Explore the vast range of titles available.

Wnts are evolutionarily conserved secreted signalling proteins that, in various developmental contexts, spread from their site of synthesis to form a gradient and activate target-gene expression at a distance. However, the requirement for Wnts to spread has never been directly tested. Here we used genome engineering to replace the endogenous wingless gene, which encodes the main Drosophila Wnt, with one that expresses a membrane-tethered form of the protein. Surprisingly, the resulting flies were viable and produced normally patterned appendages of nearly the right size, albeit with a delay. We show that, in the prospective wing, prolonged wingless transcription followed by memory of earlier signalling allows persistent expression of relevant target genes. We suggest therefore that the spread of Wingless is dispensable for patterning and growth even though it probably contributes to increasing cell proliferation. Replacement of the wingless ( wg ) gene in Drosophila with one that expresses a membrane-tethered form of Wg results in viable flies with normally patterned appendages of nearly the right size; early wg transcription and memory of signalling ensure continued target-gene expression in the absence of Wg release, even though the spread of Wg could boost cell proliferation. Wingless protein's long-range signalling function Morphogens are secreted proteins with important roles in growth and patterning during development. A key feature of a morphogen is that it can act at a distance from the site of origin through a diffusion gradient. These authors have directly assessed the function of the spread of the Wingless (Wg) protein in Drosophila . They found that flies in which the resident wg gene has been replaced by an engineered form encoding a membrane-tethered Wg protein are viable and fertile. These findings are consistent with a role for Wingless as a long-range signal, but raise doubts over the essential role of a diffusion gradient.

Ribosomes are multicomponent molecular machines that synthesize all of the proteins of living cells. Most of the genes that encode the protein components of ribosomes are therefore essential. A reduction in gene dosage is often viable albeit deleterious and is associated with human syndromes, which are collectively known as ribosomopathies 1 – 3 . The cell biological basis of these pathologies has remained unclear. Here, we model human ribosomopathies in Drosophila and find widespread apoptosis and cellular stress in the resulting animals. This is not caused by insufficient protein synthesis, as reasonably expected. Instead, ribosomal protein deficiency elicits proteotoxic stress, which we suggest is caused by the accumulation of misfolded proteins that overwhelm the protein degradation machinery. We find that dampening the integrated stress response 4 or autophagy increases the harm inflicted by ribosomal protein deficiency, suggesting that these activities could be cytoprotective. Inhibition of TOR activity—which decreases ribosomal protein production, slows down protein synthesis and stimulates autophagy 5 —reduces proteotoxic stress in our ribosomopathy model. Interventions that stimulate autophagy, combined with means of boosting protein quality control, could form the basis of a therapeutic strategy for this class of diseases. Recasens-Alvarez et al. model human ribosomopathies and find that apoptosis and cellular stress result from proteotoxic stress that overwhelms the degradation machinery.

Growth deceleration before growth termination is a universal feature of growth during development. Transcriptomics analysis reveals that during their two-day period of growth deceleration, wing imaginal discs of Drosophila undergo a progressive metabolic shift from oxidative phosphorylation towards glycolysis. Ultra-sensitive reporters of HIF-1α stability and activity show that imaginal discs become increasingly hypoxic during development in normoxic conditions, suggesting that limiting oxygen supply could underlie growth deceleration. We confirm the expectation that rising levels of HIF-1α dampen TOR signalling activity through transcriptional activation of REDD1. Conversely, excess TOR leads, in a tissue-size-dependent manner, to hypoxia, which boosts HIF-1α levels and activity. Thus, HIF-1α mediates a negative feedback loop whereby TOR signalling triggers hypoxia, which in turn reduces TOR signalling. Abrogation of this feedback by Sima/HIF-1α knockdown leads to cellular stress, which is alleviated by reduced TOR signalling or a modest increase in environmental oxygen. We conclude that Sima/HIF-1α prevents TOR-mediated growth from depleting local oxygen supplies during normal development. During normal development, growing fly wing precursors become hypoxic, triggering HIF-1α-mediated negative feedback on TOR signalling and growth deceleration. Without this feedback, the mismatch between oxygen supply and demand leads to tissue stress.

