto-Oncogene Proteins metabolism, T-Cell Acute Lymphocytic Leukemia Protein 1, Anemia, Diamond-Blackfan metabolism, GATA1 Transcription Factor metabolism, Mutation, Response Elements, Transcription, Genetic
- Abstract
Whole-exome sequencing has been incredibly successful in identifying causal genetic variants and has revealed a number of novel genes associated with blood and other diseases. One limitation of this approach is that it overlooks mutations in noncoding regulatory elements. Furthermore, the mechanisms by which mutations in transcriptionalcis-regulatory elements result in disease remain poorly understood. Here we used CRISPR/Cas9 genome editing to interrogate three such elements harboring mutations in human erythroid disorders, which in all cases are predicted to disrupt a canonical binding motif for the hematopoietic transcription factor GATA1. Deletions of as few as two to four nucleotides resulted in a substantial decrease (>80%) in target gene expression. Isolated deletions of the canonical GATA1 binding motif completely abrogated binding of the cofactor TAL1, which binds to a separate motif. Having verified the functionality of these three GATA1 motifs, we demonstrate strong evolutionary conservation of GATA1 motifs in regulatory elements proximal to other genes implicated in erythroid disorders, and show that targeted disruption of such elements results in altered gene expression. By modeling transcription factor binding patterns, we show that multiple transcription factors are associated with erythroid gene expression, and have created predictive maps modeling putative disruptions of their binding sites at key regulatory elements. Our study provides insight into GATA1 transcriptional activity and may prove a useful resource for investigating the pathogenicity of noncoding variants in human erythroid disorders.
- Published
- 2016
- Full Text
- View/download PDF
47. Advances in understanding erythropoiesis: evolving perspectives.
- Author
-
Nandakumar SK, Ulirsch JC, and Sankaran VG
- Subjects
- Animals, Cell Communication physiology, Cell Differentiation physiology, Embryonic Development physiology, Erythropoiesis genetics, Hematologic Diseases physiopathology, Hematopoietic Stem Cells physiology, Hemoglobins physiology, Heterografts physiology, Humans, Mice, Models, Biological, Transcription, Genetic physiology, Zebrafish, Erythropoiesis physiology
- Abstract
Red blood cells (RBCs) are generated from haematopoietic stem and progenitor cells (HSPCs) through the step-wise process of differentiation known as erythropoiesis. In this review, we discuss our current understanding of erythropoiesis and highlight recent advances in this field. During embryonic development, erythropoiesis occurs in three distinct waves comprising first, the yolk sac-derived primitive RBCs, followed sequentially by the erythro-myeloid progenitor (EMP) and HSPC-derived definitive RBCs. Recent work has highlighted the complexity and variability that may exist in the hierarchical arrangement of progenitors responsible for erythropoiesis. Using recently defined cell surface markers, it is now possible to enrich for erythroid progenitors and precursors to a much greater extent than has been possible before. While a great deal of knowledge has been gained on erythropoiesis from model organisms, our understanding of this process is currently being refined through human genetic studies. Genes mutated in erythroid disorders can now be identified more rapidly by the use of next-generation sequencing techniques. Genome-wide association studies on erythroid traits in healthy populations have also revealed new modulators of erythropoiesis. All of these recent developments have significant promise not only for increasing our understanding of erythropoiesis, but also for improving our ability to intervene when RBC production is perturbed in disease., (© 2016 John Wiley & Sons Ltd.)
- Published
- 2016
- Full Text
- View/download PDF
48. Targeted Application of Human Genetic Variation Can Improve Red Blood Cell Production from Stem Cells.
- Author
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Giani FC, Fiorini C, Wakabayashi A, Ludwig LS, Salem RM, Jobaliya CD, Regan SN, Ulirsch JC, Liang G, Steinberg-Shemer O, Guo MH, Esko T, Tong W, Brugnara C, Hirschhorn JN, Weiss MJ, Zon LI, Chou ST, French DL, Musunuru K, and Sankaran VG
- Subjects
- CRISPR-Cas Systems, Cell Differentiation, Cytokines metabolism, Embryonic Stem Cells cytology, Fetal Blood cytology, Genetic Techniques, Genetic Variation, Genome, Human, Hematopoietic Stem Cells cytology, Hemoglobins analysis, Humans, Mutation, Pluripotent Stem Cells cytology, Regenerative Medicine methods, Erythrocytes cytology, Stem Cells cytology
- Abstract
Multipotent and pluripotent stem cells are potential sources for cell and tissue replacement therapies. For example, stem cell-derived red blood cells (RBCs) are a potential alternative to donated blood, but yield and quality remain a challenge. Here, we show that application of insight from human population genetic studies can enhance RBC production from stem cells. The SH2B3 gene encodes a negative regulator of cytokine signaling and naturally occurring loss-of-function variants in this gene increase RBC counts in vivo. Targeted suppression of SH2B3 in primary human hematopoietic stem and progenitor cells enhanced the maturation and overall yield of in-vitro-derived RBCs. Moreover, inactivation of SH2B3 by CRISPR/Cas9 genome editing in human pluripotent stem cells allowed enhanced erythroid cell expansion with preserved differentiation. Our findings therefore highlight the potential for combining human genome variation studies with genome editing approaches to improve cell and tissue production for regenerative medicine., (Copyright © 2016 Elsevier Inc. All rights reserved.)
