37 results on '"Reilly SK"'
Search Results
2. Transmission from vaccinated individuals in a large SARS-CoV-2 Delta variant outbreak
- Author
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Siddle, KJ, Krasilnikova, LA, Moreno, GK, Schaffner, SF, Vostok, J, Fitzgerald, NA, Lemieux, JE, Barkas, N, Loreth, C, Specht, I, Tomkins-Tinch, CH, Paull, JS, Schaeffer, B, Taylor, BP, Loftness, B, Johnson, H, Schubert, PL, Shephard, HM, Doucette, M, Fink, T, Lang, AS, Baez, S, Beauchamp, J, Hennigan, S, Buzby, E, Ash, S, Brown, J, Clancy, S, Cofsky, S, Gagne, L, Hall, J, Harrington, R, Gionet, GL, DeRuff, KC, Vodzak, ME, Adams, GC, Dobbins, ST, Slack, SD, Reilly, SK, Anderson, LM, Cipicchio, MC, DeFelice, MT, Grimsby, JL, Anderson, SE, Blumenstiel, BS, Meldrim, JC, Rooke, HM, Vicente, G, Smith, NL, Messer, KS, Reagan, FL, Mandese, ZM, Lee, MD, Ray, MC, Fisher, ME, Ulcena, MA, Nolet, CM, English, SE, Larkin, KL, Vernest, K, Chaluvadi, S, Arvidson, D, Melchiono, M, Covell, T, Harik, V, Brock-Fisher, T, Dunn, M, Kearns, A, Hanage, WP, Bernard, C, Philippakis, A, Lennon, NJ, Gabriel, SB, Gallagher, GR, Smole, S, Madoff, LC, Brown, CM, Park, DJ, MacInnis, BL, Sabeti, PC, Siddle, KJ, Krasilnikova, LA, Moreno, GK, Schaffner, SF, Vostok, J, Fitzgerald, NA, Lemieux, JE, Barkas, N, Loreth, C, Specht, I, Tomkins-Tinch, CH, Paull, JS, Schaeffer, B, Taylor, BP, Loftness, B, Johnson, H, Schubert, PL, Shephard, HM, Doucette, M, Fink, T, Lang, AS, Baez, S, Beauchamp, J, Hennigan, S, Buzby, E, Ash, S, Brown, J, Clancy, S, Cofsky, S, Gagne, L, Hall, J, Harrington, R, Gionet, GL, DeRuff, KC, Vodzak, ME, Adams, GC, Dobbins, ST, Slack, SD, Reilly, SK, Anderson, LM, Cipicchio, MC, DeFelice, MT, Grimsby, JL, Anderson, SE, Blumenstiel, BS, Meldrim, JC, Rooke, HM, Vicente, G, Smith, NL, Messer, KS, Reagan, FL, Mandese, ZM, Lee, MD, Ray, MC, Fisher, ME, Ulcena, MA, Nolet, CM, English, SE, Larkin, KL, Vernest, K, Chaluvadi, S, Arvidson, D, Melchiono, M, Covell, T, Harik, V, Brock-Fisher, T, Dunn, M, Kearns, A, Hanage, WP, Bernard, C, Philippakis, A, Lennon, NJ, Gabriel, SB, Gallagher, GR, Smole, S, Madoff, LC, Brown, CM, Park, DJ, MacInnis, BL, and Sabeti, PC
- Abstract
An outbreak of over 1,000 COVID-19 cases in Provincetown, Massachusetts (MA), in July 2021-the first large outbreak mostly in vaccinated individuals in the US-prompted a comprehensive public health response, motivating changes to national masking recommendations and raising questions about infection and transmission among vaccinated individuals. To address these questions, we combined viral genomic and epidemiological data from 467 individuals, including 40% of outbreak-associated cases. The Delta variant accounted for 99% of cases in this dataset; it was introduced from at least 40 sources, but 83% of cases derived from a single source, likely through transmission across multiple settings over a short time rather than a single event. Genomic and epidemiological data supported multiple transmissions of Delta from and between fully vaccinated individuals. However, despite its magnitude, the outbreak had limited onward impact in MA and the US overall, likely due to high vaccination rates and a robust public health response.
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- 2022
3. HCR-FlowFISH: A flexible CRISPR screening method to identify cis-regulatory elements and their target genes
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Reilly, SK, primary, Gosai, SJ, additional, Gutierrez, A, additional, Ulirsch, JC, additional, Kanai, M, additional, Berenzy, D, additional, Kales, S, additional, Butler, GB, additional, Gladden-Young, A, additional, Finucane, HK, additional, Sabeti, PC, additional, and Tewhey, R, additional
- Published
- 2020
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4. Massively parallel approaches for characterizing noncoding functional variation in human evolution.
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Rong S, Root E, and Reilly SK
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- Humans, Genetic Variation, Animals, Regulatory Sequences, Nucleic Acid genetics, Phenotype, Machine Learning, Evolution, Molecular, Genome, Human
- Abstract
The genetic differences underlying unique phenotypes in humans compared to our closest primate relatives have long remained a mystery. Similarly, the genetic basis of adaptations between human groups during our expansion across the globe is poorly characterized. Uncovering the downstream phenotypic consequences of these genetic variants has been difficult, as a substantial portion lies in noncoding regions, such as cis-regulatory elements (CREs). Here, we review recent high-throughput approaches to measure the functions of CREs and the impact of variation within them. CRISPR screens can directly perturb CREs in the genome to understand downstream impacts on gene expression and phenotypes, while massively parallel reporter assays can decipher the regulatory impact of sequence variants. Machine learning has begun to be able to predict regulatory function from sequence alone, further scaling our ability to characterize genome function. Applying these tools across diverse phenotypes, model systems, and ancestries is beginning to revolutionize our understanding of noncoding variation underlying human evolution., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)
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- 2024
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5. 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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6. Multicenter integrated analysis of noncoding CRISPRi screens.
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Yao D, Tycko J, Oh JW, Bounds LR, Gosai SJ, Lataniotis L, Mackay-Smith A, Doughty BR, Gabdank I, Schmidt H, Guerrero-Altamirano T, Siklenka K, Guo K, White AD, Youngworth I, Andreeva K, Ren X, Barrera A, Luo Y, Yardımcı GG, Tewhey R, Kundaje A, Greenleaf WJ, Sabeti PC, Leslie C, Pritykin Y, Moore JE, Beer MA, Gersbach CA, Reddy TE, Shen Y, Engreitz JM, Bassik MC, and Reilly SK
- Subjects
- Humans, Genome, K562 Cells, RNA, Guide, CRISPR-Cas Systems, Clustered Regularly Interspaced Short Palindromic Repeats genetics, CRISPR-Cas Systems genetics
- Abstract
The ENCODE Consortium's efforts to annotate noncoding cis-regulatory elements (CREs) have advanced our understanding of gene regulatory landscapes. Pooled, noncoding CRISPR screens offer a systematic approach to investigate cis-regulatory mechanisms. The ENCODE4 Functional Characterization Centers conducted 108 screens in human cell lines, comprising >540,000 perturbations across 24.85 megabases of the genome. Using 332 functionally confirmed CRE-gene links in K562 cells, we established guidelines for screening endogenous noncoding elements with CRISPR interference (CRISPRi), including accurate detection of CREs that exhibit variable, often low, transcriptional effects. Benchmarking five screen analysis tools, we find that CASA produces the most conservative CRE calls and is robust to artifacts of low-specificity single guide RNAs. We uncover a subtle DNA strand bias for CRISPRi in transcribed regions with implications for screen design and analysis. Together, we provide an accessible data resource, predesigned single guide RNAs for targeting 3,275,697 ENCODE SCREEN candidate CREs with CRISPRi and screening guidelines to accelerate functional characterization of the noncoding genome., (© 2024. The Author(s).)
