Engineered miniature CRISPR-Cas system for mammalian genome regulation and editing

Molecular Cell, Journal Year: 2021, Volume and Issue: 81(20), P. 4333 - 4345.e4

Published: Sept. 3, 2021

Compact and versatile CRISPR-Cas systems will enable genome engineering applications through high-efficiency delivery in a wide variety of contexts. Here, we create an efficient miniature Cas system (CasMINI) engineered from the type V-F Cas12f (Cas14) by guide RNA protein engineering, which is less than half size currently used CRISPR (Cas9 or Cas12a). We demonstrate that CasMINI can drive high levels gene activation (up to thousands-fold increases), while natural fails function mammalian cells. show has comparable activities Cas12a for activation, highly specific, allows robust base editing editing. expect be broadly useful cell therapy ex vivo vivo.

Language: Английский

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The CRISPR-Cas toolbox and gene editing technologies

Molecular Cell, Journal Year: 2021, Volume and Issue: 82(2), P. 333 - 347

Published: Dec. 29, 2021

Language: Английский

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A new era in functional genomics screens

Nature Reviews Genetics, Journal Year: 2021, Volume and Issue: 23(2), P. 89 - 103

Published: Sept. 20, 2021

Language: Английский

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Advances in CRISPR therapeutics

Nature Reviews Nephrology, Journal Year: 2022, Volume and Issue: 19(1), P. 9 - 22

Published: Oct. 24, 2022

Language: Английский

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Ethical implications of epigenetics in the era of personalized medicine

Clinical Epigenetics, Journal Year: 2022, Volume and Issue: 14(1)

Published: March 25, 2022

Abstract Given the increasing research activity on epigenetics to monitor human diseases and its connection with lifestyle environmental expositions, field of has attracted a great deal interest also at ethical societal level. In this review, we will identify discuss current ethical, legal social issues in context personalized medicine. The review covers aspects such as how epigenetic information should impact patient autonomy ability generate an intentional voluntary decision, measures data protection related privacy confidentiality derived from epigenome studies (e.g., risk discrimination, re-identification unexpected findings) or debate distribution responsibilities for health (i.e., personal versus public responsibilities). We pay special attention discrimination stigmatization consequence inferring exposures potentially contained data. Furthermore, environment individual habits do not affect all populations equally, violation principle distributive justice access benefits clinical is discussed. regard, represents opportunity integration policy aimed create healthier living environments. Whether these policies coexist or, contrast, compete strategies reinforcing medicine interventions needs be considered. ends reflection main challenges research, some them technical dimension assessing causality establishing reference epigenomes) but sphere add determinism top genetic one). sum, into life science investigation experiences exposure risk, nutritional habits, prejudice stigma, imperative understand variation disease. This pragmatic approach required locate out experimental laboratories facilitate implementation society.

Language: Английский

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Predicting transcriptional outcomes of novel multigene perturbations with GEARS

Nature Biotechnology, Journal Year: 2023, Volume and Issue: 42(6), P. 927 - 935

Published: Aug. 17, 2023

Abstract Understanding cellular responses to genetic perturbation is central numerous biomedical applications, from identifying interactions involved in cancer developing methods for regenerative medicine. However, the combinatorial explosion number of possible multigene perturbations severely limits experimental interrogation. Here, we present graph-enhanced gene activation and repression simulator (GEARS), a method that integrates deep learning with knowledge graph gene–gene relationships predict transcriptional both single using single-cell RNA-sequencing data perturbational screens. GEARS able outcomes perturbing combinations consisting genes were never experimentally perturbed. exhibited 40% higher precision than existing approaches predicting four distinct interaction subtypes screen identified strongest twice as well prior approaches. Overall, can phenotypically effects thus guide design experiments.

Language: Английский

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Mitigation of chromosome loss in clinical CRISPR-Cas9-engineered T cells

Cell, Journal Year: 2023, Volume and Issue: 186(21), P. 4567 - 4582.e20

Published: Oct. 1, 2023

CRISPR-Cas9 genome editing has enabled advanced T cell therapies, but occasional loss of the targeted chromosome remains a safety concern. To investigate whether Cas9-induced is universal phenomenon and evaluate its clinical significance, we conducted systematic analysis in primary human cells. Arrayed pooled CRISPR screens revealed that was generalizable across resulted partial entire chromosome, including preclinical chimeric antigen receptor cells with persisted for weeks culture, implying potential to interfere use. A modified manufacturing process, employed our first-in-human trial Cas9-engineered (NCT03399448), reduced while largely preserving efficacy. Expression p53 correlated protection from observed this protocol, suggesting both mechanism strategy engineering mitigates genotoxicity clinic.