Dpp, a member of the BMP family, is a morphogen that specifies positional information in Drosophila wing precursors. In this tissue, Dpp expressed along the anterior-posterior boundary forms a concentration gradient that controls the expression domains of target genes, which in turn specify the position of wing veins. Dpp also promotes growth in this tissue. The relationship between the spatio-temporal profile of Dpp signalling and growth has been the subject of debate, which has intensified recently with the suggestion that the stripe of Dpp is dispensable for growth. With two independent conditional alleles of dpp, we find that the stripe of Dpp is essential for wing growth. We then show that this requirement, but not patterning, can be fulfilled by uniform, low level, Dpp expression. Thus, the stripe of Dpp ensures that signalling remains above a pro-growth threshold, while at the same time generating a gradient that patterns cell fates. From the wings of a butterfly to the fingers of a human hand, living tissues often have complex and intricate patterns. Developmental biologists have long been fascinated by the signals – called morphogens – that guide how these kinds of pattern develop. Morphogens are substances that are produced by groups of cells and spread to the rest of the tissue to form a gradient. Depending on where they sit along this gradient, cells in the tissue activate different sets of genes, and the resulting pattern of gene activity ultimately defines the position of the different parts of the tissue. Decades worth of studies into how limbs develop in animals from mice to fruit flies have revealed common principles of morphogen gradients that regulate the development of tissue patterns. Morphogens have been shown to help regulate the growth of tissues in a number of different animals as well. However, how the morphogens regulate tissue size and what role their gradients play in this process remain topics of intense debate in the field of developmental biology. In the developing wing of a fruit fly, a morphogen called Dpp is expressed in a thin stripe located in the centre and spreads to the rest of the tissue to form a gradient. Bosch, Ziukaite, Alexandre et al. have now characterised where and when the Dpp morphogen must be produced to regulate both the final size of the fly’s wing and the number of cells the wing eventually contains. The experiments involved preventing the production of Dpp in the developing wing in specific cells and at specific stages of development. This approach confirmed that Dpp must be produced in the central stripe for the wing to grow. Matsuda and Affolter and, independently, Barrio and Milán report the same findings in two related studies. Moreover, Bosch et al. and Barrio and Milán also conclude that the gradient of Dpp throughout the wing is not required for growth. Further work will be needed to explain how the Dpp signal regulates the growth of the wing. The answer to this question will contribute to a better understanding of the role of morphogens in regulating the size of human organs and how a failure to do so might cause developmental disorders.

A genetic multicolor cell-labeling technique for Droshophila melanogaster , Flybow, is described and applied to the study of neural circuits. This method implements a variant of the mouse Brainbow strategy in combination with specific neuronal targeting using the Gal-4–upstream activating sequence system to select for membrane-tethered fluorescent proteins. Also in this issue, Hampel et al . report a similar strategy, Drosophila Brainbow, to select for epitope-tagged proteins detectable via immunofluorescence. To facilitate studies of neural network architecture and formation, we generated three Drosophila melanogaster variants of the mouse Brainbow-2 system, called Flybow. Sequences encoding different membrane-tethered fluorescent proteins were arranged in pairs within cassettes flanked by recombination sites. Flybow combines the Gal4-upstream activating sequence binary system to regulate transgene expression and an inducible modified Flp- FRT system to drive inversions and excisions of cassettes. This provides spatial and temporal control over the stochastic expression of one of two or four reporters within one sample. Using the visual system, the embryonic nervous system and the wing imaginal disc, we show that Flybow in conjunction with specific Gal4 drivers can be used to visualize cell morphology with high resolution. Finally, we demonstrate that this labeling approach is compatible with available Flp- FRT -based techniques, such as mosaic analysis with a repressible cell marker; this could further support the genetic analysis of neural circuit assembly and function.

Tumors adapt their metabolism to sustain increased proliferation, rendering them particularly vulnerable to fluctuations in nutrient availability. However, the role of the tumor microenvironment in modulating sensitivity to nutrient restriction (NR) remains poorly understood. Using a Drosophila brain dedifferentiation neural stem cell (NSC) tumor model induced by Prospero (Pros) inhibition, we show that tumor sensitivity to NR is governed by the blood–brain barrier (BBB) glia. We found that the SLC36 amino acid transporter Pathetic (Path) regulates brain branched-chain amino acids (BCAAs) levels. Under NR, while wild-type buffers against low nutrient levels by upregulating Path, tumor glia down-regulate Path. Furthermore, Path is specifically required by the tumor (but not wildtype) BBB; its downregulation causes reduced cell cycle progression of BBB glial cells and, in turn, restricts NSC tumor growth. Path influences BBB glial cell cycle via the BCAA-mTor-S6K pathway, and its expression is controlled by Ilp6 levels and the Insulin/PI3K pathway. Overexpression of Path is sufficient to counteract the inhibitory effects of NR on tumor growth. These findings suggest that Path levels at the glial niche BBB play a key role in determining tumor sensitivity to NR.

