Conserved roles of engrailed: patterning tissues and specifying cell types.

More than 40 years ago, studies of the Drosophila engrailed and Hox genes led to major discoveries that shaped the history of developmental biology. We learned that these genes define the state of determination of cells that populate particular spatially defined regions: the identity of segmental domains by Hox genes, and the identity of posterior developmental compartments by engrailed. Hence, the boundaries that delimit spatial domains depend on engrailed.

Metabolic control of progenitor cell propagation during Drosophila tracheal remodeling.

Adult progenitor cells in the trachea of Drosophila larvae are activated and migrate out of niches when metamorphosis induces tracheal remodeling. Here we show that in response to metabolic deficiency in decaying tracheal branches, signaling by the insulin pathway controls the progenitor cells by regulating Yorkie (Yki)-dependent proliferation and migration. Yki, a transcription coactivator that is regulated by Hippo signaling, promotes transcriptional activation of cell cycle regulators and components of the extracellular matrix in tracheal progenitor cells.

Fly Cell Atlas: A single-nucleus transcriptomic atlas of the adult fruit fly.

For more than 100 years, the fruit fly has been one of the most studied model organisms. Here, we present a single-cell atlas of the adult fly, Tabula , that includes 580,000 nuclei from 15 individually dissected sexed tissues as well as the entire head and body, annotated to >250 distinct cell types. We provide an in-depth analysis of cell type-related gene signatures and transcription factor markers, as well as sexual dimorphism, across the whole animal.

Protocol for time lapse imaging of cytonemes.

This protocol describes how to image time and spatially resolved time lapses of Drosophila air sac primordium (ASP) cytonemes in cultures of wing imaginal discs. It describes how to manually measure the length of cytonemes using custom-made FIJI/ImageJ tools, and to analyze data using R/R-Studios pipeline. It can also be used for studies of cell division, organelle localization, and protein trafficking as well as other cellular materials that can be fluorescently tagged and imaged with minimal phototoxicity.

Regulated delivery controls Hedgehog, Wingless, and Decapentaplegic signaling.

Morphogen signaling proteins disperse across tissues to activate signal transduction in target cells. We investigated dispersion of Hedgehog (Hh), Wnt homolog Wingless (Wg), and Bone morphogenic protein homolog Decapentaplegic (Dpp) in the wing imaginal disc. We discovered that delivery of Hh, Wg, and Dpp to their respective targets is regulated.

Elimination of nurse cell nuclei that shuttle into oocytes during oogenesis.

Drosophila oocytes develop together with 15 sister germline nurse cells (NCs), which pass products to the oocyte through intercellular bridges. The NCs are completely eliminated during stages 12-14, but we discovered that at stage 10B, two specific NCs fuse with the oocyte and extrude their nuclei through a channel that opens in the anterior face of the oocyte. These nuclei extinguish in the ooplasm, leaving 2 enucleated and 13 nucleated NCs.

The conserved transmembrane protein TMEM-39 coordinates with COPII to promote collagen secretion and regulate ER stress response.

Dysregulation of collagen production and secretion contributes to aging and tissue fibrosis of major organs. How procollagen proteins in the endoplasmic reticulum (ER) route as specialized cargos for secretion remains to be fully elucidated. Here, we report that TMEM39, an ER-localized transmembrane protein, regulates production and secretory cargo trafficking of procollagen.

Hedgehog produced by the wing imaginal disc induces distinct responses in three target tissues.

Hedgehog (Hh) is an evolutionarily conserved signaling protein that has essential roles in animal development and homeostasis. We investigated Hh signaling in the region of the wing imaginal disc that produces Hh and is near the tracheal air sac primordium (ASP) and myoblasts. Hh distributes in concentration gradients in the anterior compartment of the wing disc, ASP and myoblasts, and activates genes in each tissue.

Cytoneme-mediated signaling essential for tumorigenesis.

Communication between neoplastic cells and cells of their microenvironment is critical to cancer progression. To investigate the role of cytoneme-mediated signaling as a mechanism for distributing growth factor signaling proteins between tumor and tumor-associated cells, we analyzed EGFR and RET Drosophila tumor models and tested several genetic loss-of-function conditions that impair cytoneme-mediated signaling. Neuroglian, capricious, Irk2, SCAR, and diaphanous are genes that cytonemes require during normal development.

Designing a Green Fluorogenic Protease Reporter by Flipping a Beta Strand of GFP for Imaging Apoptosis in Animals.

