Project description:Enteroendocrine cells (EEs) in the intestinal epithelium have important endocrine functions, yet this cell lineage exhibits great local and regional variations that have hampered detailed characterization of EE subtypes. Through single cell RNA-sequencing analysis, combined with a collection of neuropeptide/receptor knock-in strains, here we provide a comprehensive analysis of cellular diversity, spatial distribution and transcription factor (TF) code of EEs in adult Drosophila midgut. We identify 10 major EE subtypes that totally produced 14 different classes of hormone peptides. Each EE on average co-produces 2-5 different classes of hormone peptides. Functional screen with subtype-enriched TFs suggests a combinatorial TF code that controls EE cell diversity, with class-specific TFs Mirr and Ptx1that respectively define two major classes of EEs, and regional TFs such as Esg, Drm, Exex and Fer1that further define regional EE identity. Our single-cell data should greatly facilitate Drosophila modeling of EE differentiation and function.
Project description:Post-developmental organ resizing improves organismal fitness under constantly changing nutrient environments. Although stem cell abundance is a fundamental determinant of adaptive resizing, our understanding of its underlying mechanisms remains primarily limited to the regulation of stem cell division. Here, we demonstrate that nutrient fluctuation induces dedifferentiation in the Drosophila adult midgut to drive adaptive intestinal growth. From lineage tracing and single-cell RNA sequencing, we identify a subpopulation of enteroendocrine (EE) cells that convert into functional intestinal stem cells (ISCs) in response to dietary glucose and amino acids by activating the JAK-STAT pathway. Genetic ablation of EE-derived ISCs severely impairs ISC expansion and midgut growth despite the retention of resident ISCs, and in silico modeling further indicates that EE dedifferentiation enables an efficient increase in the midgut cell number while maintaining epithelial cell composition. Our findings identify a physiologically induced dedifferentiation that ensures ISC expansion during adaptive organ growth in concert with nutrient conditions.
Project description:Methionine restriction is known to extend lifespan in various model organisms including Drosophila melanogaster. In this analysis, we performed scRNAseq of Drosophila female midgut samples to understand the cell type specific response to methionine restriction.
Project description:Homeostatic regulation of the intestinal enteroendocrine lineage hierarchy is a poorly understood process. We resolved transcriptional changes during enteroendocrine differentiation in real-time at single-cell level using a novel knock-in allele of Neurog3, the master regulator gene briefly expressed at the onset of enteroendocrine specification. A bi-fluorescent reporter, Neurog3Chrono, measures time from the onset of enteroendocrine differentiation and enables precise positioning of single-cell transcriptomes along an absolute time axis. This approach yielded a definitive description of the enteroendocrine hierarchy and its sub-lineages, uncovered differential kinetics between sub-lineages, and revealed time-dependent hormonal plasticity in enterochromaffin and L-cells. The time-resolved map of transcriptional changes predicted multiple novel molecular regulators. Nine of these were validated by conditional knockout in mice or CRISPR-modification in intestinal organoids. Six novel candidate regulators (Sox4, Rfx6, Tox3, Myt1, Runx1t1 and Zcchc12) yielded specific enteroendocrine phenotypes. Our time-resolved single cell transcriptional map presents a rich resource to unravel enteroendocrine differentiation.
Project description:Enteroendocrine cells (EECs) alter hormone expression while migrating along the crypt-villus axis. We report this swich is regulated by a BMP morphogen gradient. Single cell RNA sequencing of control and BMP4 stimulated EECs was performed using organoids from different hormone reporter mice. We show that BMP4 stimulation can switch different subsets of EECs from their crypt to villus state.
Project description:Comparisons between EcR mutant midgut and a reference control sample from mixed-stage normal midgut Keywords = Drosophila, ecdysone, network, genomic, microarray, organogenesis, EcR, midgut, central nervous system, salivary gland, epidermis, imaginal disc, development Keywords: other
Project description:A midgut reference sample is compared to midgut samples taken at various stages before (18h, 4h), at (0h), and after (2h, 3h, 4h, 5h, 6h, 8h, 10h, 12h) puparium formation. Keywords = Drosophila, ecdysone, network, genomic, microarray, organogenesis, EcR, midgut, central nervous system, salivary gland, epidermis, imaginal disc, development Keywords: other
Project description:We generated pluripotent stem cell-derived human intestinal organoids with (n=2) and without (n=1) EECs and performed single-cell sequencing after their transplantation under the mouse kidney capsule to evaluate similarities and differences in the absence of enteroendocrine cells.
Project description:Bulk RNAseq from whole adult Drosophila melanogaster guts expressing either mCherry-RNAi (control) or RNAi against ChAT (Choline acteyltransferase) under control of a enteroendocrine-specific driver to assess the role of epithelial acetylcholine production in intestinal barrier function.