A robust and tunable system for targeted cell ablation in developing embryos

Labbaf, Zahra; Petratou, Kleio; Ermlich, Laura; Backer, Wilko; Tarbashevich, Katsiaryna; Reichman-Fried, Michal; Luschnig, Stefan; Schulte‐Merker, Stefan; Raz, Erez

doi:10.1016/j.devcel.2022.07.008

Cited by 7 publications

(7 citation statements)

References 61 publications

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“…The transgene transcripts are expressed in the otxG+ in predominantly ectoderm and in the endomesoderm from early blastula stage continues to gastrula. The applications of this two plasmid Tet-ON system are in principle tunable to a vast range of ectopic gene expressions, such as (fluorescence reporter, calcium indicators, recombinases, endonucleases), targeted cell ablation (Labbaf et al, 2022), controllable functional inactivation of genes by dominant negative mutants. These tools could accelerate the use of sea star as a scalable invertebrate model in developmental, regeneration, aging studies and cancer biology.…”

Section: Discussionmentioning

confidence: 99%

Induciblein vivogenome editing in the sea starPatiria miniata

Zueva

Hinman

2023

Preprint

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For centuries, echinoderms, a marine-invertebrate phylum, have fascinated scientists for their developmental and postembryonic phenomen. Experimentation on their eggs and embryos in particular have contributed foundation scientific advances. However, powerful molecular genetic studies are restricted to embryonic developmental stages which are amenable to genetic perturbation by microinjection of reagents into the zygotes. This represents a significant bottleneck to the study of postembryonic processes in where the earliest function of a gene must remain intact. We therefore sought to establish a spatio-temporal turnable gene editing tool for these species. Here, using the sea star Patiria miniata as a model we introduce a chemically inducible, Tet-ON, gene expression system. Pairing this Tet-ON system with CRISPR-mediated gene alteration technology we show as a proof-of-principle demonstration an inducible gene editing in the sea star transgenic cell populations for the first time in echinoderm biology. The approach we show here can be adapted for use in other species of echinoderms and will also extend experimental possibilities tremendously.

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Section: Discussionmentioning

confidence: 99%

Induciblein vivogenome editing in the sea starPatiria miniata

Zueva

Hinman

2023

Preprint

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“…If this is the case, then a reduction in spinal cord morphogenesis anterior to the tailbud should also affect the length of the tail tissues. To reduce spinal cord elongation along the whole body we turned to a genetic ablation technique for tissue-specific expression of the bacterial Kid toxin (Labbaf et al, 2022). The expression of UAS:Kid in the spinal cord was driven by an Adamts3:GAL4FF (ATS3) transgene which has strong expression in motor neurons (Asakawa et al, 2013; Wang et al, 2020).…”

Section: Resultsmentioning

confidence: 99%

“…ATS3:GAL4 (Wang et al, 2020) were crossed with either UAS:Kid (Labbaf et al, 2022) or UAS:GFP fish to produce embryos that ATS3:GAL4;UAS:Kid and ATS3:GAL4;UAS:GFP. This causes the expression of the bacterial Kid toxin or GFP in AdamTS3 expressing cells of the spinal cord.…”

Section: Methodsmentioning

confidence: 99%

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Spinal Cord Elongation Enables Proportional Regulation of the Zebrafish Posterior Body

Saunders,

Camacho,

Steventon

2024

Preprint

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Early embryos display a remarkable ability to regulate the patterning of tissues in response to changes in tissue size. However, it is not clear whether this ability continues into post-gastrulation stages upon cell commitment to distinct germ layers. Here, we performed targeted removal of neural fated cells in the zebrafish tailbud using multi-photon ablation. This led to a proportional reduction in the length of both the spinal cord and paraxial mesoderm in the tail, revealing a capacity to regulate tissue morphogenesis across multiple tissues to build a well-proportioned posterior body. Following analysis of cell proliferation, gene expression, signalling and cell movements, we found no evidence of cell fate switching from mesoderm to neural fate to compensate for neural progenitor loss. Furthermore, we found that tail paraxial mesoderm length is not reduced upon direct removal of an equivalent number of mesoderm progenitors, ruling out the hypothesis that neuromesodermal competent cells enable proportional regulation. Instead, reduction in the numbers of cells across the spinal cord reduces both spinal cord and paraxial mesoderm length. We conclude that spinal cord elongation is a driver of paraxial mesoderm elongation in the zebrafish posterior body and that this can explain proportional regulation of both tissues upon neural progenitor reduction.

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“…In vertebrates, the notochord exchanges patterning cues with the sclerotome, which gives rise to the vertebral bodies; as the vertebral column develops, the notochord regresses and its remnants form the nuclei pulposi , the central-most region of the intervertebral discs, which are highly hydrated and possess shock-absorbing properties 4 . Although its evolutionary origins and its relation to analogous structures in non-chordate phyla remain still debated 5 , 6 , the severe effects of notochord ablation 7 , 8 and malformations 9 – 12 unequivocally assert the role of the notochord in embryonic development. The widespread pivotal role of the notochord in chordate development renders studies of the gene regulatory network (GRN) underlying its formation both compelling and broadly informative.…”

Section: Introductionmentioning

confidence: 99%

Cis-regulatory interfaces reveal the molecular mechanisms underlying the notochord gene regulatory network of Ciona

Negrón-Piñeiro,

Wu,

Popsuj

et al. 2024

Nat Commun

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Tissue-specific gene expression is fundamental in development and evolution, and is mediated by transcription factors and by the cis-regulatory regions (enhancers) that they control. Transcription factors and their respective tissue-specific enhancers are essential components of gene regulatory networks responsible for the development of tissues and organs. Although numerous transcription factors have been characterized from different organisms, the knowledge of the enhancers responsible for their tissue-specific expression remains fragmentary. Here we use Ciona to study the enhancers associated with ten transcription factors expressed in the notochord, an evolutionary hallmark of the chordate phylum. Our results illustrate how two evolutionarily conserved transcription factors, Brachyury and Foxa2, coordinate the deployment of other notochord transcription factors. The results of these detailed cis-regulatory analyses delineate a high-resolution view of the essential notochord gene regulatory network of Ciona, and provide a reference for studies of transcription factors, enhancers, and their roles in development, disease, and evolution.

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A robust and tunable system for targeted cell ablation in developing embryos

Cited by 7 publications

References 61 publications

Induciblein vivogenome editing in the sea starPatiria miniata

Induciblein vivogenome editing in the sea starPatiria miniata

Spinal Cord Elongation Enables Proportional Regulation of the Zebrafish Posterior Body

Cis-regulatory interfaces reveal the molecular mechanisms underlying the notochord gene regulatory network of Ciona

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