CRISPR-Cas9 genome editing articles within Nature Communications

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  • Article
    | Open Access

    CRISPR gene activation and inhibition has become a powerful synthetic tool for influencing the expression of native genes for foundational studies, cellular reprograming, and metabolic engineering. Here the authors demonstrate near leak-free, inducible expression of a polycistronic array containing up to 24 gRNAs from two orthogonal CRISPR/Cas systems.

    • William M. Shaw
    • , Lucie Studená
    •  & Rodrigo Ledesma-Amaro

  • Article
    | Open Access

    As CRISPR-based therapies enter the clinic, evaluation of safety remains a critical and active area of study. Here the authors use next generation sequencing to achieve high sequencing depth and demonstrate that clinically relevant delivery of high-fidelity Cas9 to primary HSPCs and ex vivo culture up to 10 days does not introduce or enrich for tumorigenic variants.

    • M. Kyle Cromer
    • , Valentin V. Barsan
    •  & Matthew H. Porteus

  • Article
    | Open Access

    Base editing is promising for gene therapy, but in vivo delivery has been limiting. Here the authors perform structure-based rational engineering of the cytosine base editing system Target-AID to minimise off-target effects and decrease its size.

    • Ang Li
    • , Hitoshi Mitsunobu
    •  & Keiji Nishida

  • Article
    | Open Access

    CRISPR-Cas induced HDR methods tend to have a low efficiency. Here the authors report an HDR improvement strategy, Recursive Editing, that selectively retargets undesired indel outcomes to create additional opportunities for HDR; they introduce REtarget, a tool for Recursive Editing experimental design.

    • Lukas Möller
    • , Eric J. Aird
    •  & Jacob E. Corn

  • Article
    | Open Access

    Paeniclostridium sordellii is an opportunistic pathogen that can occur and be fatal in women undergoing abortion or childbirth. The pathogenesis of a hemorrhagic toxin, TcsH, produced by this bacteria, remains unknown. Here, authors carry out genome-wide screens to identify pathologically relevant host factors of TcsH.

    • Xingxing Li
    • , Liuqing He
    •  & Liang Tao

  • Article
    | Open Access

    Understanding how DNA double strand breaks (DSBs) form and are repaired in the genome depends on their accurate measurement. Here the authors describe INDUCE-seq; a DSB-detection method that simultaneously measures physiological and induced breaks throughout the genome.

    • Felix M. Dobbs
    • , Patrick van Eijk
    •  & Simon H. Reed

  • Article
    | Open Access

    DNAPKcs and its kinase activity are required for blunt DNA break end joining when the bridging factor XLF is weakened, but for homologous recombination and radiation resistance, the influence of DNAPKcs is not further enhanced with loss of XLF.

    • Metztli Cisneros-Aguirre
    • , Felicia Wednesday Lopezcolorado
    •  & Jeremy M. Stark

  • Article
    | Open Access

    The identification of events occurring at target sites is critical to determine the safety of CRISPRbased DNA editing tools. Here, the authors show that LINE-1 retrotranspositions can occur frequently at canonical CRISPR/Cas9 editing sites, but are rare with prime editors and base editors.

    • Jianli Tao
    • , Qi Wang
    •  & Roberto Chiarle

  • Article
    | Open Access

    The CRISPR/Cas system has emerged as a powerful and versatile genome engineering tool. Here the authors couple Cas9 to effector protein Exonuclease III via coiled-coil mediated interactions, termed CCExo, leading to increased deletion sizes and enhanced gene knock-out efficiencies in cell lines, primary cells and in vivo.

    • Duško Lainšček
    • , Vida Forstnerič
    •  & Roman Jerala

  • Article
    | Open Access

    Prime editing enables search-and-replace genome editing but is limited by low editing efficiency. Here the authors present PepSEq, a high-throughput method for screening a large library of peptides that influence prime editing efficiency.

    • Minja Velimirovic
    • , Larissa C. Zanetti
    •  & Richard I. Sherwood

  • Article
    | Open Access

    The recent development of PAM-less base editors makes it possible to assess the functional impact and pathogenicity of nucleotide mutations in animals. Here the authors show that SpG and SpRY could edit NGN and NNN PAMs in zebrafish using the purified protein with synthetically modified gRNA, respectively.

