Series on CRISPR technology

Qi and colleagues review CRISPR-based epigenome engineering technologies to modulate histone and DNA modifications and to perturb DNA and RNA regulatory elements and chromatin organization.

RNA-targeting CRISPR. In this Perspective, Smargon, Shi and Yeo discuss the rapid development of the RNA-targeting CRISPR–Cas engineering system and highlight how this can be leveraged to further understand RNA biology.

In this Perspective, Lea and Niakan describe advances in CRISPR/Cas9 genome editing techniques and discuss ethical questions and potential clinical implications of this technology.

Harnessing DNA repair pathways in genome editing In this Review, Yeh, Richardson and Corn discuss the DNA repair pathways that underlie genome editing and recent improvements and strategies to yield desired genomic alterations in cells and organisms.

Advances in CRISPR-based systems have greatly expanded the molecular toolbox for biologists. In this issue, we present the first of a Series of commissioned Review articles that highlight the progress made using CRISPR–Cas9 technology and its relevance for cell biological research.

Wang et al. developed a transformer base editor system in which the enzyme activity of the base editor is turned on only at the on-target site, therefore minimizing genome-wide and transcriptome-wide off-target mutations.

Gao et al. developed a CRISPR–Cas9-based system in which sgRNA production is controlled by the endogenous promoter to monitor the expression of weakly expressed genes and long non-coding RNAs in mammalian cells.

Moghadam et al. developed a CRISPR transcriptional repressor to silence MyD88 expression in vivo to modulate immune response against AAV gene therapy and septicaemia.

Li and colleagues report base editor variants with improved targeting efficiencies and broader editing windows by fusing the original base editors with the single-stranded DNA-binding domain of Rad51.

CRISPR base editors can induce single-base-pair changes in the genome, although they are often inefficient. A study now shows that fusion of the DNA-binding domain of RAD51 to base editors enhances both the efficiency and the targeting range of optimized enzymes. These ‘hyper-editors’ offer effective tools for disease modeling and gene therapy.

Artegiani, Hendriks et al. describe a CRISPR–Cas9-based method to efficiently generate human knock-in organoids using non-homologous end joining to study rare intestinal cell types and human hepatocyte division.

Li et al. demonstrate the efficacy of correcting the mutated TERT promoter using a programmable base editing, highlighting its ability to induce senescence, arrest and regression in brain tumour models.

Fu et al. report the transition of ESCs into a 2-cell-embyro-like state induced by Dux involves two steps and can be prevented by Myc and Dnmt1, which inhibit the downregulation of pluripotency genes and the activation of 2C⁺-upregulated elements.

Atlasi et al. demonstrate that transcriptome and epigenome resetting during serum-to-2i pluripotency conversion involves minimal enhancer–promoter rewiring and is instead linked to 2i-enhancer activation via Esrrb-mediated H3K27 acetylation.

MicroRNAs (miRNAs) repress target mRNAs, often with exquisite tissue specificity. Wang et al. exploit the specific expression of miRNAs to regulate guide production for Cas9. Their method enables novel strategies to simultaneously measure the activity of multiple miRNAs and restrict Cas9 binding or genome editing to precisely defined cell types.

Wang et al. developed an inducible CRISPR–Cas9 system, in which guide RNA release is controlled by specific microRNAs, and demonstrated its application as a microRNA sensor and cell-type-specific genome regulator.

Li and colleagues develop a CRISPR–Cas9-based screen strategy that combines base editing and haploid embryonic stem cell technologies to identify amino acids critical for protein function in mice.

Functional genetic screening of mice and other mammals is exceedingly challenging. A CRISPR-based mutagenesis screen in mice has successfully revealed amino acids vital for protein function of the DND1 gene, missense mutations of which lead to defects in primordial germ cell development.

Through CRISPR–Cas9 screen, Dev et al. identified that SHLD1/2 inhibition contributes to PARP-inhibitor resistance. Mechanistically, SHLDs promote non-homologous end-joining and antagonize homologous recombination.

Using a multi-tier scRNA-seq and CRISP-seq approach, Giladi et al. define a transcriptional signature for the naive haematopoietic stem cell state, and follow progenitor plasticity and fate commitment under the influence of cytokines and growth factors.

Yilmaz et al. generate a genome-wide loss-of-function library using human haploid embryonic stem cells and define genes that are essential for cell survival, growth and pluripotency maintenance, as well as growth-restricting genes.

Hicks et al. compare human pluripotent stem cell (hPSC)-derived muscle progenitors to fetal muscle cells, identify ERBB3/NGFR⁺ populations with improved myogenic potential in vivo and enhance cell maturation by inhibiting TGF-β signalling during directed differentiation.