Using single-cell RNA sequencing to generate predictive cell-type-specific split-GAL4 reagents throughout development

Proceedings of the National Academy of Sciences, Год журнала: 2023, Номер 120(32)

Опубликована: Июль 31, 2023

Cell-type-specific tools facilitate the identification and functional characterization of distinct cell types that form complexity neuronal circuits. A large collection existing genetic in Drosophila relies on enhancer activity to label different subsets cells has been extremely useful analyzing circuits adults. However, these enhancer-based GAL4 lines often do not reflect expression nearby gene(s) as they only represent a small portion full gene regulatory elements. While intersectional techniques such split-GAL4 system further improve cell-type-specificity, it requires significant time resources screen through combinations patterns. Here, we use developmental single-cell RNA sequencing (scRNAseq) datasets select pairs for provide highly efficient predictive pipeline (scMarco) generate cell-type-specific at any during development, based native These gene-specific can be generated from coding intronic MiMIC/CRIMIC or by CRISPR knock-in. We developing visual model demonstrate high power scRNAseq-guided targeting known types, annotating clusters scRNAseq well identifying novel types. Lastly, are broadly applicable other tissue. Our work opens new avenues generating targeted manipulation throughout development represents valuable resource community.

Язык: Английский

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A split-GAL4 driver line resource forDrosophilaneuron types

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2024, Номер unknown

Опубликована: Янв. 10, 2024

Abstract Techniques that enable precise manipulations of subsets neurons in the fly central nervous system have greatly facilitated our understanding neural basis behavior. Split-GAL4 driver lines allow specific targeting cell types Drosophila melanogaster and other species. We describe here a collection 3060 range adult 1373 characterized third-instar larvae. These tools functional, transcriptomic, proteomic studies based on anatomical targeting. NeuronBridge search relate light microscopy images these split-GAL4 to connectomes reconstructed from electron images. The collections are result screening over 77,000 split hemidriver combinations. Previously published new included, all validated for expression curated optimal type specificity across diverse types. In addition stocks well-characterized lines, we make available 300,000 3D lines.

Язык: Английский

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Split Proteins and Reassembly Modules for Biological Applications

ChemBioChem, Год журнала: 2024, Номер 25(10)

Опубликована: Март 26, 2024

Abstract Split systems, modular entities enabling controlled biological processes, have become instrumental in research. This review highlights their utility across applications like gene regulation, protein interaction identification, and biosensor development. Covering significant progress over the last decade, it revisits traditional split proteins such as GFP, luciferase, inteins, explores advancements technologies Cas base editors. We also examine reassembly modules diverse fields, from regulation to therapeutic innovation. offers a comprehensive perspective on recent evolution of systems

Язык: Английский

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A split-GAL4 driver line resource for Drosophila neuron types

Опубликована: Янв. 6, 2025

Techniques that enable precise manipulations of subsets neurons in the fly central nervous system have greatly facilitated our understanding neural basis behavior. Split-GAL4 driver lines allow specific targeting cell types Drosophila melanogaster and other species. We describe here a collection 3060 range adult 1373 characterized third-instar larvae. These tools functional, transcriptomic, proteomic studies based on anatomical targeting. NeuronBridge search relate light microscopy images these split-GAL4 to connectomes reconstructed from electron images. The collections are result screening over 77,000 split hemidriver combinations. Previously published new included, all validated for expression curated optimal type specificity across diverse types. In addition stocks well-characterized lines, we make available 300,000 3D lines.

Язык: Английский

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Conditional Split Inteins: Adaptable Tools for Programming Protein Functions

International Journal of Molecular Sciences, Год журнала: 2025, Номер 26(2), С. 586 - 586

Опубликована: Янв. 11, 2025

Split inteins are biological mechanisms for the operation of spatiotemporal control protein activities. They function through trans-splicing, in which their N- and C-terminal fragments expressed contiguously with two halves. The subsequent self-excision upon recognition complimentary fragment yields a mature, complete, functional protein. conditional regulation splicing environmental factors or attachment regulatory modules can be used to determine when where will operate, providing potential novel approaches engineering biology applications. This review discuss current split intein applications mechanistic basis species classification. These considerations provide guidance extein activation strategies, design spatial arrangements, taking advantage unique reaction environments. pave way future implementation discoveries selection appropriate aid designing strategies.

