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Ecdysteroid control of cell cycle and cellular commitment in insect wing imaginal discs.

The wing imaginal disc shows two different developmental responses to the steroid hormone 20-hydroxyecdysone (20E) in conjunction with cell cycle, and the responses depend on the developmental stage and the concentration to which it is exposed. The time of head capsule slippage (HCS) in late fourth instar, which signals the onset of molting process into the fifth instar, is the boundary for the appearance of 20E-dependency. Before HCS, 20E was not effective in maintaining the cell cycle, while, after HCS, 20E drove the cell cycle (20E-dependent cell cycle). In the 20E-dependent cell cycle, S phase was induced by 20E above a threshold concentration, whereas M phase was induced in a range of concentrations, above which it was suppressed. The 20E-dependent cell cycle first appeared concomitantly with the entrance of the discs into the reversible stage of pupal commitment. The composite data suggest that the control mechanism of cell cycle is altered in the pupally committed cells so that a high ecdysteroid titer to induce a stationary molt in young instars does not affect the cell division of uncommitted imaginal disc cells while that to induce pupal ecdysis suppresses it in the committed discs, which occurs before the onset of pupal differentiation.

Animals↗

The tumor suppressor gene fat modulates the EGFR-mediated proliferation control in the imaginal tissues of Drosophila melanogaster.

Molecules involved in cell adhesion can regulate both early signal transduction events, triggered by soluble factors, and downstream events involved in cell cycle progression. Correct integration of these signals allows appropriate cellular growth, differentiation and ultimately tissue morphogenesis, but incorrect interpretation contributes to pathologies such as tumor growth. The Fat cadherin is a tumor suppressor protein required in Drosophila for epithelial morphogenesis, proliferation control and epithelial planar polarization, and its loss results in a hyperplastic growth of imaginal tissues. While several molecular events have been characterized through which fat participates in the establishment of the epithelial planar polarity, little is known about mechanisms underlying fat-mediated control of cell proliferation. Here we provide evidence that fat specifically cooperates with the epidermal growth factor receptor (EGFR) pathway in controlling cell proliferation in developing imaginal epithelia. Hyperplastic larval and adult fat structures indeed undergo an amazing, synergistic enlargement following to EGFR oversignalling. We further show that such a strong functional interaction occurs downstream of MAPK activation through the transcriptional regulation of genes involved in the EGFR nuclear signalling. Considering that fat mutation shows di per se a hyperplastic phenotype, we suggest a model in which fat acts in parallel to EGFR pathway in transducing different cell communication signals; furthermore its function is requested downstream of MAPK for a correct rendering of the growth signals converging to the epidermal growth factor receptor.

Animals↗

Independent roles of Drosophila Moesin in imaginal disc morphogenesis and hedgehog signalling.

The three ERM proteins (Ezrin, Radixin and Moesin) form a conserved family required in many developmental processes involving regulation of the cytoskeleton. In general, the molecular function of ERM proteins is to link specific membrane proteins to the actin cytoskeleton. In Drosophila, loss of moesin (moe) activity causes incorrect localisation of maternal determinants during oogenesis, failures in rhabdomere differentiation in the eye and alterations of epithelial integrity in the wing imaginal disc. Some aspects of Drosophila Moe are related to the activity of the small GTPase RhoA, because the reduction of RhoA activity corrects many phenotypes of moe mutant embryos and imaginal discs. We have analysed the phenotype of moesin loss-of-function alleles in the wing disc and adult wing, and studied the effects of reduced Moesin activity on signalling mediated by the Notch, Decapentaplegic, Wingless and Hedgehog pathways. We found that reductions in Moesin levels in the wing disc cause the formation of wing-tissue vesicles and large thickenings of the vein L3, corresponding to breakdowns of epithelial continuity in the wing base and modifications of Hedgehog signalling in the wing blade, respectively. We did not observe any effect on signalling pathways other than Hedgehog, indicating that the moe defects in epithelial integrity have not generalised effects on cell signalling. The effects of moe mutants on Hedgehog signalling depend on the correct gene-dose of rhoA, suggesting that the requirements for Moesin in disc morphogenesis and Hh signalling in the wing disc are mediated by its regulation of RhoA activity. The mechanism linking Moesin activity with RhoA function and Hedgehog signalling remains to be elucidated.

Animals↗

JNK signaling pathway required for wound healing in regenerating Drosophila wing imaginal discs.