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- 2024
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7. Vocal learning-associated convergent evolution in mammalian proteins and regulatory elements.
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Wirthlin ME, Schmid TA, Elie JE, Zhang X, Kowalczyk A, Redlich R, Shvareva VA, Rakuljic A, Ji MB, Bhat NS, Kaplow IM, Schäffer DE, Lawler AJ, Wang AZ, Phan BN, Annaldasula S, Brown AR, Lu T, Lim BK, Azim E, Clark NL, Meyer WK, Pond SLK, Chikina M, Yartsev MM, Pfenning AR, Andrews G, Armstrong JC, Bianchi M, Birren BW, Bredemeyer KR, Breit AM, Christmas MJ, Clawson H, Damas J, Di Palma F, Diekhans M, Dong MX, Eizirik E, Fan K, Fanter C, Foley NM, Forsberg-Nilsson K, Garcia CJ, Gatesy J, Gazal S, Genereux DP, Goodman L, Grimshaw J, Halsey MK, Harris AJ, Hickey G, Hiller M, Hindle AG, Hubley RM, Hughes GM, Johnson J, Juan D, Kaplow IM, Karlsson EK, Keough KC, Kirilenko B, Koepfli KP, Korstian JM, Kowalczyk A, Kozyrev SV, Lawler AJ, Lawless C, Lehmann T, Levesque DL, Lewin HA, Li X, Lind A, Lindblad-Toh K, Mackay-Smith A, Marinescu VD, Marques-Bonet T, Mason VC, Meadows JRS, Meyer WK, Moore JE, Moreira LR, Moreno-Santillan DD, Morrill KM, Muntané G, Murphy WJ, Navarro A, Nweeia M, Ortmann S, Osmanski A, Paten B, Paulat NS, Pfenning AR, Phan BN, Pollard KS, Pratt HE, Ray DA, Reilly SK, Rosen JR, Ruf I, Ryan L, Ryder OA, Sabeti PC, Schäffer DE, Serres A, Shapiro B, Smit AFA, Springer M, Srinivasan C, Steiner C, Storer JM, Sullivan KAM, Sullivan PF, Sundström E, Supple MA, Swofford R, Talbot JE, Teeling E, Turner-Maier J, Valenzuela A, Wagner F, Wallerman O, Wang C, Wang J, Weng Z, Wilder AP, Wirthlin ME, Xue JR, and Zhang X
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- Animals, Chiroptera genetics, Chiroptera physiology, Chromatin metabolism, Larynx physiology, Epigenesis, Genetic, Genome, Amino Acid Sequence, Machine Learning, Vocalization, Animal physiology, Motor Cortex cytology, Motor Cortex physiology, Enhancer Elements, Genetic, Motor Neurons physiology, Gene Expression Regulation, Evolution, Molecular, Proteins genetics, Proteins metabolism, Eutheria genetics, Eutheria physiology
- Abstract
Vocal production learning ("vocal learning") is a convergently evolved trait in vertebrates. To identify brain genomic elements associated with mammalian vocal learning, we integrated genomic, anatomical, and neurophysiological data from the Egyptian fruit bat ( Rousettus aegyptiacus ) with analyses of the genomes of 215 placental mammals. First, we identified a set of proteins evolving more slowly in vocal learners. Then, we discovered a vocal motor cortical region in the Egyptian fruit bat, an emergent vocal learner, and leveraged that knowledge to identify active cis-regulatory elements in the motor cortex of vocal learners. Machine learning methods applied to motor cortex open chromatin revealed 50 enhancers robustly associated with vocal learning whose activity tended to be lower in vocal learners. Our research implicates convergent losses of motor cortex regulatory elements in mammalian vocal learning evolution.
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- 2024
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8. Genome-wide association study identifies human genetic variants associated with fatal outcome from Lassa fever.
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Kotliar D, Raju S, Tabrizi S, Odia I, Goba A, Momoh M, Sandi JD, Nair P, Phelan E, Tariyal R, Eromon PE, Mehta S, Robles-Sikisaka R, Siddle KJ, Stremlau M, Jalloh S, Gire SK, Winnicki S, Chak B, Schaffner SF, Pauthner M, Karlsson EK, Chapin SR, Kennedy SG, Branco LM, Kanneh L, Vitti JJ, Broodie N, Gladden-Young A, Omoniwa O, Jiang PP, Yozwiak N, Heuklom S, Moses LM, Akpede GO, Asogun DA, Rubins K, Kales S, Happi AN, Iruolagbe CO, Dic-Ijiewere M, Iraoyah K, Osazuwa OO, Okonkwo AK, Kunz S, McCormick JB, Khan SH, Honko AN, Lander ES, Oldstone MBA, Hensley L, Folarin OA, Okogbenin SA, Günther S, Ollila HM, Tewhey R, Okokhere PO, Schieffelin JS, Andersen KG, Reilly SK, Grant DS, Garry RF, Barnes KG, Happi CT, and Sabeti PC
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- Humans, Genome-Wide Association Study, Seroepidemiologic Studies, Lassa virus genetics, Fever, Human Genetics, Lassa Fever genetics, Lassa Fever diagnosis, Lassa Fever epidemiology
- Abstract
Infection with Lassa virus (LASV) can cause Lassa fever, a haemorrhagic illness with an estimated fatality rate of 29.7%, but causes no or mild symptoms in many individuals. Here, to investigate whether human genetic variation underlies the heterogeneity of LASV infection, we carried out genome-wide association studies (GWAS) as well as seroprevalence surveys, human leukocyte antigen typing and high-throughput variant functional characterization assays. We analysed Lassa fever susceptibility and fatal outcomes in 533 cases of Lassa fever and 1,986 population controls recruited over a 7 year period in Nigeria and Sierra Leone. We detected genome-wide significant variant associations with Lassa fever fatal outcomes near GRM7 and LIF in the Nigerian cohort. We also show that a haplotype bearing signatures of positive selection and overlapping LARGE1, a required LASV entry factor, is associated with decreased risk of Lassa fever in the Nigerian cohort but not in the Sierra Leone cohort. Overall, we identified variants and genes that may impact the risk of severe Lassa fever, demonstrating how GWAS can provide insight into viral pathogenesis., (© 2024. The Author(s).)
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- 2024
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9. Nonviral In Vivo Delivery of CRISPR-Cas9 Using Protein-Agnostic, High-Loading Porous Silicon and Polymer Nanoparticles.