Tumors adapt their metabolism to sustain increased proliferation, rendering them particularly vulnerable to fluctuations in nutrient availability. However, the role of the tumor microenvironment in modulating sensitivity to nutrient restriction (NR) remains poorly understood. Using a Drosophila brain dedifferentiation neural stem cell (NSC) tumor model induced by Prospero (Pros) inhibition, we show that tumor sensitivity to NR is governed by the blood-brain barrier (BBB) glia. We found that the SLC36 amino acid transporter Pathetic (Path) regulates brain branched-chain amino acids (BCAAs) levels. Under NR, while wild-type buffers against low nutrient levels by upregulating Path, tumor glia down-regulate Path. Furthermore, Path is specifically required by the tumor (but not wildtype) BBB; its downregulation causes reduced cell cycle progression of BBB glial cells and, in turn, restricts NSC tumor growth. Path influences BBB glial cell cycle via the BCAA-mTor-S6K pathway, and its expression is controlled by Ilp6 levels and the Insulin/PI3K pathway. Overexpression of Path is sufficient to counteract the inhibitory effects of NR on tumor growth. These findings suggest that Path levels at the glial niche BBB play a key role in determining tumor sensitivity to NR.

Vincent and colleagues show in Drosophila wing imaginal discs that the signalling molecule Wingless is synthesized and secreted at the apical surface, and is re-internalized to be transcytosed basally, where its signalling occurs. The apical and basolateral membranes of epithelia are insulated from each other, preventing the transfer of extracellular proteins from one side to the other 1 . Thus, a signalling protein produced apically is not expected to reach basolateral receptors. Evidence suggests that Wingless, the main Drosophila Wnt, is secreted apically in the embryonic epidermis 2 , 3 . However, in the wing imaginal disc epithelium, Wingless is mostly seen on the basolateral membrane where it spreads from secreting to receiving cells 4 , 5 . Here we examine the apico-basal movement of Wingless in Wingless-producing cells of wing imaginal discs. We find that it is presented first on the apical surface before making its way to the basolateral surface, where it is released and allowed to interact with signalling receptors. We show that Wingless transcytosis involves dynamin-dependent endocytosis from the apical surface. Subsequent trafficking from early apical endosomes to the basolateral surface requires Godzilla, a member of the RNF family of membrane-anchored E3 ubiquitin ligases. Without such transport, Wingless signalling is strongly reduced in this tissue.

Transposable elements have been used in Drosophila to detect gene expression, inactivate gene function, and induce ectopic expression or overexpression. We have combined all of these features in a single construct. A promoterless GAL4 cDNA is expressed when the construct inserts within a transcriptional unit, and GAL4 activates a GFP-encoding gene present in the same transposon. In a primary screen, patterned gene expression is detected as GFP fluorescence in the live progeny of dysgenic males. Many animals expressing GFP in distinct patterns can be recovered with relatively little effort. As expected, many insertions cause loss of function. After insertion at a genomic location, specific parts of the transposon can be excised by FLP recombinase, thus allowing it to induce conditional misexpression of the tagged gene. Therefore, both gain- and loss-of-function studies can be carried out with a single insertion in a gene identified by virtue of its expression pattern. Using this promoter trap approach, we have identified a group of cells that innervate the calyx of the mushroom body and could thus define a previously unrecognized memory circuit.

Morphogens are produced by a subset of cells to trigger a signalling gradient that provides positional information to surrounding tissues. At increasing distances from the source, the dwindling number of morphogen molecules is expected to constrain the useful range of morphogen gradients. We have identified a genetic circuit that counteracts this limitation in developing wings of Drosophila by boosting BMP signalling at the distal end of the gradient without amplifying the signal near the source. This circuit involves Brinker, a transcription factor that represses BMP target genes while itself being repressed by BMP signalling. We suggest that temporal averaging inherent to the production of the inverse Brk gradient contributes to the enhancement of the positional information far from the Dpp source. Despite being a core component of BMP signalling in flies, Brinker is exclusively found in insects, likely in all insect species. Genomic analysis across a wide range of insects and gene expression analysis in limb primordia of the apterygote Thermobia domestica suggests that Brinker is an insect-specific innovation that was subsequently wired into the BMP signalling network in pterygotes, perhaps to enable wing development.Competing Interest StatementThe authors have declared no competing interest.