Project description:Cell diversity of the brain and how it is affected by starvation, remains largely unknown. Here, we introduce a single cell transcriptome atlas of the entire Drosophila first instar larval brain. We first assigned cell-type identity based on known marker genes, distinguishing five major groups: neural progenitors, differentiated neurons, glia, undifferentiated neurons and non-neural cells. All major classes were further subdivided into multiple subtypes, revealing biological features of various cell-types. We further assessed transcriptional changes in response to starvation at the single-cell level. While after starvation the composition of the brain remains unaffected, transcriptional profile of several cell clusters changed. Intriguingly, different cell-types show very distinct responses to starvation, suggesting the presence of cell-specific programs for nutrition availability. Establishing a single-cell transcriptome atlas of the larval brain provides a powerful tool to explore cell diversity and assess genetic profiles from developmental, functional and behavioral perspectives.
Project description:The cellular composition of the brain and how it is affected by starvation, remains largely unknown. Here we introduce a single-cell transcriptome atlas of the entire Drosophila melanogaster first instar larval brain. We first assigned cell type identity based on the expression of previously characterized marker genes, allowing us to distinguish five major groups: neural progenitors cells, differentiated neurons, glial cells, undifferentiated neurons as well as non-neural cells corresponding to organs and structures located adjacent to the brain. All major classes were further subdivided into multiple subtypes based on cluster analysis, revealing critical biological features of various cell types. Moreover, we included two different feeding conditions: normal fed versus starved. After starvation, the transcriptional profile of several cell clusters were altered, while the overall composition of the brain remains unaffected. Intriguingly, different cell clusters show very distinct responses to starvation, suggesting the presence of cell-specific programs for nutrition availability. Establishing a single-cell transcriptome atlas of the larval brain provides a powerful tool to explore cell diversity, assess genetic profiles of neurogenic, neuronal and glial cell types. The analysis of neurotransmitters, neuropeptides and their respective receptors may further open the doors for functional studies. Overall design: scRNA-seq (10x Genomics ) of the Drosophila larval brain at different feeding conditions
Project description:The diversity of cell types and regulatory states in the brain, and how these change during aging, remains largely unknown. We present a single-cell transcriptome atlas of the entire adult Drosophila melanogaster brain sampled across its lifespan. Cell clustering identified 87 initial cell clusters that are further subclustered and validated by targeted cell-sorting. Our data show high granularity and identify a wide range of cell types. Gene network analyses using SCENIC revealed regulatory heterogeneity linked to energy consumption. During aging, RNA content declines exponentially without affecting neuronal identity in old brains. This single-cell brain atlas covers nearly all cells in the normal brain and provides the tools to study cellular diversity alongside other Drosophila and mammalian single-cell datasets in our unique single-cell analysis platform: SCope (http://scope.aertslab.org). These results, together with SCope, allow comprehensive exploration of all transcriptional states of an entire aging brain.
Project description:Fixed lineages derived from unique, genetically specified neuroblasts form the anatomical building blocks of the Drosophila brain. Neurons belonging to the same lineage project their axons in a common tract, which is labeled by neuronal markers. In this paper, we present a detailed atlas of the lineage-associated tracts forming the brain of the early Drosophila larva, based on the use of global markers (anti-Neuroglian, anti-Neurotactin, inscuteable-Gal4>UAS-chRFP-Tub) and lineage-specific reporters. We describe 68 discrete fiber bundles that contain axons of one lineage or pairs/small sets of adjacent lineages. Bundles enter the neuropil at invariant locations, the lineage tract entry portals. Within the neuropil, these fiber bundles form larger fascicles that can be classified, by their main orientation, into longitudinal, transverse, and vertical (ascending/descending) fascicles. We present 3D digital models of lineage tract entry portals and neuropil fascicles, set into relationship to commonly used, easily recognizable reference structures such as the mushroom body, the antennal lobe, the optic lobe, and the Fasciclin II-positive fiber bundles that connect the brain and ventral nerve cord. Correspondences and differences between early larval tract anatomy and the previously described late larval and adult lineage patterns are highlighted. Our L1 neuro-anatomical atlas of lineages constitutes an essential step towards following morphologically defined lineages to the neuroblasts of the early embryo, which will ultimately make it possible to link the structure and connectivity of a lineage to the expression of genes in the particular neuroblast that gives rise to that lineage. Furthermore, the L1 atlas will be important for a host of ongoing work that attempts to reconstruct neuronal connectivity at the level of resolution of single neurons and their synapses.
Project description:The late larval brain consists of embryonically produced primary neurons forming a deep core cortex, surrounded at the surface by approximately 100 secondary lineages. Each secondary lineage forms a tract (secondary lineage tract) with an invariant and characteristic trajectory. Within the neuropile, tracts of neighboring lineages bundle together to form secondary tract systems. In this paper, we visualized secondary lineages by the global marker BP106 (neurotactin), as well as green fluorescent protein-labeled clones and thereby establish a comprehensive digital atlas of secondary lineages. The information contained in this atlas is the location of the lineage within the cortex, the neuropile compartment contacted by the lineage tract, and the projection pattern of the lineage tract within the neuropile. We have digitally mapped the expression pattern of three genes, sine oculis, period, and engrailed into the lineage atlas. The atlas will enable us and others to analyze the phenotype of mutant clones in the larval brain. Mutant clones can only be interpreted if the corresponding wild-type clone is well characterized, and our lineage atlas, which visualizes all wild-type lineages, will provide this information. Secondly, secondary lineage tracts form a scaffold of connections in the neuropile that foreshadows adult nerve connections. Thus, starting from the larval atlas and proceeding forward through pupal development, one will be able to reconstruct adult brain connectivity at a high level of resolution. Third, the atlas can serve as a repository for genes expressed in lineage-specific patterns.