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Fletcher RB, Stokes LD, Kelly IB 3rd, Henderson KM, Vallecillo-Viejo IC, Colazo JM, Wong BV, Yu F, d'Arcy R, Struthers MN, Evans BC, Ayers J, Castanon M, Weirich MJ, Reilly SK, Patel SS, Ivanova YI, Silvera Batista CA, Weiss SM, Gersbach CA, Brunger JM, and Duvall CL
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- Mice, Animals, Silicon, Porosity, Polymers, CRISPR-Cas Systems genetics, Nanoparticles
- Abstract
The complexity of CRISPR machinery is a challenge to its application for nonviral in vivo therapeutic gene editing. Here, we demonstrate that proteins, regardless of size or charge, efficiently load into porous silicon nanoparticles (PSiNPs). Optimizing the loading strategy yields formulations that are ultrahigh loading─>40% cargo by volume─and highly active. Further tuning of a polymeric coating on the loaded PSiNPs yields nanocomposites that achieve colloidal stability under cryopreservation, endosome escape, and gene editing efficiencies twice that of the commercial standard Lipofectamine CRISPRMAX. In a mouse model of arthritis, PSiNPs edit cells in both the cartilage and synovium of knee joints, and achieve 60% reduction in expression of the therapeutically relevant MMP13 gene. Administered intramuscularly, they are active over a broad dose range, with the highest tested dose yielding nearly 100% muscle fiber editing at the injection site. The nanocomposite PSiNPs are also amenable to systemic delivery. Administered intravenously in a model that mimics muscular dystrophy, they edit sites of inflamed muscle. Collectively, the results demonstrate that the PSiNP nanocomposites are a versatile system that can achieve high loading of diverse cargoes and can be applied for gene editing in both local and systemic delivery applications.
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- 2023
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10. Machine-guided design of synthetic cell type-specific cis -regulatory elements.
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Gosai SJ, Castro RI, Fuentes N, Butts JC, Kales S, Noche RR, Mouri K, Sabeti PC, Reilly SK, and Tewhey R
- Abstract
Cis -regulatory elements (CREs) control gene expression, orchestrating tissue identity, developmental timing, and stimulus responses, which collectively define the thousands of unique cell types in the body. While there is great potential for strategically incorporating CREs in therapeutic or biotechnology applications that require tissue specificity, there is no guarantee that an optimal CRE for an intended purpose has arisen naturally through evolution. Here, we present a platform to engineer and validate synthetic CREs capable of driving gene expression with programmed cell type specificity. We leverage innovations in deep neural network modeling of CRE activity across three cell types, efficient in silico optimization, and massively parallel reporter assays (MPRAs) to design and empirically test thousands of CREs. Through in vitro and in vivo validation, we show that synthetic sequences outperform natural sequences from the human genome in driving cell type-specific expression. Synthetic sequences leverage unique sequence syntax to promote activity in the on-target cell type and simultaneously reduce activity in off-target cells. Together, we provide a generalizable framework to prospectively engineer CREs and demonstrate the required literacy to write regulatory code that is fit-for-purpose in vivo across vertebrates., Competing Interests: Competing Interests PCS is a co-founder of and consultant to Sherlock Biosciences and Board Member of Danaher Corporation. PCS and RT have filed intellectual property related to MPRA. SJG, RIC, SKR, PCS, and RT have filed a provisional patent application related to work described here.
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- 2023
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11. Evolutionary constraint and innovation across hundreds of placental mammals.
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Christmas MJ, Kaplow IM, Genereux DP, Dong MX, Hughes GM, Li X, Sullivan PF, Hindle AG, Andrews G, Armstrong JC, Bianchi M, Breit AM, Diekhans M, Fanter C, Foley NM, Goodman DB, Goodman L, Keough KC, Kirilenko B, Kowalczyk A, Lawless C, Lind AL, Meadows JRS, Moreira LR, Redlich RW, Ryan L, Swofford R, Valenzuela A, Wagner F, Wallerman O, Brown AR, Damas J, Fan K, Gatesy J, Grimshaw J, Johnson J, Kozyrev SV, Lawler AJ, Marinescu VD, Morrill KM, Osmanski A, Paulat NS, Phan BN, Reilly SK, Schäffer DE, Steiner C, Supple MA, Wilder AP, Wirthlin ME, Xue JR, Birren BW, Gazal S, Hubley RM, Koepfli KP, Marques-Bonet T, Meyer WK, Nweeia M, Sabeti PC, Shapiro B, Smit AFA, Springer MS, Teeling EC, Weng Z, Hiller M, Levesque DL, Lewin HA, Murphy WJ, Navarro A, Paten B, Pollard KS, Ray DA, Ruf I, Ryder OA, Pfenning AR, Lindblad-Toh K, and Karlsson EK
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- Animals, Female, Humans, Conserved Sequence genetics, Genome, Human, Eutheria genetics, Evolution, Molecular
- Abstract
Zoonomia is the largest comparative genomics resource for mammals produced to date. By aligning genomes for 240 species, we identify bases that, when mutated, are likely to affect fitness and alter disease risk. At least 332 million bases (~10.7%) in the human genome are unusually conserved across species (evolutionarily constrained) relative to neutrally evolving repeats, and 4552 ultraconserved elements are nearly perfectly conserved. Of 101 million significantly constrained single bases, 80% are outside protein-coding exons and half have no functional annotations in the Encyclopedia of DNA Elements (ENCODE) resource. Changes in genes and regulatory elements are associated with exceptional mammalian traits, such as hibernation, that could inform therapeutic development. Earth's vast and imperiled biodiversity offers distinctive power for identifying genetic variants that affect genome function and organismal phenotypes.
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- 2023
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12. The functional and evolutionary impacts of human-specific deletions in conserved elements.
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Xue JR, Mackay-Smith A, Mouri K, Garcia MF, Dong MX, Akers JF, Noble M, Li X, Lindblad-Toh K, Karlsson EK, Noonan JP, Capellini TD, Brennand KJ, Tewhey R, Sabeti PC, and Reilly SK
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- Humans, Conserved Sequence genetics, Genome, Genomics, RNA-Binding Proteins genetics, Evolution, Molecular, Sequence Deletion, Brain growth & development, Gene Expression Regulation, Developmental
- Abstract
Conserved genomic sequences disrupted in humans may underlie uniquely human phenotypic traits. We identified and characterized 10,032 human-specific conserved deletions (hCONDELs). These short (average 2.56 base pairs) deletions are enriched for human brain functions across genetic, epigenomic, and transcriptomic datasets. Using massively parallel reporter assays in six cell types, we discovered 800 hCONDELs conferring significant differences in regulatory activity, half of which enhance rather than disrupt regulatory function. We highlight several hCONDELs with putative human-specific effects on brain development, including HDAC5 , CPEB4 , and PPP2CA . Reverting an hCONDEL to the ancestral sequence alters the expression of LOXL2 and developmental genes involved in myelination and synaptic function. Our data provide a rich resource to investigate the evolutionary mechanisms driving new traits in humans and other species.
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- 2023
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13. Leveraging base-pair mammalian constraint to understand genetic variation and human disease.