    • Fang Liang
    • , Yu Zhang
    •  & Wei Qin

  • Article
    | Open Access

    The need for diverse chromosomal modifications in biotechnology, synthetic biology and basic research requires the development of new technologies. Here the authors present CRISPR SWAPnDROP, which extends the limits of genome editing to large-scale in-vivo DNA transfer between bacterial species.

    • Marc Teufel
    • , Carlo A. Klein
    •  & Patrick Sobetzko

  • Article
    | Open Access

    The ability to alter the genomes of living cells is key to understanding how genes influence the functions of organisms and will be critical to modify living systems for useful purposes. Here, the authors use computational design to discover Cas9 enzymes with increased activity.

    • Pascal D. Vos
    • , Giulia Rossetti
    •  & Oliver Rackham

  • Article
    | Open Access

    Cas12i is a genome editing platform with compact size that fits in AAV vector with short 43-mer gRNA, absence of tracrRNA, ability to process pre-crRNA, and high specificity. Here the authors present an unbiased mutational scanning approach to engineer Cas12i, which shows low activity in mammalian cells, and identify single substitutions that significantly improve indel activity.

    • Colin McGaw
    • , Anthony J. Garrity
    •  & Shaorong Chong

  • Article
    | Open Access

    Current base- and prime-editing technologies lack efficient strategies to edit multiple genomic loci simultaneously, limiting their applications in complex genomics and polygenic diseases. Here the authors describe drive-and-process CRISPR array architectures for multiplex base-editing and multiplex prime-editing in human cells.

    • Qichen Yuan
    •  & Xue Gao

  • Article
    | Open Access

    CRISPR gene drives are genetic elements capable of quickly spreading through populations and they offer promising solutions for curbing the spread of vector-borne diseases and controlling crop pest and invasive species populations. Here the authors present a method for overcoming resistance alleles “double-tap,” that encodes additional gRNAs in the gene drive that target the most common generated resistance alleles.

    • Alena L. Bishop
    • , Víctor López Del Amo
    •  & Valentino M. Gantz

  • Article
    | Open Access

    Programmable double-strand DNA breaks (DSBs) can be harnessed for precision genome editing through manipulation of the homology-directed repair (HDR) pathway. Here the authors report the development of the double tap - double tap implements secondary gRNAs which target Cas9 to common indel sequences and provides a second chance at HDR.

    • Zsolt Bodai
    • , Alena L. Bishop
    •  & Alexis C. Komor

  • Article
    | Open Access

    Combinatorial CRISPR screens can be utilized to identify genetic interactions and functional redundancies of multiple genes. Here, the authors benchmark ten digenic CRISPR technologies and identify novel Cas9 tracrRNA combinations that show superior performance.

    • Ruitong Li
    • , Olaf Klingbeil
    •  & William R. Sellers

  • Article
    | Open Access

    Here, Mac Kain and Maarifi et al. perform a functional CRISPR/Cas9 screen to identify SARS-CoV-2 restriction factors in A549 cells. They identify DAXX, a scaffold protein of nuclear bodies with diverse functions, that has anti-viral activity post SARS-CoV-2 entry, while SARS-CoV-2 has evolved a mechanism to counteract its action via PLpro-mediated proteasomal degradation.

    • Alice Mac Kain
    • , Ghizlane Maarifi
    •  & Ferdinand Roesch

  • Article
    | Open Access

    Targeting integrin-mediated retention of malignant B cells in their protective microenvironment is an efficacious treatment for lymphoma and leukemia. Here, the authors present an unbiased loss-of-adhesion CRISPR screening method, identifying therapeutic targets for these B-cell malignancies.

    • Martin F. M. de Rooij
    • , Yvonne J. Thus
    •  & Marcel Spaargaren

  • Article
    | Open Access

    RNA modifications, including N6-methyladenosine (m6A), have been reported to regulate fundamental RNA processes and properties, and directly linked to various human diseases. Here, the authors develop a chemically inducible and reversible RNA m6A modification editing platform integrating chemically induced proximity (CIP) and CRISPR methods.