- Published
- 2016
- Full Text
- View/download PDF
49. Association of Epidemiologic Factors and Genetic Variants Influencing Hypothalamic-Pituitary-Adrenocortical Axis Function With Postconcussive Symptoms After Minor Motor Vehicle Collision.
- Author
-
Auvergne L, Bortsov AV, Ulirsch JC, Peak DA, Jones JS, Swor RA, Domeier RM, Lee DC, Rathlev NK, Hendry PL, and McLean SA
- Subjects
- Accidents, Traffic, Adolescent, Adult, Aged, Craniocerebral Trauma epidemiology, Craniocerebral Trauma etiology, Emergency Service, Hospital, Female, Genotype, Humans, Male, Middle Aged, Pain epidemiology, Pain etiology, Post-Concussion Syndrome epidemiology, Post-Concussion Syndrome physiopathology, Post-Concussion Syndrome psychology, Stress, Psychological epidemiology, Stress, Psychological etiology, Tacrolimus Binding Proteins blood, Young Adult, Hypothalamo-Hypophyseal System physiopathology, Pituitary-Adrenal System physiopathology, Polymorphism, Single Nucleotide, Post-Concussion Syndrome genetics, Tacrolimus Binding Proteins genetics
- Abstract
Objectives: To determine the influence of epidemiologic factors and the influence of genetic variants affecting FKBP5, a protein known to modulate hypothalamic-pituitary-adrenocortical axis function, on the severity of somatic symptoms commonly termed "postconcussive" 6 and 12 months after motor vehicle collision (MVC)., Methods: European Americans 18 to 65 years of age who presented to one of eight emergency departments (EDs) after MVC were enrolled. Exclusion criteria included hospital admission. Blood samples were collected in the ED for genotyping. Participants completed evaluations including an adapted Rivermead Post-Concussive Symptoms Questionnaire in the ED and at 6 weeks, 6 months, and 1 year. Repeated-measures analysis of covariance was used to evaluate the association between epidemiologic factors (sociodemographic, pre-MVC health, collision characteristics, head injury, peritraumatic pain, and stress), FKBP5 genetic variants, and postconcussive symptom severity., Results: Among 943 patients recruited in the ED, follow-up was completed on 835 (88%) at 6 months and 857 (90%) at 1 year. Self-reported head impact during collision was not associated with chronic postconcussive symptom severity. After correction for multiple testing, three FKBP5 single-nucleotide polymorphisms (rs3800373, rs7753746, and rs9380526) predicted chronic postconcussive symptom severity, with an average symptom severity of 1.10 (95% confidence interval = 0.96-1.24), 1.36 (1.21-1.51), and 1.55 (1.23-1.88) for one, two, or three copies of minor allele at rs3800373 (p = .001). Similar effect sizes were observed for the minor alleles of rs7753746 and rs9380526., Conclusions: Postconcussive symptoms after minor MVC are not generally related to the severity of mild brain injury. This study shows that neurobiologic stress systems may play a role in the pathogenesis of postconcussive symptoms.
- Published
- 2016
- Full Text
- View/download PDF
50. Inducible Gata1 suppression expands megakaryocyte-erythroid progenitors from embryonic stem cells.
- Author
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Noh JY, Gandre-Babbe S, Wang Y, Hayes V, Yao Y, Gadue P, Sullivan SK, Chou ST, Machlus KR, Italiano JE Jr, Kyba M, Finkelstein D, Ulirsch JC, Sankaran VG, French DL, Poncz M, and Weiss MJ
- Subjects
- Animals, Cell Differentiation genetics, Embryonic Stem Cells cytology, GATA1 Transcription Factor genetics, Gene Expression Regulation drug effects, Gene Expression Regulation genetics, Humans, Megakaryocyte-Erythroid Progenitor Cells cytology, Megakaryocytes cytology, Megakaryocytes metabolism, Mice, Anti-Bacterial Agents pharmacology, Cell Differentiation drug effects, Doxycycline pharmacology, Embryonic Stem Cells metabolism, GATA1 Transcription Factor biosynthesis, Megakaryocyte-Erythroid Progenitor Cells metabolism, Thrombopoietin pharmacology
- Abstract
Transfusion of donor-derived platelets is commonly used for thrombocytopenia, which results from a variety of clinical conditions and relies on a constant donor supply due to the limited shelf life of these cells. Embryonic stem (ES) and induced pluripotent stem (iPS) cells represent a potential source of megakaryocytes and platelets for transfusion therapies; however, the majority of current ES/iPS cell differentiation protocols are limited by low yields of hematopoietic progeny. In both mice and humans, mutations in the gene-encoding transcription factor GATA1 cause an accumulation of proliferating, developmentally arrested megakaryocytes, suggesting that GATA1 suppression in ES and iPS cell-derived hematopoietic progenitors may enhance megakaryocyte production. Here, we engineered ES cells from WT mice to express a doxycycline-regulated (dox-regulated) shRNA that targets Gata1 transcripts for degradation. Differentiation of these cells in the presence of dox and thrombopoietin (TPO) resulted in an exponential (at least 10¹³-fold) expansion of immature hematopoietic progenitors. Dox withdrawal in combination with multilineage cytokines restored GATA1 expression, resulting in differentiation into erythroblasts and megakaryocytes. Following transfusion into recipient animals, these dox-deprived mature megakaryocytes generated functional platelets. Our findings provide a readily reproducible strategy to exponentially expand ES cell-derived megakaryocyte-erythroid progenitors that have the capacity to differentiate into functional platelet-producing megakaryocytes.
- Published
- 2015
- Full Text
- View/download PDF
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