Язык: Английский

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A split-GAL4 driver line resource for Drosophila neuron types

eLife, Год журнала: 2025, Номер 13

Опубликована: Янв. 24, 2025

Techniques that enable precise manipulations of subsets neurons in the fly central nervous system (CNS) have greatly facilitated our understanding neural basis behavior. Split-GAL4 driver lines allow specific targeting cell types Drosophila melanogaster and other species. We describe here a collection 3060 range adult CNS 1373 characterized third-instar larvae. These tools functional, transcriptomic, proteomic studies based on anatomical targeting. NeuronBridge search relate light microscopy images these split-GAL4 to connectomes reconstructed from electron images. The collections are result screening over 77,000 split hemidriver combinations. Previously published new included, all validated for expression curated optimal cell-type specificity across diverse types. In addition stocks well-characterized lines, we make available 300,000 3D lines.

Язык: Английский

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Cellular Dissociation of Ecdysteroid-DopEcR Signaling for Toxin Avoidance and Detoxification

Опубликована: Янв. 1, 2025

Язык: Английский

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eLife assessment: A split-GAL4 driver line resource for Drosophila CNS cell types

Опубликована: Июль 30, 2024

Techniques that enable precise manipulations of subsets neurons in the fly central nervous system have greatly facilitated our understanding neural basis behavior. Split-GAL4 driver lines allow specific targeting cell types Drosophila melanogaster and other species. We describe here a collection 3060 range adult 1373 characterized third-instar larvae. These tools functional, transcriptomic, proteomic studies based on anatomical targeting. NeuronBridge search relate light microscopy images these split-GAL4 to connectomes reconstructed from electron images. The collections are result screening over 77,000 split hemidriver combinations. In addition stocks for well-characterized lines, we make available 300,000 new 3D lines.

Язык: Английский

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Hox gene–specific cellular targeting using split intein Trojan exons

Proceedings of the National Academy of Sciences, Год журнала: 2024, Номер 121(17)

Опубликована: Апрель 11, 2024

The Trojan exon method, which makes use of intronically inserted T2A-Gal4 cassettes, has been widely used in Drosophila to create thousands gene-specific Gal4 driver lines. These dual-purpose lines provide genetic access specific cell types based on their expression a native gene while simultaneously mutating one allele the enable loss-of-function analysis homozygous animals. While this dual is often an advantage, truncation mutations produced by exons are sometimes deleterious heterozygotes, perhaps creating translation products with dominant negative effects. Such mutagenic effects can cause developmental lethality as observed genes encoding essential transcription factors. Given importance factors specifying type, alternative techniques for generating that target them required. Here, we introduce modified method retains targeting fidelity and plug-and-play modularity original but mitigates its exploiting self-splicing capabilities split inteins. “Split Intein exons” (siTrojans) ensure two generated from interrupted trans-spliced full-length protein. We demonstrate efficacy siTrojans comprehensive toolkit Split segmentally expressed Hox illustrate neural circuit mapping neurons according position along anterior–posterior axis. Both gene–specific introduced here should be broadly useful.

Язык: Английский

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A split-GAL4 driver line resource for Drosophila CNS cell types

Опубликована: Июль 30, 2024

Techniques that enable precise manipulations of subsets neurons in the fly central nervous system have greatly facilitated our understanding neural basis behavior. Split-GAL4 driver lines allow specific targeting cell types Drosophila melanogaster and other species. We describe here a collection 3060 range adult 1373 characterized third-instar larvae. These tools functional, transcriptomic, proteomic studies based on anatomical targeting. NeuronBridge search relate light microscopy images these split-GAL4 to connectomes reconstructed from electron images. The collections are result screening over 77,000 split hemidriver combinations. In addition stocks for well-characterized lines, we make available 300,000 new 3D lines.

Язык: Английский

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