Blockade of the release of the neuropeptide leucokinin to determine its possible functions in fly behavior: Chemoreception assays

Previous studies have revealed leucokinin (LK) expression in the brain and ventral ganglion of Drosophila CNS. One pair of protocerebrum neurons located in the lateral horn area (LHLK) surrounds the peduncles of the mushroom bodies while two pairs of subesophageal neurons (SELKs) project extended processes to the tritocerebrum and through a cervical connection to the ventral ganglion. There, axons of eight or nine pairs of abdominal (ABLK) neurons leave the CNS through the abdominal nerves and processes connecting each other ipsilaterally and contralaterally. The neural functions of LK remain largely unknown, especially those related to Drosophila behavior. Here, we have studied the role of LK in olfactory and gustatory perception by keeping the LK neurons electrically silent through targeted expression of inward rectifier K⁺ channels. In order to examine the effects of LK failure, we first analyzed the dehydration response, comparing the leucokinin-silent individuals with their parents as a control. Our results showed significant differences that demonstrate the effectiveness of the method. We then tested the olfactory behavioral response to a set of odorants over a range of concentrations in a T-maze paradigm in which flies were allowed to choose between the odorant and solvent compartments. The feeding preference assays were carried out on microplates in which flies were allowed to choose between two colored tastes. Our results show that the blockade of LK release alters both olfactory and gustatory responses, and are therefore evidence that this neuropeptide also modulates chemosensory responses through LHLK and SELK neurons.     

Different ways to make neurons: parallel evolution in the SoxB family

Combining genome-wide analyses of binding sites and expression profiles generates a model for the functional evolution of two SOXB paralogous proteins in neurogenesis.     

Development of the Drosophila entero-endocrine lineage and its specification by the Notch signaling pathway

In this paper we have investigated the developmental–genetic steps that shape the entero-endocrine system of Drosophila melanogaster from the embryo to the adult. The process starts in the endoderm of the early embryo where precursors of endocrine cells and enterocytes of the larval midgut, as well as progenitors of the adult midgut, are specified by a Notch signaling-dependent mechanism. In a second step that occurs during the late larval period, enterocytes and endocrine cells of a transient pupal midgut are selected from within the clusters of adult midgut progenitors. As in the embryo, activation of the Notch pathway triggers enterocyte differentiation and inhibits cells from further proliferation or choosing the endocrine fate. The third step of entero-endocrine cell development takes place at a mid-pupal stage. Before this time point, the epithelial layer destined to become the adult midgut is devoid of endocrine cells. However, precursors of the intestinal midgut stem cells (pISCs) are already present. After an initial phase of symmetric divisions which causes an increase in their own population size, pISCs start to spin off cells that become postmitotic and express the endocrine fate marker, Prospero. Activation of Notch in pISCs forces these cells into an enterocyte fate. Loss of Notch function causes an increase in the proliferatory activity of pISCs, as well as a higher ratio of Prospero-positive cells.     

Slit and Robo regulate dendrite branching and elongation of space-filling neurons in Drosophila

Space-filling neurons extensively sample their receptive fields with fine dendritic branches. In this study we show that a member of the conserved Robo receptor family, Robo, and its ligand Slit regulate the dendritic differentiation of space-filling neurons. Loss of Robo or Slit function leads to faster elongating and less branched dendrites of the complex and space-filling class IV multi-dendritic dendrite-arborization (md-da) neurons in the Drosophila embryonic peripheral nervous system, but not of the simpler class I neurons. The total dendrite length of Class IV neurons is not modified in robo or slit mutant embryos. Robo mediates this process cell-autonomously. Upon Robo over-expression in md-da neurons the dendritic tree is simplified and time-lapse analysis during larval stages indicates that this is due to reduction in the number of newly formed branches. We propose that Slit, through Robo, provides an extrinsic signal to coordinate the growth rate and the branching level of space-filling neurons, thus allowing them to appropriately cover their target field.     