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Sullivan PF, Meadows JRS, Gazal S, Phan BN, Li X, Genereux DP, Dong MX, Bianchi M, Andrews G, Sakthikumar S, Nordin J, Roy A, Christmas MJ, Marinescu VD, Wang C, Wallerman O, Xue J, Yao S, Sun Q, Szatkiewicz J, Wen J, Huckins LM, Lawler A, Keough KC, Zheng Z, Zeng J, Wray NR, Li Y, Johnson J, Chen J, Paten B, Reilly SK, Hughes GM, Weng Z, Pollard KS, Pfenning AR, Forsberg-Nilsson K, Karlsson EK, and Lindblad-Toh K
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- Animals, Humans, Biological Evolution, Genome, Human, Genome-Wide Association Study, Genomics, Molecular Sequence Annotation, Polymorphism, Single Nucleotide, Genetic Variation, Disease genetics
- Abstract
Thousands of genomic regions have been associated with heritable human diseases, but attempts to elucidate biological mechanisms are impeded by an inability to discern which genomic positions are functionally important. Evolutionary constraint is a powerful predictor of function, agnostic to cell type or disease mechanism. Single-base phyloP scores from 240 mammals identified 3.3% of the human genome as significantly constrained and likely functional. We compared phyloP scores to genome annotation, association studies, copy-number variation, clinical genetics findings, and cancer data. Constrained positions are enriched for variants that explain common disease heritability more than other functional annotations. Our results improve variant annotation but also highlight that the regulatory landscape of the human genome still needs to be further explored and linked to disease.
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- 2023
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14. Transmission from vaccinated individuals in a large SARS-CoV-2 Delta variant outbreak.
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Siddle KJ, Krasilnikova LA, Moreno GK, Schaffner SF, Vostok J, Fitzgerald NA, Lemieux JE, Barkas N, Loreth C, Specht I, Tomkins-Tinch CH, Paull JS, Schaeffer B, Taylor BP, Loftness B, Johnson H, Schubert PL, Shephard HM, Doucette M, Fink T, Lang AS, Baez S, Beauchamp J, Hennigan S, Buzby E, Ash S, Brown J, Clancy S, Cofsky S, Gagne L, Hall J, Harrington R, Gionet GL, DeRuff KC, Vodzak ME, Adams GC, Dobbins ST, Slack SD, Reilly SK, Anderson LM, Cipicchio MC, DeFelice MT, Grimsby JL, Anderson SE, Blumenstiel BS, Meldrim JC, Rooke HM, Vicente G, Smith NL, Messer KS, Reagan FL, Mandese ZM, Lee MD, Ray MC, Fisher ME, Ulcena MA, Nolet CM, English SE, Larkin KL, Vernest K, Chaluvadi S, Arvidson D, Melchiono M, Covell T, Harik V, Brock-Fisher T, Dunn M, Kearns A, Hanage WP, Bernard C, Philippakis A, Lennon NJ, Gabriel SB, Gallagher GR, Smole S, Madoff LC, Brown CM, Park DJ, MacInnis BL, and Sabeti PC
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- Adolescent, Adult, Aged, Aged, 80 and over, COVID-19 virology, Child, Child, Preschool, Contact Tracing methods, Disease Outbreaks, Female, Genome, Viral, Humans, Infant, Infant, Newborn, Male, Massachusetts epidemiology, Middle Aged, Molecular Epidemiology, Phylogeny, SARS-CoV-2 classification, Vaccination, Whole Genome Sequencing, Young Adult, COVID-19 epidemiology, COVID-19 immunology, COVID-19 transmission, SARS-CoV-2 genetics, SARS-CoV-2 immunology
- Abstract
An outbreak of over 1,000 COVID-19 cases in Provincetown, Massachusetts (MA), in July 2021-the first large outbreak mostly in vaccinated individuals in the US-prompted a comprehensive public health response, motivating changes to national masking recommendations and raising questions about infection and transmission among vaccinated individuals. To address these questions, we combined viral genomic and epidemiological data from 467 individuals, including 40% of outbreak-associated cases. The Delta variant accounted for 99% of cases in this dataset; it was introduced from at least 40 sources, but 83% of cases derived from a single source, likely through transmission across multiple settings over a short time rather than a single event. Genomic and epidemiological data supported multiple transmissions of Delta from and between fully vaccinated individuals. However, despite its magnitude, the outbreak had limited onward impact in MA and the US overall, likely due to high vaccination rates and a robust public health response., Competing Interests: Declaration of interests P.C.S. is a co-founder of, shareholder in, and scientific advisor to Sherlock Biosciences, as well as a board member of and shareholder in Danaher Corporation. J.E.L. has received consulting fees from Sherlock Biosciences. A.P. is a venture partner at Google Ventures. W.P.H. is a member of the scientific advisory board of Biobot. Other authors report no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)
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- 2022
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15. Detection of Neanderthal Adaptively Introgressed Genetic Variants That Modulate Reporter Gene Expression in Human Immune Cells.
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Jagoda E, Xue JR, Reilly SK, Dannemann M, Racimo F, Huerta-Sanchez E, Sankararaman S, Kelso J, Pagani L, Sabeti PC, and Capellini TD
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- Animals, Gene Expression, Humans, Inflammation, Genetic Variation, Genome, Human, Immunity, Innate genetics, Neanderthals genetics
- Abstract
Although some variation introgressed from Neanderthals has undergone selective sweeps, little is known about its functional significance. We used a Massively Parallel Reporter Assay (MPRA) to assay 5,353 high-frequency introgressed variants for their ability to modulate the gene expression within 170 bp of endogenous sequence. We identified 2,548 variants in active putative cis-regulatory elements (CREs) and 292 expression-modulating variants (emVars). These emVars are predicted to alter the binding motifs of important immune transcription factors, are enriched for associations with neutrophil and white blood cell count, and are associated with the expression of genes that function in innate immune pathways including inflammatory response and antiviral defense. We combined the MPRA data with other data sets to identify strong candidates to be driver variants of positive selection including an emVar that may contribute to protection against severe COVID-19 response. We endogenously deleted two CREs containing expression-modulation variants linked to immune function, rs11624425 and rs80317430, identifying their primary genic targets as ELMSAN1, and PAN2 and STAT2, respectively, three genes differentially expressed during influenza infection. Overall, we present the first database of experimentally identified expression-modulating Neanderthal-introgressed alleles contributing to potential immune response in modern humans., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)
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- 2022
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16. Synthetic DNA spike-ins (SDSIs) enable sample tracking and detection of inter-sample contamination in SARS-CoV-2 sequencing workflows.
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Lagerborg KA, Normandin E, Bauer MR, Adams G, Figueroa K, Loreth C, Gladden-Young A, Shaw BM, Pearlman LR, Berenzy D, Dewey HB, Kales S, Dobbins ST, Shenoy ES, Hooper D, Pierce VM, Zachary KC, Park DJ, MacInnis BL, Tewhey R, Lemieux JE, Sabeti PC, Reilly SK, and Siddle KJ
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- COVID-19 diagnosis, DNA Primers chemical synthesis, Genome, Viral genetics, Humans, Quality Control, RNA, Viral genetics, Reproducibility of Results, Sequence Analysis methods, Whole Genome Sequencing, Workflow, DNA Primers standards, SARS-CoV-2 genetics, Sequence Analysis standards
- Abstract
The global spread and continued evolution of SARS-CoV-2 has driven an unprecedented surge in viral genomic surveillance. Amplicon-based sequencing methods provide a sensitive, low-cost and rapid approach but suffer a high potential for contamination, which can undermine laboratory processes and results. This challenge will increase with the expanding global production of sequences across a variety of laboratories for epidemiological and clinical interpretation, as well as for genomic surveillance of emerging diseases in future outbreaks. We present SDSI + AmpSeq, an approach that uses 96 synthetic DNA spike-ins (SDSIs) to track samples and detect inter-sample contamination throughout the sequencing workflow. We apply SDSIs to the ARTIC Consortium's amplicon design, demonstrate their utility and efficiency in a real-time investigation of a suspected hospital cluster of SARS-CoV-2 cases and validate them across 6,676 diagnostic samples at multiple laboratories. We establish that SDSI + AmpSeq provides increased confidence in genomic data by detecting and correcting for relatively common, yet previously unobserved modes of error, including spillover and sample swaps, without impacting genome recovery., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)
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- 2022
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17. Evidence of transmission from fully vaccinated individuals in a large outbreak of the SARS-CoV-2 Delta variant in Provincetown, Massachusetts.