    • Huaxia Shi
    • , Ying Xu
    •  & Fu-Sen Liang

  • Article
    | Open Access

    The prime editors (PEs) have shown great promise for precise genome modification. Here the authors place a stabilizing viral xrRNA motif to the 3′ of pegRNAs to enhance editing efficiencies.

    • Guiquan Zhang
    • , Yao Liu
    •  & Jianghuai Liu

  • Article
    | Open Access

    Prime editors can mediate all twelve types of base substitutions and small insertions or deletions in living cells but its efficiency remains low. Here the authors introduce same-sense mutations into pegRNAs to increase base-editing efficiency and the pegRNA secondary structure was altered to increase indel-editing efficiency.

    • Xiaosa Li
    • , Lina Zhou
    •  & Jia Chen

  • Article
    | Open Access

    Base editors are genome engineering tools that can generate nucleotide substitutions without introducing double-stranded breaks. Here the authors show that a phage-derived peptidyl CRISPR inhibitor can be employed to modulate the activity and targeting scope of CRISPR base editor for precision base editing applications.

    • Kun Jia
    • , Yan-ru Cui
    •  & Jia Liu

  • Article
    | Open Access

    Gene editing tools have tremendous potential for biomedical and basic research. Here the authors report a Cas9 from Faecalibaculum rodentium (FrCas9) that achieves efficient and specific gene editing in human cells with a NNTA palindrome PAMs for targeting optimal sites at TATA-boxes to enhance CRISPRa/i screening.

    • Zifeng Cui
    • , Rui Tian
    •  & Zheng Hu

  • Article
    | Open Access

    Cas9 off-target sites can be predicted by many bioinformatics tools. Here the authors present low complexity mechanistic model that characterizes SpCas9 kinetics in free-energy terms, allowing quantitative prediction of off-target activity in bulk-biochemistry, single molecule, and whole-genome profiling experiments.

    • Behrouz Eslami-Mossallam
    • , Misha Klein
    •  & Martin Depken

  • Article
    | Open Access

    Numerous rationally-designed and directed-evolution variants of SpCas9 have been reported to expand the utility of CRISPR technology. Here the authors make comparisons of numerous Cas9 variants, nominate options for base editing screens with denser coverage with A>G and C>T base editing screens and identify loss-of-function mutations in BRCA1 and Venetoclax-resistant mutations in BCL2.

    • Annabel K. Sangree
    • , Audrey L. Griffith
    •  & John G. Doench

  • Article
    | Open Access

    The off-target effects of CRISPR-Cas9 are thought to be mediated by its cognate guide RNA. Here the authors show that Cas9 independently interacts with the human transcriptome, correlating with elevated RNA editing even under guide RNA co-expression.

    • Aaron A. Smargon
    • , Assael A. Madrigal
    •  & Gene W. Yeo

  • Article
    | Open Access

    While prime editing is a promising technique, some genomic sites remain difficult to edit. Here the authors present fluoPEER, fluorescent prime editing and enrichment reporter, to rank the efficiency of pegRNAs and prime editor variants.

    • I. F. Schene
    • , I. P. Joore
    •  & S. A. Fuchs

  • Article
    | Open Access

    T follicular helper (Tfh) and T help type 1 (Th1) cells both arise from naïve CD4 T cells, but detailed knowledge of their differentiation remains incomplete. Here the authors pursue an in vivo CRISPR screen to identify genes, focusing on druggable targets, regulating Tfh versus Th1 to provide a resource for related studies, while also implicating HIF-1α and VHL in this regulation.

    • Bonnie Huang
    • , James D. Phelan
    •  & Pamela L. Schwartzberg

  • Article
    | Open Access

    Biocontainment is a key to developing safe genetically-engineered microbes (GEMs). Here the authors demonstrate genetically stable CRISPR-based kill switches that control GEMs’ viability in animal hosts, enabling their safe biomedical applications.

    • Austin G. Rottinghaus
    • , Aura Ferreiro
    •  & Tae Seok Moon

  • Article
    | Open Access

    Anti-deaminases can inhibit APOBEC3, a component of cytosine base editors. Here Zhanjun Li and colleagues repurposed anti-deaminase proteins derived from viruses to inhibit base editors for use in efficient regulation of base editors’ activity in gene modification and therapeutic applications.

    • Zhiquan Liu
    • , Siyu Chen
    •  & Zhanjun Li

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