Generation of a targeted expression system for branchless and characterization of novel cellular expression patterns of the gene in Drosophila
| Autor: | Du, Lijuan, Zhou, Amy, Patel, Akshay, Rao, Mishal, Anderson, Kelsey, Roy, Sougata |
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| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
Microscopy
Confocal Reverse Transcriptase Polymerase Chain Reaction Gene Expression Profiling Gene Expression Regulation Developmental Protein-Tyrosine Kinases Embryo Mammalian Receptors Fibroblast Growth Factor Time-Lapse Imaging Article Animals Genetically Modified Fibroblast Growth Factors Trachea Drosophila melanogaster Organ Culture Techniques Cell Movement Larva Morphogenesis Animals Drosophila Proteins CRISPR-Cas Systems Hypoxia In Situ Hybridization |
| Popis: | Fibroblast growth factors (FGF) are essential signaling proteins that regulate diverse cellular functions in developmental and metabolic processes. In Drosophila, the FGF homolog, branchless (bnl) is expressed in a dynamic and spatiotemporally restricted pattern to induce branching morphogenesis of the trachea, which expresses the Bnl-receptor, breathless (btl). Here we have developed a new strategy to determine bnl- expressing cells and study their interactions with the btl-expressing cells in the range of tissue patterning during Drosophila development. To enable targeted gene expression specifically in the bnl expressing cells, a new LexA based bnl enhancer trap line was generated using CRISPR/Cas9 based genome editing. Analyses of the spatiotemporal expression of the reporter in various embryonic stages, larval or adult tissues and in metabolic hypoxia, confirmed its target specificity and versatility. With this tool, new bnl expressing cells, their unique organization and functional interactions with the btl-expressing cells were uncovered in a larval tracheoblast niche in the leg imaginal discs, in larval photoreceptors of the developing retina, and in the embryonic central nervous system. The targeted expression system also facilitated live imaging of simultaneously labeled Bnl sources and tracheal cells, which revealed a unique morphogenetic movement of the embryonic bnl- source. Migration of bnl- expressing cells may create a dynamic spatiotemporal pattern of the signal source necessary for the directional growth of the tracheal branch. The genetic tool and the comprehensive profile of expression, organization, and activity of various types of bnl-expressing cells described in this study provided us with an important foundation for future research investigating the mechanisms underlying Bnl signaling in tissue morphogenesis. |
| Databáze: | OpenAIRE |
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