We have examined wound healing during regeneration of Drosophila wing imaginal discs fragments by confocal microscopy and assessed the role of components of the JNK pathway in this process. After cutting, columnar and peripodial epithelia cells at the wound edge start to close the wound through formation and contraction of an actin cable. This is followed by a zipping process through filopodial protrusions from both epithelia knitting the wound edges from proximal to distal areas of the disc. Activation of the JNK pathway is involved in such process. puckered (puc) expression is induced in several rows of cells at the edge of the wound, whereas absence of JNK pathway activity brought about by hemipterous, basket, and Dfos mutants impair wound healing. These defects are accompanied by lowered or loss of expression of puc. In support of a role of puc in wound healing, hep mutant phenotypes are rescued by reducing puc function, whereas overexpression of puc inhibits wound healing. Altogether, these results demonstrate a role for the JNK pathway in imaginal disc wound healing, similar to that reported for other healing processes such as embryonic dorsal closure, thoracic closure, and adult epithelial wound healing in Drosophila. Differences with such processes are also highlighted.

Animals↗

Modulation of the Suppressor of fused protein regulates the Hedgehog signaling pathway in Drosophila embryo and imaginal discs.

The Suppressor of fused (Su(fu)) protein is known to be a negative regulator of Hedgehog (Hh) signal transduction in Drosophila imaginal discs and embryonic development. It is antagonized by the kinase Fused (Fu) since Su(fu) null mutations fully suppress the lack of Fu kinase activity. In this study, we overexpressed the Su(fu) gene in imaginal discs and observed opposing effects depending on the position of the cells, namely a repression of Hh target genes in cells receiving Hh and their ectopic expression in cells not receiving Hh. These effects were all enhanced in a fu mutant context and were suppressed by cubitus interruptus (Ci) overexpression. We also show that the Su(fu) protein is poly-phosphorylated during embryonic development and these phosphorylation events are altered in fu mutants. This study thus reveals an unexpected role for Su(fu) as an activator of Hh target gene expression in absence of Hh signal. Both negative and positive roles of Su(fu) are antagonized by Fused. Based on these results, we propose a model in which Su(fu) protein levels and isoforms are crucial for the modulation of the different Ci states that control Hh target gene expression.

Animals↗

Reproductive and worker castes in the primitively eusocial wasp Belonogaster petiolata (DeGeer) (Hymenoptera: Vespidae): evidence for pre-imaginal differentiation.

Until recently, morphological differences between castes in independent-founding polistine wasps were considered absent. This paper investigates the extent of morphological and physiological differences between reproductive (foundress and gyne) and worker castes of Belonogaster petiolata, and tests the hypothesis that caste differentiation in this species occurs pre-imaginally.Foundresses were significantly larger than workers, to the extent that foundress/worker ratios were comparable with those between queens and workers in some swarm-founding Polistinae. Early emerging workers were small, but body size increased over the colony cycle such that late-season workers were similar in size to gynes. In proportion to body size, workers possessed broader heads while foundresses and gynes had broader thoraces and gasters. All queens, 98% of subordinate foundresses, and 95% of over-wintering gynes were inseminated. Workers were never inseminated and lacked mature ovaries in colonies with active queens. However, in the absence of the queen (and other foundresses), 11% of workers developed mature ovaries. Ovarian size and fat content of foundresses and gynes was significantly greater than that of workers. The differences in external morphology and reproductive physiology between castes support the hypothesis that differentiation occurs pre-imaginally. However, imaginal factors, in particular social dominance of the queen, maintain the reproductive subordinance of workers.

Journal Article↗

Within grasp but out of reach: evidence for a double dissociation between imagined hand and arm movements in the left cerebral hemisphere.

What roles are played by the cerebral hemispheres in planning object-oriented reaching and grasping movements? In an attempt to address this question, we compared the abilities of the left and right hemispheres of commissurotomy patient J.W. to imagine hand manipulation (i.e., grasp) or arm transportation (i.e., reach) movements. A graphically rendered manipulandum (dowel) was briefly presented to the left (LVF) or right (RVF) visual fields in a variety of different orientations. In the grasp selection task (experiment 1), J.W. was required to determine which side of a dowel his thumb would be on if he were to engage the stimulus in a power grip using either his dominant (right) or non-dominant hand. In the reach selection task (experiment 3), J.W. judged which end his elbow would be on if he treated the dowel as an armrest for his dominant or non-dominant forearm. No actual movements were allowed in either task. Movements selected in the imagery tasks were compared with those chosen during actual motor control under comparable circumstances. These comparisons revealed a left hemisphere advantage for representing grasping movements involving the right hand, and reaching movements involving the left arm. The right hemisphere, by contrast, displayed moderate accuracy when representing grasping movements with the left hand, but appeared incapable of imagining reaching movements with either arm. The double dissociation between imagery for hand and arm movements in the left cerebral hemispere is consistent with the hypothesis that grasping and reaching components of prehension involve dissociable planning mechanisms.

Arm↗

A sequence conserved in vertebrate Hox gene introns functions as an enhancer regulated by posterior homeotic genes in Drosophila imaginal discs.