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Siddle KJ, Krasilnikova LA, Moreno GK, Schaffner SF, Vostok J, Fitzgerald NA, Lemieux JE, Barkas N, Loreth C, Specht I, Tomkins-Tinch CH, Silbert J, Schaeffer B, Taylor BP, Loftness B, Johnson H, Schubert PL, Shephard HM, Doucette M, Fink T, Lang AS, Baez S, Beauchamp J, Hennigan S, Buzby E, Ash S, Brown J, Clancy S, Cofsky S, Gagne L, Hall J, Harrington R, Gionet GL, DeRuff KC, Vodzak ME, Adams GC, Dobbins ST, Slack SD, Reilly SK, Anderson LM, Cipicchio MC, DeFelice MT, Grimsby JL, Anderson SE, Blumenstiel BS, Meldrim JC, Rooke HM, Vicente G, Smith NL, Messer KS, Reagan FL, Mandese ZM, Lee MD, Ray MC, Fisher ME, Ulcena MA, Nolet CM, English SE, Larkin KL, Vernest K, Chaluvadi S, Arvidson D, Melchiono M, Covell T, Harik V, Brock-Fisher T, Dunn M, Kearns A, Hanage WP, Bernard C, Philippakis A, Lennon NJ, Gabriel SB, Gallagher GR, Smole S, Madoff LC, Brown CM, Park DJ, MacInnis BL, and Sabeti PC
- Abstract
Multiple summer events, including large indoor gatherings, in Provincetown, Massachusetts (MA), in July 2021 contributed to an outbreak of over one thousand COVID-19 cases among residents and visitors. Most cases were fully vaccinated, many of whom were also symptomatic, prompting a comprehensive public health response, motivating changes to national masking recommendations, and raising questions about infection and transmission among vaccinated individuals. To characterize the outbreak and the viral population underlying it, we combined genomic and epidemiological data from 467 individuals, including 40% of known outbreak-associated cases. The Delta variant accounted for 99% of sequenced outbreak-associated cases. Phylogenetic analysis suggests over 40 sources of Delta in the dataset, with one responsible for a single cluster containing 83% of outbreak-associated genomes. This cluster was likely not the result of extensive spread at a single site, but rather transmission from a common source across multiple settings over a short time. Genomic and epidemiological data combined provide strong support for 25 transmission events from, including many between, fully vaccinated individuals; genomic data alone provides evidence for an additional 64. Together, genomic epidemiology provides a high-resolution picture of the Provincetown outbreak, revealing multiple cases of transmission of Delta from fully vaccinated individuals. However, despite its magnitude, the outbreak was restricted in its onward impact in MA and the US, likely due to high vaccination rates and a robust public health response.
- Published
- 2021
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18. Author Correction: Direct characterization of cis-regulatory elements and functional dissection of complex genetic associations using HCR-FlowFISH.
- Author
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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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19. Genome-wide functional screen of 3'UTR variants uncovers causal variants for human disease and evolution.
- Author
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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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20. 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).)
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- 2021
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21. 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.)
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- 2021
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22. DNA spike-ins enable confident interpretation of SARS-CoV-2 genomic data from amplicon-based sequencing.
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Lagerborg KA, Normandin E, Bauer MR, Adams G, Figueroa K, Loreth C, Gladden-Young A, Shaw B, Pearlman L, Shenoy ES, Hooper D, Pierce VM, Zachary KC, Park DJ, MacInnis BL, Lemieux JE, Sabeti PC, Reilly SK, and Siddle KJ
- Abstract
The rapid global spread and continued evolution of SARS-CoV-2 has highlighted an unprecedented need for viral genomic surveillance and clinical viral sequencing. Amplicon-based sequencing methods provide a sensitive, low-cost and rapid approach but suffer a high potential for contamination, which can undermine lab processes and results. This challenge will only increase with expanding global production of sequences by diverse research groups for epidemiological and clinical interpretation. We present an approach which uses synthetic DNA spike-ins (SDSIs) to track samples and detect inter-sample contamination through a sequencing workflow. Applying this approach to the ARTIC Consortium's amplicon design, we define a series of best practices for Illumina-based sequencing and provide a detailed characterization of approaches to increase sensitivity for low-viral load samples incorporating the SDSIs. We demonstrate the utility and efficiency of the SDSI method amidst a real-time investigation of a suspected hospital cluster of SARS-CoV-2 cases., Competing Interests: Competing interests J.E.L. has received consulting fees from Sherlock Biosciences. P.C.S. is a co-founder of, shareholder in, and scientific advisor to Sherlock Biosciences, Inc., as well as a Board member of and shareholder in Danaher Corporation. M.R.B., K.A.L., E.N., S.K.R., K.J.S., P.C.S are co-inventors on a patent application filed by Broad Institute relating to methods of this manuscript.
- Published
- 2021
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23. Phylogenetic analysis of SARS-CoV-2 in Boston highlights the impact of superspreading events.
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Lemieux JE, Siddle KJ, Shaw BM, Loreth C, Schaffner SF, Gladden-Young A, Adams G, Fink T, Tomkins-Tinch CH, Krasilnikova LA, DeRuff KC, Rudy M, Bauer MR, Lagerborg KA, Normandin E, Chapman SB, Reilly SK, Anahtar MN, Lin AE, Carter A, Myhrvold C, Kemball ME, Chaluvadi S, Cusick C, Flowers K, Neumann A, Cerrato F, Farhat M, Slater D, Harris JB, Branda JA, Hooper D, Gaeta JM, Baggett TP, O'Connell J, Gnirke A, Lieberman TD, Philippakis A, Burns M, Brown CM, Luban J, Ryan ET, Turbett SE, LaRocque RC, Hanage WP, Gallagher GR, Madoff LC, Smole S, Pierce VM, Rosenberg E, Sabeti PC, Park DJ, and MacInnis BL
- Subjects
- Boston epidemiology, COVID-19 transmission, Disease Outbreaks, Epidemiological Monitoring, Humans, COVID-19 epidemiology, Genome, Viral, Phylogeny, SARS-CoV-2 genetics
- Abstract
Analysis of 772 complete severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomes from early in the Boston-area epidemic revealed numerous introductions of the virus, a small number of which led to most cases. The data revealed two superspreading events. One, in a skilled nursing facility, led to rapid transmission and significant mortality in this vulnerable population but little broader spread, whereas other introductions into the facility had little effect. The second, at an international business conference, produced sustained community transmission and was exported, resulting in extensive regional, national, and international spread. The two events also differed substantially in the genetic variation they generated, suggesting varying transmission dynamics in superspreading events. Our results show how genomic epidemiology can help to understand the link between individual clusters and wider community spread., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)
- Published
- 2021
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24. Massively parallel discovery of human-specific substitutions that alter enhancer activity.