A family of proteases called caspases mediate apoptosis signaling in animals. We report a GFP-based fluorogenic protease reporter, dubbed "FlipGFP", by flipping a beta strand of the GFP. Upon protease activation and cleavage, the beta strand is restored, leading to reconstitution of the GFP and fluorescence.

Glutamate signaling at cytoneme synapses.

We investigated the roles of components of neuronal synapses for development of the air sac primordium (ASP). The ASP, an epithelial tube, extends specialized signaling filopodia called cytonemes that take up signals such as Dpp (Decapentaplegic, a homolog of the vertebrate bone morphogenetic protein) from the wing imaginal disc. Dpp signaling in the ASP was compromised if disc cells lacked Synaptobrevin and Synaptotagmin-1 (which function in vesicle transport at neuronal synapses), the glutamate transporter, and a voltage-gated calcium channel, or if ASP cells lacked Synaptotagmin-4 or the glutamate receptor GluRII.

Visualizing Dynamics of Cell Signaling In Vivo with a Phase Separation-Based Kinase Reporter.

Visualizing dynamics of kinase activity in living animals is essential for mechanistic understanding of cell and developmental biology. We describe GFP-based kinase reporters that phase-separate upon kinase activation via multivalent protein-protein interactions, forming intensively fluorescent droplets. Called SPARK (separation of phases-based activity reporter of kinase), these reporters have large dynamic range (fluorescence change), high brightness, fast kinetics, and are reversible.

Scripting a place in time.

Evidence in many experimental systems supports the idea that non-uniform distributions of morphogen proteins encode positional information in developing tissues. There is also strong evidence that morphogen dispersal is mediated by cytonemes and that morphogen proteins transfer from producing to receiving cells at morphogenetic synapses that form at sites of cytoneme contacts. This essay considers some implications of this mechanism and its relevance to various contexts including large single cells such as the pre-cellular Drosophila embryo and the ciliate Stentor.

Essential basal cytonemes take up Hedgehog in the wing imaginal disc.

Morphogen concentration gradients that extend across developmental fields form by dispersion from source cells. In the wing disc, Hedgehog (Hh) produced by posterior compartment cells distributes in a concentration gradient to adjacent cells of the anterior compartment. We monitored Hh:GFP after pulsed expression, and analyzed the movement and colocalization of Hh, Patched (Ptc) and Smoothened (Smo) proteins tagged with GFP or mCherry and expressed at physiological levels from bacterial artificial chromosome transgenes.

Hyperactive locomotion in a model is a functional readout for the synaptic abnormalities underlying fragile X syndrome.

Fragile X syndrome (FXS) is the most common cause of heritable intellectual disability and autism and affects ~1 in 4000 males and 1 in 8000 females. The discovery of effective treatments for FXS has been hampered by the lack of effective animal models and phenotypic readouts for drug screening. FXS ensues from the epigenetic silencing or loss-of-function mutation of the () gene, which encodes an RNA binding protein that associates with and represses the translation of target mRNAs.

Macrophages Help Cells Connect to Pattern Zebrafish Stripes.

Mechanisms that disseminate the proteins that orchestrate organ and tissue development have been a major focus of cell and developmental biology. Reporting in Science, Eom and Parichy (2017) characterize the role that macrophages play in facilitating long-distance signaling between the cells that make stripes in the adult zebrafish.

Distributing signaling proteins in space and time: the province of cytonemes.

During development, cells use specialized filopodia called cytonemes to deploy the signaling proteins that coordinate growth and direct morphogenesis. Cytonemes are dynamic structures that can extend long distances across tissues to either deliver or take up signaling proteins. Signaling proteins transfer between cells at the tips of cytonemes where specific contacts termed morphogenetic synapses form.

Cells must express components of the planar cell polarity system and extracellular matrix to support cytonemes.

Drosophila dorsal air sac development depends on Decapentaplegic (Dpp) and Fibroblast growth factor (FGF) proteins produced by the wing imaginal disc and transported by cytonemes to the air sac primordium (ASP). Dpp and FGF signaling in the ASP was dependent on components of the planar cell polarity (PCP) system in the disc, and neither Dpp- nor FGF-receiving cytonemes extended over mutant disc cells that lacked them. ASP cytonemes normally navigate through extracellular matrix (ECM) composed of collagen, laminin, Dally and Dally-like (Dlp) proteins that are stratified in layers over the disc cells.