2023-07-31 ニューヨーク大学 (NYU)
Depiction of a fruit fly head and the clusters of single-cell RNA sequencing of developing visual system. Image credit: Monika Jasnauskaite
◆従来は、ショウジョウバエの視覚系に約200種類の細胞タイプがあることが知られていましたが、そのうち約半数しか特定できていませんでした。しかし、この新しい手法を使うことで、残りの細胞タイプも特定することができました。
◆今後、これらの新しいツールを使って他の生物の研究にも応用できる可能性があります。また、この研究は、脳の発達と機能に関する理解を進める上で重要な知見をもたらすものとされています。
<関連情報>
- https://www.nyu.edu/about/news-publications/news/2023/july/how-flies-develop-sight.html
- https://www.pnas.org/doi/10.1073/pnas.2307451120
単一細胞RNA配列決定法を用いて、発生過程における細胞型特異的split-GAL4試薬を予測可能にする Using single-cell RNA sequencing to generate predictive cell-type-specific split-GAL4 reagents throughout development
Yu-Chieh David Chen, Yen-Chung Chen, Raghuvanshi Rajesh, Nathalie Shoji, Maisha Jacy, Haluk Lacin, Ted Erclik, and Claude Desplan
Proceedings of the National Academy of Sciences Published:July 31, 2023
DOI:https://doi.org/10.1073/pnas.2307451120
Significance
Understanding the function of individual cell types in the nervous systems has remained a major challenge for neuroscience researchers, partly due to the incomplete identification and characterization of neuronal cell types. To study the development of individual cell types and their function in health and disease, specific experimental access to each cell type is an essential prerequisite. Here, we establish a pipeline to generate gene-specific split-GAL4 lines guided by single-cell RNA sequencing datasets. These intersectional lines show high accuracy for labeling targeted cell types and can be applied to any tissue in Drosophila. The gene-specific split-GAL4 and scMarco, a Graphical User Interface designed to enable identification of marker gene pairs, will represent valuable resources to the fly research community.
Abstract
Cell-type-specific tools facilitate the identification and functional characterization of the distinct cell types that form the complexity of neuronal circuits. A large collection of existing genetic tools in Drosophila relies on enhancer activity to label different subsets of cells and has been extremely useful in analyzing functional circuits in adults. However, these enhancer-based GAL4 lines often do not reflect the expression of nearby gene(s) as they only represent a small portion of the full gene regulatory elements. While genetic intersectional techniques such as the split-GAL4 system further improve cell-type-specificity, it requires significant time and resources to screen through combinations of enhancer expression patterns. Here, we use existing developmental single-cell RNA sequencing (scRNAseq) datasets to select gene pairs for split-GAL4 and provide a highly efficient and predictive pipeline (scMarco) to generate cell-type-specific split-GAL4 lines at any time during development, based on the native gene regulatory elements. These gene-specific split-GAL4 lines can be generated from a large collection of coding intronic MiMIC/CRIMIC lines or by CRISPR knock-in. We use the developing Drosophila visual system as a model to demonstrate the high predictive power of scRNAseq-guided gene-specific split-GAL4 lines in targeting known cell types, annotating clusters in scRNAseq datasets as well as in identifying novel cell types. Lastly, the gene-specific split-GAL4 lines are broadly applicable to any other Drosophila tissue. Our work opens new avenues for generating cell-type-specific tools for the targeted manipulation of distinct cell types throughout development and represents a valuable resource for the Drosophila community.