Project description:The diversity of cell types and regulatory states in the brain, and how these change during ageing, remains largely unknown. We present a single-cell transcriptome atlas of the entire adult Drosophila melanogaster brain sampled across its lifespan. Cell clustering identified 87 initial cell clusters that are further subclustered and validated by targeted cell-sorting. Our data shows high granularity and identifies a wide range of cell types. Gene network analyses using SCENIC revealed regulatory heterogeneity linked to energy consumption. During ageing, RNA content declines exponentially without affecting neuronal identity in old brains. This single-cell brain atlas covers nearly all cells in the normal brain and provides the tools to study cellular diversity alongside other Drosophila and mammalian single-cell datasets in our unique single-cell analysis platform. These results allow comprehensive exploration of all transcriptional states of an entire ageing brain. Overall design: scRNA-seq (10x Genomics CHROMIUM Single Cell 3’ Solution V2 Chemistry and Drop-seq) of adult brains from Drosophila melanogaster at various ages; SMART-seq2 on FAC-sorted R23E10-Gal4 positive neurons of adult brains from Drosophila melanogaster; Adapted SMART-seq2 on FAC-sorted R23E10-Gal4 positive neurons of adult brains from Drosophila melanogaster; CEL-seq2 on FAC-sorted R23E10-Gal4 positive neurons of adult brains from Drosophila melanogaster; Bulk RNA-seq of adult brains from Drosophila melanogaster; ATAC-seq of young (0 days) and old (50 days) adult brains from Drosophila melanogaster
Project description:Imaging the expression patterns of reporter constructs is a powerful tool to dissect the neuronal circuits of perception and behavior in the adult brain of Drosophila, one of the major models for studying brain functions. To date, several Drosophila brain templates and digital atlases have been built to automatically analyze and compare collections of expression pattern images. However, there has been no systematic comparison of performances between alternative atlasing strategies and registration algorithms. Here, we objectively evaluated the performance of different strategies for building adult Drosophila brain templates and atlases. In addition, we used state-of-the-art registration algorithms to generate a new group-wise inter-sex atlas. Our results highlight the benefit of statistical atlases over individual ones and show that the newly proposed inter-sex atlas outperformed existing solutions for automated registration and annotation of expression patterns. Over 3,000 images from the Janelia Farm FlyLight collection were registered using the proposed strategy. These registered expression patterns can be searched and compared with a new version of the BrainBaseWeb system and BrainGazer software. We illustrate the validity of our methodology and brain atlas with registration-based predictions of expression patterns in a subset of clock neurons. The described registration framework should benefit to brain studies in Drosophila and other insect species.
Project description:Our study focuses on understanding the early transcriptional changes taking place during the divergence of the adult muscle precursors that give rise to indirect flight muscles and direct flight muscles in Drosophila. We analyzed the heterogenous cell population of the adult muscle precursors by scRNA-seq and build an integrated single-cell reference atlas. We addressed the differences among muscle-type and different cell state during myoblast differentiation. Also, our dataset includes the transcriptional profile of the epithelial cells localized in the presumptive hinge and notum of third instar larval wing discs. In addition we studied the functional relevance of Amalgam in flight muscle development by depleting Ama expression specifically in the adult muscle precursors. We determined the transcriptional changes and perturbations in AMP cell identity upon Ama knockdown.
Project description:Developmental regulatory proteins are commonly utilized in multiple cell types throughout development. The Drosophila single-minded (sim) gene acts as master regulator of embryonic CNS midline cell development and transcription. However, it is also expressed in the brain during larval development. In this paper, we demonstrate that sim is expressed in three clusters of anterior central brain neurons: DAMv1/2, BAmas1/2, and TRdm and in three clusters of posterior central brain neurons: a subset of DPM neurons, and two previously unidentified clusters, which we term PLSC and PSC. In addition, sim is expressed in the lamina and medulla of the optic lobes. MARCM studies confirm that sim is expressed at high levels in neurons but is low or absent in neuroblasts (NBs) and ganglion mother cell (GMC) precursors. In the anterior brain, sim(+) neurons are detected in 1st and 2nd instar larvae but rapidly increase in number during the 3rd instar stage. To understand the regulation of sim brain transcription, 12 fragments encompassing 5'-flanking, intronic, and 3'-flanking regions were tested for the presence of enhancers that drive brain expression of a reporter gene. Three of these fragments drove expression in sim(+) brain cells, including all sim(+) neuronal clusters in the central brain and optic lobes. One fragment upstream of sim is autoregulatory and is expressed in all sim(+) brain cells. One intronic fragment drives expression in only the PSC and laminar neurons. Another downstream intronic fragment drives expression in all sim(+) brain neurons, except the PSC and lamina. Thus, together these two enhancers drive expression in all sim(+) brain neurons. Sequence analysis of existing sim mutant alleles identified three likely null alleles to utilize in MARCM experiments to examine sim brain function. Mutant clones of DAMv1/2 neurons revealed a consistent axonal fasciculation defect. Thus, unlike the embryonic roles of sim that control CNS midline neuron and glial formation and differentiation, postembryonic sim, instead, controls aspects of axon guidance in the brain. This resembles the roles of vertebrate sim that have an early role in neuronal migration and a later role in axonogenesis.