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Uebbing S, Gockley J, Reilly SK, Kocher AA, Geller E, Gandotra N, Scharfe C, Cotney J, and Noonan JP
- Subjects
- Animals, Humans, Neocortex, Pan troglodytes genetics, Biological Evolution, Enhancer Elements, Genetic, Genome, Human, Neural Stem Cells metabolism, Transcription Factors metabolism
- Abstract
Genetic changes that altered the function of gene regulatory elements have been implicated in the evolution of human traits such as the expansion of the cerebral cortex. However, identifying the particular changes that modified regulatory activity during human evolution remain challenging. Here we used massively parallel enhancer assays in neural stem cells to quantify the functional impact of >32,000 human-specific substitutions in >4,300 human accelerated regions (HARs) and human gain enhancers (HGEs), which include enhancers with novel activities in humans. We found that >30% of active HARs and HGEs exhibited differential activity between human and chimpanzee. We isolated the effects of human-specific substitutions from background genetic variation to identify the effects of genetic changes most relevant to human evolution. We found that substitutions interacted in both additive and nonadditive ways to modify enhancer function. Substitutions within HARs, which are highly constrained compared to HGEs, showed smaller effects on enhancer activity, suggesting that the impact of human-specific substitutions is buffered in enhancers with constrained ancestral functions. Our findings yield insight into how human-specific genetic changes altered enhancer function and provide a rich set of candidates for studies of regulatory evolution in humans., Competing Interests: The authors declare no competing interest., (Copyright © 2020 the Author(s). Published by PNAS.)
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- 2021
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25. Phylogenetic analysis of SARS-CoV-2 in the Boston area highlights the role of recurrent importation and superspreading events.
- Author
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Lemieux JE, Siddle KJ, Shaw BM, Loreth C, Schaffner SF, Gladden-Young A, Adams G, Fink T, Tomkins-Tinch CH, Krasilnikova LA, DeRuff KC, Rudy M, Bauer MR, Lagerborg KA, Normandin E, Chapman SB, Reilly SK, Anahtar MN, Lin AE, Carter A, Myhrvold C, Kemball ME, Chaluvadi S, Cusick C, Flowers K, Neumann A, Cerrato F, Farhat M, Slater D, Harris JB, Branda J, Hooper D, Gaeta JM, Baggett TP, O'Connell J, Gnirke A, Lieberman TD, Philippakis A, Burns M, Brown CM, Luban J, Ryan ET, Turbett SE, LaRocque RC, Hanage WP, Gallagher GR, Madoff LC, Smole S, Pierce VM, Rosenberg E, Sabeti PC, Park DJ, and Maclnnis BL
- Abstract
SARS-CoV-2 has caused a severe, ongoing outbreak of COVID-19 in Massachusetts with 111,070 confirmed cases and 8,433 deaths as of August 1, 2020. To investigate the introduction, spread, and epidemiology of COVID-19 in the Boston area, we sequenced and analyzed 772 complete SARS-CoV-2 genomes from the region, including nearly all confirmed cases within the first week of the epidemic and hundreds of cases from major outbreaks at a conference, a nursing facility, and among homeless shelter guests and staff. The data reveal over 80 introductions into the Boston area, predominantly from elsewhere in the United States and Europe. We studied two superspreading events covered by the data, events that led to very different outcomes because of the timing and populations involved. One produced rapid spread in a vulnerable population but little onward transmission, while the other was a major contributor to sustained community transmission, including outbreaks in homeless populations, and was exported to several other domestic and international sites. The same two events differed significantly in the number of new mutations seen, raising the possibility that SARS-CoV-2 superspreading might encompass disparate transmission dynamics. Our results highlight the failure of measures to prevent importation into MA early in the outbreak, underscore the role of superspreading in amplifying an outbreak in a major urban area, and lay a foundation for contact tracing informed by genetic data., Competing Interests: Competing interests: J.E.L. has received consulting fees from Sherlock Biosciences. J.B. has been a consultant for T2 Biosystems, DiaSorin, and Roche Diagnostics. A.P. is a Venture Partner at Google Ventures. P.C.S. is a co-founder and shareholder of Sherlock Biosciences, and a Board member and shareholder of Danaher Corporation.
- Published
- 2020
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26. 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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27. Direct Identification of Hundreds of Expression-Modulating Variants using a Multiplexed Reporter Assay.
- Author
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Tewhey R, Kotliar D, Park DS, Liu B, Winnicki S, Reilly SK, Andersen KG, Mikkelsen TS, Lander ES, Schaffner SF, and Sabeti PC
- Published
- 2018
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28. Evolution of Gene Regulation in Humans.
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Reilly SK and Noonan JP
- Subjects
- Animals, Genome, Humans, Mice, Mice, Transgenic genetics, Primates genetics, Evolution, Molecular, Gene Expression Regulation genetics, Induced Pluripotent Stem Cells
- Abstract
As a species, we possess unique biological features that distinguish us from other primates. Here, we review recent efforts to identify changes in gene regulation that drove the evolution of novel human phenotypes. We discuss genotype-directed comparisons of human and nonhuman primate genomes to identify human-specific genetic changes that may encode new regulatory functions. We also review phenotype-directed approaches, which use comparisons of gene expression or regulatory function in homologous human and nonhuman primate cells and tissues to identify changes in expression levels or regulatory activity that may be due to genetic changes in humans. Together, these studies are beginning to reveal the landscape of regulatory innovation in human evolution and point to specific regulatory changes for further study. Finally, we highlight two novel strategies to model human-specific regulatory functions in vivo: primate induced pluripotent stem cells and the generation of humanized mice by genome editing.
- Published
- 2016
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29. Direct Identification of Hundreds of Expression-Modulating Variants using a Multiplexed Reporter Assay.
- Author
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Tewhey R, Kotliar D, Park DS, Liu B, Winnicki S, Reilly SK, Andersen KG, Mikkelsen TS, Lander ES, Schaffner SF, and Sabeti PC
- Subjects
- Alleles, Gene Library, Hep G2 Cells, Humans, Quantitative Trait Loci, Sensitivity and Specificity, Spondylitis, Ankylosing genetics, Gene Expression Regulation, Genes, Reporter, Genetic Diseases, Inborn genetics, Genetic Techniques, Genetic Variation
- Abstract
Although studies have identified hundreds of loci associated with human traits and diseases, pinpointing causal alleles remains difficult, particularly for non-coding variants. To address this challenge, we adapted the massively parallel reporter assay (MPRA) to identify variants that directly modulate gene expression. We applied it to 32,373 variants from 3,642 cis-expression quantitative trait loci and control regions. Detection by MPRA was strongly correlated with measures of regulatory function. We demonstrate MPRA's capabilities for pinpointing causal alleles, using it to identify 842 variants showing differential expression between alleles, including 53 well-annotated variants associated with diseases and traits. We investigated one in detail, a risk allele for ankylosing spondylitis, and provide direct evidence of a non-coding variant that alters expression of the prostaglandin EP4 receptor. These results create a resource of concrete leads and illustrate the promise of this approach for comprehensively interrogating how non-coding polymorphism shapes human biology., (Copyright © 2016 Elsevier Inc. All rights reserved.)
