Comprehensive spatiotemporal mapping of single-cell lineages in developing mouse brain by CRISPR-based barcoding

Xie, Lulu; Liu, Hengxin; You, Zhiwen; Wang, Luyue; Li, Yiwen; Zhang, Xinyue; Ji, Xiaoshan; He, Hui; Yuan, Tingli; Zheng, Wei; Wu, Ziyan; Xiong, Man; Wu, Wei; Chen, Yuejun

doi:10.1038/s41592-023-01947-3

Cited by 7 publications

(3 citation statements)

References 69 publications

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“…Recently, CRISPR/Cas9 has been used in combination with DNA barcoding to construct the dynamic trajectories of cell division, which have made it possible to reconstruct the development of lineage relationships at a single-cell resolution [ 135 , 136 , 137 , 138 , 139 ]. In a recent study, researchers developed a lineage-tracing system called CREST (CRISPR editing-based lineage-specific tracing) by the tissue-specific induction of Cas9 expression [ 140 ]. In this way, they precisely mapped the single-cell lineages in developing mouse ventral midbrain, identified the origin of dopaminergic neurons, and demonstrated that the transcriptome defined progenitor cell types with distinct clonal fates and molecular features.…”

Section: Joint Applications With Other Technologiesmentioning

confidence: 99%

The Current Situation and Development Prospect of Whole-Genome Screening

Yang,

Lei,

Ren

et al. 2024

IJMS

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High-throughput genetic screening is useful for discovering critical genes or gene sequences that trigger specific cell functions and/or phenotypes. Loss-of-function genetic screening is mainly achieved through RNA interference (RNAi), CRISPR knock-out (CRISPRko), and CRISPR interference (CRISPRi) technologies. Gain-of-function genetic screening mainly depends on the overexpression of a cDNA library and CRISPR activation (CRISPRa). Base editing can perform both gain- and loss-of-function genetic screening. This review discusses genetic screening techniques based on Cas9 nuclease, including Cas9-mediated genome knock-out and dCas9-based gene activation and interference. We compare these methods with previous genetic screening techniques based on RNAi and cDNA library overexpression and propose future prospects and applications for CRISPR screening.

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Section: Joint Applications With Other Technologiesmentioning

confidence: 99%

The Current Situation and Development Prospect of Whole-Genome Screening

Yang,

Lei,

Ren

et al. 2024

IJMS

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“…This approach enables the unique labeling of each transduced cell, facilitating the study of clonal dynamics [ 120 ], such as in glioma progression [ 121 ]. With future technological advancements, IUE may be adopted for genetic barcode labeling and other recently developed molecular recorders to track various cellular events [ 122 , 123 , 124 , 125 , 126 ].…”

Section: Perspectivesmentioning

confidence: 99%

Advanced Techniques Using In Vivo Electroporation to Study the Molecular Mechanisms of Cerebral Development Disorders

Yang,

Shitamukai,

Yang

et al. 2023

IJMS

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The mammalian cerebral cortex undergoes a strictly regulated developmental process. Detailed in situ visualizations, imaging of these dynamic processes, and in vivo functional gene studies significantly enhance our understanding of brain development and related disorders. This review introduces basic techniques and recent advancements in in vivo electroporation for investigating the molecular mechanisms underlying cerebral diseases. In utero electroporation (IUE) is extensively used to visualize and modify these processes, including the forced expression of pathological mutants in human diseases; thus, this method can be used to establish animal disease models. The advent of advanced techniques, such as genome editing, including de novo knockout, knock-in, epigenetic editing, and spatiotemporal gene regulation, has further expanded our list of investigative tools. These tools include the iON expression switch for the precise control of timing and copy numbers of exogenous genes and TEMPO for investigating the temporal effects of genes. We also introduce the iGONAD method, an improved genome editing via oviductal nucleic acid delivery approach, as a novel genome-editing technique that has accelerated brain development exploration. These advanced in vivo electroporation methods are expected to provide valuable insights into pathological conditions associated with human brain disorders.

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“…mDA neurons are born amongst radial glia (Rgl) in the floor plate (FP) region of the ventral midbrain (VM), and their development is controlled by the interplay of two signaling centers, the isthmic organizer at the midbrain-hindbrain boundary (Joyner et al, 2000), and the midbrain floor plate (mFP) itself (Placzek and Briscoe, 2005). In recent years, single-cell RNA-sequencing (scRNA-seq) technologies have revealed the cellular diversity of the developing VM and provided novel insights into the molecular programs that regulate mDA neuron development (Braun et al, 2023; Kee et al, 2017; La Manno et al, 2021, 2016; Tiklová et al, 2019; Xie et al, 2023). However, while our understanding of cell types in the VM has grown considerably, the specific functions of signaling factors and extracellular matrix (ECM) derived from individual cell types on mDA neuron development and the components that make up the cellular microenvironment or dopaminergic niche remain to be elucidated.…”

Section: Introductionmentioning

confidence: 99%

Midbrain Dopaminergic Neuron Development is Regulated by Two Molecularly Distinct Subtypes of Radial Glia Cells

Ásgrímsdóttir,

Bassini,

di Val Cervo

et al. 2024

Preprint

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Understanding midbrain dopaminergic (mDA) neuron development is key to advancing cell replacement therapies for Parkinson's disease (PD). Recent single-cell RNA-sequencing (scRNA-seq) studies have identified different subtypes of transient radial glia (Rgl) cell types in the developing mouse and human ventral midbrain. However, their individual functions and their impact on mDA neuron development are unclear. Here we analyze the transcriptome of endogenous mouse and human ventral midbrain Rgl and assess the function of key midbrain floor plate Rgl factors in human stem cells during mDA neuron differentiation. We find that Rgl1 is defined by a neurogenic network centered on Arntl, and that this transcription factor regulates human mDA neurogenesis. Conversely, the transcriptome of Rgl3 is dominated by signaling and extracellular matrix molecules that control different aspects of human mDA neuron development. Thus, our results suggest a function of Rgl1 as a mDA progenitor, and of Rgl3 as a signaling mDA niche cell. Moreover, using human stem cells we demonstrate that new knowledge of cell-type specific intrinsic and extrinsic developmental factors can readily be applied to improve the generation of cells with therapeutic interest, such as human mDA neurons for PD.

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Comprehensive spatiotemporal mapping of single-cell lineages in developing mouse brain by CRISPR-based barcoding

Cited by 7 publications

References 69 publications

The Current Situation and Development Prospect of Whole-Genome Screening

The Current Situation and Development Prospect of Whole-Genome Screening

Advanced Techniques Using In Vivo Electroporation to Study the Molecular Mechanisms of Cerebral Development Disorders

Midbrain Dopaminergic Neuron Development is Regulated by Two Molecularly Distinct Subtypes of Radial Glia Cells

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