The Hox gene Ultrabithorax modulates the shape and size of the third leg of Drosophila by influencing diverse mechanisms

The developmental mechanisms that regulate the relative size and shape of organs have remained obscure despite almost a century of interest in the problem and the fact that changes in relative size represent the dominant mode of evolutionary change. Here, I investigate how the Hox gene Ultrabithorax (Ubx) instructs the legs on the third thoracic segment of Drosophila melanogaster to develop with a different size and shape from the legs on the second thoracic segment. Through loss-of-function and gain-of-function experiments, I demonstrate that different segments of the leg, the femur and the first tarsal segment, and even different regions of the femur, regulate their size in response to Ubx expression through qualitatively different mechanisms. In some regions, Ubx acts autonomously to specify shape and size, whereas in other regions, Ubx influences size through nonautonomous mechanisms. Loss of Ubx autonomously reduces cell size in the T3 femur, but this reduction seems to be partially compensated by an increase in cell numbers, so that it is unclear what effect cell size and number directly have on femur size. Loss of Ubx has both autonomous and nonautonomous effects on cell number in different regions of the basitarsus, but again there is not a strong correlation between cell size or number and organ size. Total organ size appears to be regulated through mechanisms that operate at the level of the entire leg segment (femur or basitarsus) relatively independently of the behavior of individual subpopulations of cells within the segment.     

Morphological differentiation of the embryonic peripheral neurons in Drosophila

Summary The stereotyped segmental and dorso-ventral organization of the peripheral nervous system (PNS) of Drosophila embryos allows the identification of all the neurons in the body wall. Distinct classes of neurons are distinguishable according to their location, the targets they innervate, the particular shape of their dendrites and their cell size. Those neurons innervating external sensory structures (es) and chordotonal organs (ch) have single dendrites and have been previously described (Ghysen et al. 1986; Dambly-Chaudiere and Ghysen 1986; Campos-Ortega and Hartenstein 1985). We describe here the identity and morphological features of three other classes of neurons in the body segments which have multiple dendrites (md neurons): 1) neurons that give rise to elaborate dendritic arborisations (da neurons); 2) neurons that have bipolar dendrites (bd neurons); 3) neurons that arborize around particular tracheal branches (td neurons). The thoracic hemisegment (T2 and T3) contains 13 da, one bd, one td, 21 es and four ch neurons; the abdominal hemisegment (A1 to A7) contains 14 da, three bd, three td, 15 es and eight ch neurons. The arrangement of the segmented peripheral neurons is highly invariant and provides a favorable assay system for the genetic analysis of neurodevelopment.     

Dopamine in Drosophila: setting arousal thresholds in a miniature brain

In mammals, the neurotransmitter dopamine (DA) modulates a variety of behaviours, although DA function is mostly associated with motor control and reward. In insects such as the fruitfly, Drosophila melanogaster, DA also modulates a wide array of behaviours, ranging from sleep and locomotion to courtship and learning. How can a single molecule play so many different roles? Adaptive changes within the DA system, anatomical specificity of action and effects on a variety of behaviours highlight the remarkable versatility of this neurotransmitter. Recent genetic and pharmacological manipulations of DA signalling in Drosophila have launched a surfeit of stories—each arguing for modulation of some aspect of the fly''s waking (and sleeping) life. Although these stories often seem distinct and unrelated, there are some unifying themes underlying DA function and arousal states in this insect model. One of the central roles played by DA may involve perceptual suppression, a necessary component of both sleep and selective attention.     

Drosophila homologue of the Rothmund-Thomson syndrome gene: essential function in DNA replication during development

Members of the RecQ family play critical roles in maintaining genome integrity. Mutations in human RecQL4 cause a rare genetic disorder, Rothmund-Thomson syndrome. Transgenic mice experiments showed that the RecQ4 null mutant causes embryonic lethality. Although biochemical evidence suggests that the Xenopus RecQ4 is required for the initiation of DNA replication in the oocyte extract, its biological functions during development remain to be elucidated. We present here our results in establishing the use of Drosophila as a model system to probe RecQ4 functions. Immunofluorescence experiments monitoring the cellular distribution of RecQ4 demonstrated that RecQ4 expression peaks during S phase, and RecQ4 is expressed only in tissues active in DNA replication, but not in quiescent cells. We have isolated Drosophila RecQ4 hypomorphic mutants, recq^EP and recq4²³, which specifically reduce chorion gene amplification of follicle cells by 4-5 fold, resulting in thin and fragile eggshells, and female sterility. Quantitative analysis on amplification defects over a 14-kb domain in chorion gene cluster suggests that RecQ4 may have a specific function at or near the origin of replication. A null allele recq4¹⁹ causes a failure in cell proliferation, decrease in DNA replication, chromosomal fragmentation, and lethality at the stage of first instar larvae. The mosaic analysis indicates that cell clones with homozygous recq4¹⁹ fail to proliferate. These results indicate that RecQ4 is essential for viability and fertility, and is required for most aspects of DNA replication during development.