- Published
- 2016
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30. Origin and evolution of developmental enhancers in the mammalian neocortex.
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Emera D, Yin J, Reilly SK, Gockley J, and Noonan JP
- Subjects
- Animals, Base Sequence, Computer Simulation, Humans, Mice, Models, Genetic, Neocortex embryology, Neocortex metabolism, Species Specificity, Biological Evolution, Enhancer Elements, Genetic genetics, Gene Expression Regulation, Developmental genetics, Morphogenesis genetics, Neocortex growth & development, Transcription Factors genetics
- Abstract
Morphological innovations such as the mammalian neocortex may involve the evolution of novel regulatory sequences. However, de novo birth of regulatory elements active during morphogenesis has not been extensively studied in mammals. Here, we use H3K27ac-defined regulatory elements active during human and mouse corticogenesis to identify enhancers that were likely active in the ancient mammalian forebrain. We infer the phylogenetic origins of these enhancers and find that ∼20% arose in the mammalian stem lineage, coincident with the emergence of the neocortex. Implementing a permutation strategy that controls for the nonrandom variation in the ages of background genomic sequences, we find that mammal-specific enhancers are overrepresented near genes involved in cell migration, cell signaling, and axon guidance. Mammal-specific enhancers are also overrepresented in modules of coexpressed genes in the cortex that are associated with these pathways, notably ephrin and semaphorin signaling. Our results also provide insight into the mechanisms of regulatory innovation in mammals. We find that most neocortical enhancers did not originate by en bloc exaptation of transposons. Young neocortical enhancers exhibit smaller H3K27ac footprints and weaker evolutionary constraint in eutherian mammals than older neocortical enhancers. Based on these observations, we present a model of the enhancer life cycle in which neocortical enhancers initially emerge from genomic background as short, weakly constrained "proto-enhancers." Many proto-enhancers are likely lost, but some may serve as nucleation points for complex enhancers to evolve.
- Published
- 2016
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31. The autism-associated chromatin modifier CHD8 regulates other autism risk genes during human neurodevelopment.
- Author
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Cotney J, Muhle RA, Sanders SJ, Liu L, Willsey AJ, Niu W, Liu W, Klei L, Lei J, Yin J, Reilly SK, Tebbenkamp AT, Bichsel C, Pletikos M, Sestan N, Roeder K, State MW, Devlin B, and Noonan JP
- Subjects
- Animals, Chromatin Assembly and Disassembly genetics, DNA-Binding Proteins genetics, Gene Expression Regulation, Developmental genetics, Gene Knockdown Techniques, Humans, Mice, Nervous System metabolism, Neural Stem Cells metabolism, Transcription Factors genetics, Autism Spectrum Disorder genetics, DNA-Binding Proteins metabolism, Gene Expression Regulation, Developmental physiology, Gene Regulatory Networks genetics, Models, Neurological, Nervous System embryology, Transcription Factors metabolism
- Abstract
Recent studies implicate chromatin modifiers in autism spectrum disorder (ASD) through the identification of recurrent de novo loss of function mutations in affected individuals. ASD risk genes are co-expressed in human midfetal cortex, suggesting that ASD risk genes converge in specific regulatory networks during neurodevelopment. To elucidate such networks, we identify genes targeted by CHD8, a chromodomain helicase strongly associated with ASD, in human midfetal brain, human neural stem cells (hNSCs) and embryonic mouse cortex. CHD8 targets are strongly enriched for other ASD risk genes in both human and mouse neurodevelopment, and converge in ASD-associated co-expression networks in human midfetal cortex. CHD8 knockdown in hNSCs results in dysregulation of ASD risk genes directly targeted by CHD8. Integration of CHD8-binding data into ASD risk models improves detection of risk genes. These results suggest loss of CHD8 contributes to ASD by perturbing an ancient gene regulatory network during human brain development.
- Published
- 2015
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32. Evolutionary genomics. Evolutionary changes in promoter and enhancer activity during human corticogenesis.
- Author
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Reilly SK, Yin J, Ayoub AE, Emera D, Leng J, Cotney J, Sarro R, Rakic P, and Noonan JP
- Subjects
- Animals, Humans, Macaca mulatta, Mice, Rats, Cerebral Cortex growth & development, Enhancer Elements, Genetic genetics, Epigenesis, Genetic, Evolution, Molecular, Gene Expression Regulation, Developmental, Organogenesis genetics, Promoter Regions, Genetic genetics
- Abstract
Human higher cognition is attributed to the evolutionary expansion and elaboration of the human cerebral cortex. However, the genetic mechanisms contributing to these developmental changes are poorly understood. We used comparative epigenetic profiling of human, rhesus macaque, and mouse corticogenesis to identify promoters and enhancers that have gained activity in humans. These gains are significantly enriched in modules of coexpressed genes in the cortex that function in neuronal proliferation, migration, and cortical-map organization. Gain-enriched modules also showed correlated gene expression patterns and similar transcription factor binding site enrichments in promoters and enhancers, suggesting that they are connected by common regulatory mechanisms. Our results reveal coordinated patterns of potential regulatory changes associated with conserved developmental processes during corticogenesis, providing insight into human cortical evolution., (Copyright © 2015, American Association for the Advancement of Science.)
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- 2015
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33. Coexpression networks implicate human midfetal deep cortical projection neurons in the pathogenesis of autism.
- Author
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Willsey AJ, Sanders SJ, Li M, Dong S, Tebbenkamp AT, Muhle RA, Reilly SK, Lin L, Fertuzinhos S, Miller JA, Murtha MT, Bichsel C, Niu W, Cotney J, Ercan-Sencicek AG, Gockley J, Gupta AR, Han W, He X, Hoffman EJ, Klei L, Lei J, Liu W, Liu L, Lu C, Xu X, Zhu Y, Mane SM, Lein ES, Wei L, Noonan JP, Roeder K, Devlin B, Sestan N, and State MW
- Subjects
- Animals, Brain embryology, Brain growth & development, Brain pathology, Child Development Disorders, Pervasive pathology, Exome, Female, Fetus metabolism, Fetus pathology, Gene Expression Profiling, Genetic Predisposition to Disease, Genome-Wide Association Study, Humans, Male, Mice, Mutation, Neurons metabolism, Prefrontal Cortex metabolism, Sequence Analysis, DNA, Brain metabolism, Child Development Disorders, Pervasive genetics, Child Development Disorders, Pervasive physiopathology
- Abstract
Autism spectrum disorder (ASD) is a complex developmental syndrome of unknown etiology. Recent studies employing exome- and genome-wide sequencing have identified nine high-confidence ASD (hcASD) genes. Working from the hypothesis that ASD-associated mutations in these biologically pleiotropic genes will disrupt intersecting developmental processes to contribute to a common phenotype, we have attempted to identify time periods, brain regions, and cell types in which these genes converge. We have constructed coexpression networks based on the hcASD "seed" genes, leveraging a rich expression data set encompassing multiple human brain regions across human development and into adulthood. By assessing enrichment of an independent set of probable ASD (pASD) genes, derived from the same sequencing studies, we demonstrate a key point of convergence in midfetal layer 5/6 cortical projection neurons. This approach informs when, where, and in what cell types mutations in these specific genes may be productively studied to clarify ASD pathophysiology., (Copyright © 2013 Elsevier Inc. All rights reserved.)
