Mutations in shaking-B prevent electrical synapse formation in the Drosophila giant fiber system

The giant fiber system (GFS) is a simple network of neurons that mediates visually elicited escape behavior in Drosophila. The giant fiber (GF), the major component of the system, is a large, descending interneuron that relays visual stimuli to the motoneurons that innervate the tergotrochanteral jump muscle (TTM) and dorsal longitudinal flight muscles (DLMs). Mutations in the neural transcript from the shaking-B locus abolish the behavioral response by disrupting transmission at some electrical synapses in the GFS. This study focuses on the role of the gene in the development of the synaptic connections. Using an enhancer-trap line that expresses lacZ in the GFs, we show that the neurons develop during the first 30 hr of metamorphosis. Within the next 15 hr, they begin to form electrical synapses, as indicated by the transfer of intracellularly injected Lucifer yellow. The GFs dye-couple to the TTM motoneuron between 30 and 45 hr of metamorphosis, to the peripherally synapsing interneuron that drives the DLM motoneurons at approximately 48 hr, and to giant commissural interneurons in the brain at approximately 55 hr. Immunocytochemistry with shaking-B peptide antisera demonstrates that the expression of shaking-B protein in the region of GFS synapses coincides temporally with the onset of synaptogenesis; expression persists thereafter. The mutation shak-B2, which eliminates protein expression, prevents the establishment of dye coupling shaking-B, therefore, is essential for the assembly and/or maintenance of functional gap junctions at electrical synapses in the GFS

Reversible regulation of stem cell niche size associated with dietary control of Notch signalling

BACKGROUND: Stem cells can respond to environmental and physiological inputs to adaptively remodel tissues. Little is known about whether stem cell niches are similarly responsive. The Drosophila ovary germline stem cell (GSC) niche is a well-studied model, which is comprised of cap cells that provide anchorage and maintenance signals for GSCs to maintain oogenesis. Previous studies have shown a strong link between diet and the regulation of oogenesis, making this a useful model system in which to investigate dietary regulation of the niche and its associated stem cells. RESULTS: We show that the Drosophila ovary GSC cap cell niche is a dynamic structure, which can contract and expand in fluctuating dietary conditions. Cap cells are lost when adult flies are shifted to nutrient poor diet and are restored after returning flies to nutrient-rich medium. Notch signalling in cap and escort cells is similarly reduced and restored by dietary shifts to nutrient poor and rich media. In old flies decreased Notch signalling is associated with decreased robustness of the niche to dietary changes. We demonstrated using a Notch temperature sensitive allele that removal and restoration of Notch signalling also leads to a reduction and re-expansion of the niche. Changes in niche size were not associated with apoptosis or cell division. We identified two distinct roles for Notch in the adult germarium. Notch can act in cap cells to prevent their loss while activation of Notch in the flanking escort cells results in expansion of the niche. CONCLUSIONS: We provide evidence that dietary changes alone are sufficient to alter Notch signalling and reversibly change niche size in the adult in wild type flies. We show Notch acts in different cells to maintain and re-expand the niche and propose a model in which cell fate transitions between cap cells and flanking somatic cells accounts for niche dynamics. These findings reveal an unexpected reversible plasticity of the GSC niche whose responses provide an integrated read out of the physiological status of the fly that is modulated by diet and age. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12861-015-0059-8) contains supplementary material, which is available to authorized users

Compensatory flux changes within an endocytic trafficking network maintain thermal robustness of Notch signaling.

SummaryDevelopmental signaling is remarkably robust to environmental variation, including temperature. For example, in ectothermic animals such as Drosophila, Notch signaling is maintained within functional limits across a wide temperature range. We combine experimental and computational approaches to show that temperature compensation of Notch signaling is achieved by an unexpected variety of endocytic-dependent routes to Notch activation which, when superimposed on ligand-induced activation, act as a robustness module. Thermal compensation arises through an altered balance of fluxes within competing trafficking routes, coupled with temperature-dependent ubiquitination of Notch. This flexible ensemble of trafficking routes supports Notch signaling at low temperature but can be switched to restrain Notch signaling at high temperature and thus compensates for the inherent temperature sensitivity of ligand-induced activation. The outcome is to extend the physiological range over which normal development can occur. Similar mechanisms may provide thermal robustness for other developmental signals

Down-regulation of Notch target gene expression by Suppressor of deltex

AbstractIn Drosophila, Suppressor of deltex (Su(dx)) mutations display a wing vein gap phenotype resembling that of Notch gain of function alleles. The Su(dx) protein may therefore act as a negative regulator of Notch but its activity on actual Notch signalling levels has not been demonstrated. Here we show that Su(dx) does regulate the level of Notch signalling in vivo, upstream of Notch target genes and in different developmental contexts, including a previously unknown role in leg joint formation. Overexpression of Su(dx) was capable of blocking both the endogenous activity of Notch and the ectopic Notch signalling induced by the overexpression of Deltex, an intracellular Notch binding protein. In addition, using the conditional phenotype of the Su(dx)sp allele, we show that loss of Su(dx) activity is rapidly followed by an up-regulation of E(spl)mβ expression, the immediate target of Notch signal activation during wing vein development. While Su(dx) adult wing vein phenotypes are quite mild, only affecting the distal tips of the veins, we show that the initial consequence of loss of Su(dx) activity is more severe than previously thought. Using a time-course experiment we show that the phenotype is buffered by feedback regulation illustrating how signalling networks can make development robust to perturbation

Mutations in i shaking-B prevent electrical synapse formation in the Drosophila giant fibre system.

The giant fiber system (GFS) is a simple network of neurons that mediates visually elicited escape behavior in Drosophila. The giant fiber (GF), the major component of the system, is a large, descending interneuron that relays visual stimuli to the motoneurons that innervate the tergotrochanteral jump muscle (TTM) and dorsal longitudinal flight muscles (DLMs). Mutations in the neural transcript from the shaking-B locus abolish the behavioral response by disrupting transmission at some electrical synapses in the GFS. This study focuses on the role of the gene in the development of the synaptic connections. Using an enhancer-trap line that expresses lacZ in the GFs, we show that the neurons develop during the first 30 hr of metamorphosis. Within the next 15 hr, they begin to form electrical synapses, as indicated by the transfer of intracellularly injected Lucifer yellow. The GFs dye-couple to the TTM motoneuron between 30 and 45 hr of metamorphosis, to the peripherally synapsing interneuron that drives the DLM motoneurons at approximately 48 hr, and to giant commissural interneurons in the brain at approximately 55 hr. Immunocytochemistry with shaking-B peptide antisera demonstrates that the expression of shaking-B protein in the region of GFS synapses coincides temporally with the onset of synaptogenesis; expression persists thereafter. The mutation shak-B2, which eliminates protein expression, prevents the establishment of dye coupling shaking-B, therefore, is essential for the assembly and/or maintenance of functional gap junctions at electrical synapses in the GFS