A programmable sequence of reporters for lineage analysis

García-Marqués, Jorge; Espinosa-Medina, Isabel; Ku, Kai-Yuan; Yang, Ching-Po; Koyama, Minoru; Yu, Hung–Hsiang; Lee, Tzumin

doi:10.1038/s41593-020-0676-9

Cited by 20 publications

(12 citation statements)

References 48 publications

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“…After Cas9 editing and SSA repair, the CaSSA reporter is activated resulting in a frame shift that inactivates the downstream reporter. To activate several of these reporters in a predefined order, we deployed a conditional guide RNA (gRNA) switch (19). Similar to CaSSA switches, a gRNA switch is a disrupted gRNA sequence that gets activated by another gRNA and Cas9 editing followed by SSA repair (Fig.…”

Section: Design and Validation Of Tempomentioning

confidence: 99%

“…1F). For the gRNA switch design, our previous version had ~50% activation efficiency by SSA both in Drosophila and zebrafish (19). However, higher efficiency is desired to avoid limited progression of the cascade.…”

Section: Design and Validation Of Tempomentioning

confidence: 99%

“…In contrast, imaging-based strategies enable spatial identification of cell clones but lack the necessary number of labels to distinguish subsequent cell generations and infer their mitotic connections (18). We recently developed an imaging-based approach in Drosophila called CLADES, which allows differential labelling of sequentially produced neurons and simultaneous visualization of anatomical position and temporal lineage information in the same sample (19).…”

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confidence: 99%

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TEMPO: A system to sequentially label and genetically manipulate vertebrate cell lineages

Espinosa-Medina

Feliciano

Belmonte-Mateos

et al. 2021

Preprint

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During development, regulatory factors appear in a precise order to determine cell fates over time. To investigate complex tissue development, one should not just label cell lineages but further visualize and manipulate cells with temporal control. Current strategies for tracing vertebrate cell lineages lack genetic access to sequentially produced cells. Here we present TEMPO (Temporal Encoding and Manipulation in a Predefined Order), an imaging-readable genetic tool allowing differential labelling and manipulation of consecutive cell generations in vertebrates. TEMPO is based on CRISPR and powered by a cascade of gRNAs that drive orderly activation/inactivation of reporters/effectors. Using TEMPO to visualize zebrafish and mouse neurogenesis, we recapitulated birth-order-dependent neuronal fates. Temporally manipulating cell-cycle regulators in mouse cortex progenitors altered the proportion and distribution of neurons and glia, revealing the effects of temporal gene perturbation on serial cell fates. Thus, TEMPO enables sequential manipulation of molecular factors, crucial to study cell-type specification.One-Sentence SummaryGaining sequential genetic access to vertebrate cell lineages.

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Section: Design and Validation Of Tempomentioning

confidence: 99%

Section: Design and Validation Of Tempomentioning

confidence: 99%

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confidence: 99%

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TEMPO: A system to sequentially label and genetically manipulate vertebrate cell lineages

Espinosa-Medina

Feliciano

Belmonte-Mateos

et al. 2021

Preprint

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“…New approaches involve recombinant DNA constructs that encode different FRs, and that can be transfected into the cells of interest, have allowed all the progeny of single cells to be traced (StarTrack, CLONE, MAGIC, iON) [73][74][75][76]. To resolve the timing of the birth of each cell within the same clone, lineage progression could be determined post hoc by the expression of a predetermined sequence of fluorophores in sibling cells in Drosophila (CLADES) [77]. These methods are based on transposable elements that are integrated into the genome, allowing the stable inheritance of the same barcode by all cell progeny, and avoiding plasmid loss as a consequence of cell division.…”

Section: Multicolor Lineage Tracingmentioning

confidence: 99%

Deciphering neural heterogeneity through cell lineage tracing

Figueres-Oñate

Sánchez-González

López‐Mascaraque

2020

Cell. Mol. Life Sci.

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Understanding how an adult brain reaches an appropriate size and cell composition from a pool of progenitors that proliferates and differentiates is a key question in Developmental Neurobiology. Not only the control of final size but also, the proper arrangement of cells of different embryonic origins is fundamental in this process. Each neural progenitor has to produce a precise number of sibling cells that establish clones, and all these clones will come together to form the functional adult nervous system. Lineage cell tracing is a complex and challenging process that aims to reconstruct the offspring that arise from a single progenitor cell. This tracing can be achieved through strategies based on genetically modified organisms, using either genetic tracers, transfected viral vectors or DNA constructs, and even single-cell sequencing. Combining different reporter proteins and the use of transgenic mice revolutionized clonal analysis more than a decade ago and now, the availability of novel genome editing tools and single-cell sequencing techniques has vastly improved the capacity of lineage tracing to decipher progenitor potential. This review brings together the strategies used to study cell lineages in the brain and the role they have played in our understanding of the functional clonal relationships among neural cells. In addition, future perspectives regarding the study of cell heterogeneity and the ontogeny of different cell lineages will also be addressed.

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“…In the 1980s new methods allowed marking all the descendants of a single cell by the injection of a dye or the expression of a marker gene. Since then, many new methods have been devised to improve cell lineage tracking, including inducible recombinases (Kretzschmar and Watt, 2012), fluorescent or genetic reporters (Kebschull and Zador, 2018;Weissman and Pan, 2015), or a combination of both (Garcia-Marques et al, 2020). However, these approaches come at the cost of resolution, meaning that lineage relationships of individual cells are not fully recovered.…”

Section: Introductionmentioning

confidence: 99%

Benchmarked approaches for reconstruction of in vitro cell lineages and in silico models of C. elegans and M. musculus developmental trees

Gong

Granados

et al. 2021

Cell Systems

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Benchmarked approaches for reconstruction of in vitro cell lineages and in silico models of C. elegans and M. musculus developmental trees Graphical abstract Highlights d We organized a DREAM challenge to benchmark methods of cell lineage reconstruction d Using experimental, in silico datasets as ground-truth trees of 10 2 , 10 3 , and 10 4 cells d Smaller trees allowed the training of a machine-learning decision tree approach d These results delineate a potential way forward for solving larger cell lineage trees

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A programmable sequence of reporters for lineage analysis

Cited by 20 publications

References 48 publications

TEMPO: A system to sequentially label and genetically manipulate vertebrate cell lineages

TEMPO: A system to sequentially label and genetically manipulate vertebrate cell lineages

Deciphering neural heterogeneity through cell lineage tracing

Benchmarked approaches for reconstruction of in vitro cell lineages and in silico models of C. elegans and M. musculus developmental trees

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