Language: Английский

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Durable and efficient gene silencing in vivo by hit-and-run epigenome editing

Nature, Journal Year: 2024, Volume and Issue: 627(8003), P. 416 - 423

Published: Feb. 28, 2024

Abstract Permanent epigenetic silencing using programmable editors equipped with transcriptional repressors holds great promise for the treatment of human diseases 1–3 . However, to unlock its full therapeutic potential, an experimental confirmation durable after delivery transient in vivo is needed. To this end, here we targeted Pcsk9 , a gene expressed hepatocytes that involved cholesterol homeostasis. In vitro screening different editor designs indicated zinc-finger proteins were best-performing DNA-binding platform efficient mouse A single administration lipid nanoparticles loaded editors’ mRNAs almost halved circulating levels PCSK9 nearly one year mice. Notably, and accompanying repressive marks also persisted forced liver regeneration, further corroborating heritability newly installed state. Improvements construct design resulted development all-in-one configuration term evolved engineered repressor (EvoETR). This design, which characterized by high specificity profile, reduced mice efficiency comparable obtained through conventional editing, but without causing DNA breaks. Our study lays foundation therapeutics are based on silencing.

Language: Английский

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Large-scale manipulation of promoter DNA methylation reveals context-specific transcriptional responses and stability

Genome biology, Journal Year: 2022, Volume and Issue: 23(1)

Published: July 26, 2022

Abstract Background Cytosine DNA methylation is widely described as a transcriptional repressive mark with the capacity to silence promoters. Epigenome engineering techniques enable direct testing of effect induced on endogenous promoters; however, downstream effects have not yet been comprehensively assessed. Results Here, we simultaneously induce at thousands promoters in human cells using an engineered zinc finger-DNMT3A fusion protein, enabling us test forced upon transcription, chromatin accessibility, histone modifications, and persistence after removal protein. We find that responses are highly context-specific, including lack repression, well cases increased gene expression, which appears be driven by eviction methyl-sensitive repressors. Furthermore, some regulatory networks can override promoter cause alternative usage. deposited distal regions rapidly erased process combining passive TET-mediated demethylation. Finally, demonstrate exist nucleosomes possess active modification H3K4me3, or bound initiated form RNA polymerase II. Conclusions These findings important implications for epigenome response more complex than previously appreciated.

Language: Английский

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Plant breeding advancements with “CRISPR-Cas” genome editing technologies will assist future food security

Frontiers in Plant Science, Journal Year: 2023, Volume and Issue: 14

Published: March 13, 2023

Genome editing techniques are being used to modify plant breeding, which might increase food production sustainably by 2050. A product made feasible genome is becoming better known, because of looser regulation and widespread acceptance. The world’s population supply would never have increased proportionally under current farming practices. development plants has been greatly impacted global warming climate change. Therefore, minimizing these effects crucial for agricultural that sustainable. Crops more resilient abiotic stress sophisticated practices a understanding the response mechanism. Both conventional molecular breeding create viable crop types both processes time-consuming. Recently, breeders shown an interest in approaches genetic manipulation use clustered regularly interspaced short palindromic repeats (CRISPR/Cas9). To ensure security future, kinds with desired traits must be developed. completely new era begun revolution based on CRISPR/CRISPR-associated nuclease (Cas9) systems. All may effectively target particular gene or group loci using Cas9 single-guide RNA (sgRNA). CRISPR/Cas9 can thereby save time labor compared methods. An easy, quick, efficient method directly altering sequences cells CRISPR CRISPR-Cas9 system, was developed from components earliest known bacterial immune allows targeted breakage variety cells/RNA guide endonuclease cleavage specificity system. Editing directed practically any genomic site (gRNA) sequence delivering it cell along endonuclease. We summarize recent research findings, investigate potential applications make predictions about likely future breakthroughs through

Language: Английский

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