- Published
- 2013
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34. The genomic landscape of cohesin-associated chromatin interactions.
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DeMare LE, Leng J, Cotney J, Reilly SK, Yin J, Sarro R, and Noonan JP
- Subjects
- Animals, Binding Sites, CCCTC-Binding Factor, Chromatin Immunoprecipitation, Enhancer Elements, Genetic, Gene Expression Regulation, Developmental, Genome, Histones metabolism, Limb Buds metabolism, Mice, Mice, Inbred C57BL, Organ Specificity, Promoter Regions, Genetic, Protein Subunits metabolism, Repressor Proteins metabolism, Cohesins, Cell Cycle Proteins metabolism, Chromatin metabolism, Chromosomal Proteins, Non-Histone metabolism
- Abstract
Cohesin is implicated in establishing tissue-specific DNA loops that target enhancers to promoters, and also localizes to sites bound by the insulator protein CTCF, which blocks enhancer-promoter communication. However, cohesin-associated interactions have not been characterized on a genome-wide scale. Here we performed chromatin interaction analysis with paired-end tag sequencing (ChIA-PET) of the cohesin subunit SMC1A in developing mouse limb. We identified 2264 SMC1A interactions, of which 1491 (65%) involved sites co-occupied by CTCF. SMC1A participates in tissue-specific enhancer-promoter interactions and interactions that demarcate regions of correlated regulatory output. In contrast to previous studies, we also identified interactions between promoters and distal sites that are maintained in multiple tissues but are poised in embryonic stem cells and resolve to tissue-specific activated or repressed chromatin states in the mouse embryo. Our results reveal the diversity of cohesin-associated interactions in the genome and highlight their role in establishing the regulatory architecture of development.
- Published
- 2013
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35. The evolution of lineage-specific regulatory activities in the human embryonic limb.
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Cotney J, Leng J, Yin J, Reilly SK, DeMare LE, Emera D, Ayoub AE, Rakic P, and Noonan JP
- Subjects
- Acetylation, Animals, Genetics, Medical, Genome-Wide Association Study, Histones metabolism, Humans, Macaca mulatta embryology, Mice embryology, Organogenesis, Transcriptome, Biological Evolution, Enhancer Elements, Genetic, Extremities embryology, Gene Expression Regulation, Developmental, Promoter Regions, Genetic
- Abstract
The evolution of human anatomical features likely involved changes in gene regulation during development. However, the nature and extent of human-specific developmental regulatory functions remain unknown. We obtained a genome-wide view of cis-regulatory evolution in human embryonic tissues by comparing the histone modification H3K27ac, which provides a quantitative readout of promoter and enhancer activity, during human, rhesus, and mouse limb development. Based on increased H3K27ac, we find that 13% of promoters and 11% of enhancers have gained activity on the human lineage since the human-rhesus divergence. These gains largely arose by modification of ancestral regulatory activities in the limb or potential co-option from other tissues and are likely to have heterogeneous genetic causes. Most enhancers that exhibit gain of activity in humans originated in mammals. Gains at promoters and enhancers in the human limb are associated with increased gene expression, suggesting they include molecular drivers of human morphological evolution., (Copyright © 2013 Elsevier Inc. All rights reserved.)
- Published
- 2013
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36. Chromatin state signatures associated with tissue-specific gene expression and enhancer activity in the embryonic limb.
- Author
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Cotney J, Leng J, Oh S, DeMare LE, Reilly SK, Gerstein MB, and Noonan JP
- Subjects
- Animals, E1A-Associated p300 Protein genetics, E1A-Associated p300 Protein metabolism, Embryo, Mammalian, Extremities physiology, Gene Expression Profiling, Histones genetics, Histones metabolism, Mice, Nucleosomes metabolism, Organ Specificity genetics, Chromatin genetics, Enhancer Elements, Genetic, Extremities embryology, Gene Expression Regulation, Developmental
- Abstract
The regulatory elements that direct tissue-specific gene expression in the developing mammalian embryo remain largely unknown. Although chromatin profiling has proven to be a powerful method for mapping regulatory sequences in cultured cells, chromatin states characteristic of active developmental enhancers have not been directly identified in embryonic tissues. Here we use whole-transcriptome analysis coupled with genome-wide profiling of H3K27ac and H3K27me3 to map chromatin states and enhancers in mouse embryonic forelimb and hindlimb. We show that gene-expression differences between forelimb and hindlimb, and between limb and other embryonic cell types, are correlated with tissue-specific H3K27ac signatures at promoters and distal sites. Using H3K27ac profiles, we identified 28,377 putative enhancers, many of which are likely to be limb specific based on strong enrichment near genes highly expressed in the limb and comparisons with tissue-specific EP300 sites and known enhancers. We describe a chromatin state signature associated with active developmental enhancers, defined by high levels of H3K27ac marking, nucleosome displacement, hypersensitivity to sonication, and strong depletion of H3K27me3. We also find that some developmental enhancers exhibit components of this signature, including hypersensitivity, H3K27ac enrichment, and H3K27me3 depletion, at lower levels in tissues in which they are not active. Our results establish histone modification profiling as a tool for developmental enhancer discovery, and suggest that enhancers maintain an open chromatin state in multiple embryonic tissues independent of their activity level.
- Published
- 2012
- Full Text
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37. Dietary histidine increases mouse skin urocanic acid levels and enhances UVB-induced immune suppression of contact hypersensitivity.
- Author
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Reilly SK and De Fabo EC
- Subjects
- Animals, Dermatitis, Contact immunology, Female, Immune Tolerance radiation effects, Mice, Mice, Inbred BALB C, Skin drug effects, Skin immunology, Skin metabolism, Ultraviolet Rays, Urocanic Acid metabolism, Histidine pharmacology, Immune Tolerance drug effects
- Abstract
Urocanic Acid (UCA) exists in mammalian skin primarily as the trans isomer and is photoisomerized to cis UCA upon UVB absorption. Our previous studies indicated that the photoisomerization of UCA is the initiating event in UBV-induced suppression of cell-mediated immunity (tUCA----cUCA----immune suppression). The purpose of this study was to verify the role of UCA in UV-induced immune suppression of contact hypersensitivity (CHS) in BALB/c mice. Since UCA is a metabolite of the amino acid L-histidine, we reasoned that increased dietary levels of histidine should raise skin tUCA levels. If skin tUCA is the UVB photoreceptor for immune suppression, this increase should enhance UV-induced suppression of CHS. HPLC analysis of skin from BALB/c mice given a histidine-rich diet (10%) showed that the total amount of UCA is significantly higher in these animals than in mice fed a normal diet. Further, levels of suppression of CHS of 3% and 49% in control fed mice, induced by 4.8 and 7.2 kJ/m2 UVB were significantly increased to 21% and 71% respectively in histidine-fed animals at these same UVB doses. These findings provide additional support for the UCA model for immune suppression, and provide the first evidence that UV-induced immune suppression can be enhanced by a dietary component, L-histidine.
- Published
- 1991
- Full Text
- View/download PDF
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