The intron of the mouse Hoxa-4 gene acts as a strong homeotic response element in Drosophila melanogaster leg imaginal discs. This activity depends on homeodomain binding sites present within a 30 bp conserved element, HB1, in the intron. A similar arrangement of homeodomain binding sites is found in many other potential homeotic target genes. HB1 activity in Drosophila imaginal discs is activated by Antennapedia and more posterior homeotic genes, but is not activated by more anterior genes. Testing a reporter gene construct with mutated binding sites in mouse embryos shows that HB1 is also active in the expression domains of posterior Hox genes in the mouse neural tube.

Animals↗

The Drosophila homeotic gene moira regulates expression of engrailed and HOM genes in imaginal tissues.

moira is a member of the trithorax group of homeotic gene regulators in Drosophila melanogaster. We show that moira is required for the function of multiple homeotic genes of the Antennapedia and bithorax complexes (HOM genes) in most imaginal tissues and that the requirement for moira function is at the level of transcription. moira is also required for transcription of the engrailed segmentation gene in the imaginal wing disc. The abnormalities caused by the loss of moira function in germ cells suggests that at least one other target gene requires moira for normal oogenesis.

Animals↗

Effect of bisacylhydrazine ecdysteroid mimics (RH-5849 and RH-5992) on chromosomal puffing, imaginal disc proliferation and pupariation in larvae of Drosophila melanogaster.

Two bisacylhydrazine insecticides with ecdysone-mimetic action, RH-5849 and RH-5992, have been subjected to several bioassay procedures that are prerequisites for ecdysone action in Drosophila larvae: (a) induction of early ecdysone-specific puffs on the polytene chromosomes of the larval salivary glands; (b) secretion of glycoprotein glue into the lumen of the salivary glands; (c) evagination of imaginal discs of adult wings and legs; and (d) partial rescue of wild-type phenotypic expression in ecdysone-deficient mutants (ecdysoneless1 (ecd1) and suppressor of forkedts67g (su(f)ts67g). In all these bioassays on Drosophila larvae, the two purely synthetic hydrazines exhibited similar dose-response relationships as did the natural steroid hormone, 20-hydroxyecdysone. In assays involving induction of early chromosomal puffs (74EF, 75B) or regression of the preexisting puffs (25AC, 68C), the dosages required for induction of standard ED-50 effects were one order of magnitude larger for the hydrazines in comparison with 20-hydroxyecdysone. In the assays related to glycoprotein glue secretion, evagination of imaginal discs, or rescue of phenotypic expression in ecdysone-deficient mutants, 20-hydroxyecdysone was two orders of magnitude more active than RH-5849 and RH-5992. We conclude that, in spite of these quantitative differences, the two hydrazine compounds studied are able to duplicate in Drosophila larvae the complex of qualitative biological effects that are a prerequisite for ecdysteroid hormones. The hormonomimetic stimulus of RH-compounds has been given at very low, intracellular, chromosomal level.

Animals↗

Hedgehog signal transduction in the posterior compartment of the Drosophila wing imaginal disc.

Drosophila Hedgehog (Hh) is secreted by Posterior (P) compartment cells and induces Anterior (A) cells to create a developmental organizer at the AP compartment border. Hh signaling converts Fused (Fu) to a hyperphosphorylated form, Fu*. We show that A border cells of wing imaginal discs contain Fu*. Unexpectedly, P cells also produce Fu*, in a Hh-dependent and Ptc-independent manner. Increasing Ptc, the putative Hh receptor expressed specifically by A cells, reduced Fu*. These results are consistent with proposals that Ptc downregulates Hh signaling and suggest that a receptor other than Ptc mediates Hh signaling in P cells of imaginal discs. We conclude that Hh signals in these P cells and that the outputs of the pathway are blocked by transcriptional repression.

Animals↗

Lumenal transmission of decapentaplegic in Drosophila imaginal discs.

Drosophila imaginal discs are sac-like appendage primordia comprising apposed peripodial and columnar cell layers. Cell survival in disc columnar epithelia requires the secreted signal Decapentaplegic (DPP), which also acts as a gradient morphogen during pattern formation. The distribution mechanism by which secreted DPP mediates global cell survival and graded patterning is poorly understood. Here we report detection of DPP in the lumenal cavity between apposed peripodial and columnar cell layers of both wing and eye discs. We show that peripodial cell survival hinges upon DPP signal reception and implicate DPP-dependent viability of the peripodial epithelium in growth of the entire disc. These results are consistent with lumenal transmission of the DPP survival signal during imaginal disc development.

Animals↗

Induction of smoking urge through imaginal and in vivo procedures: physiological and self-report manifestations.

This experiment assessed reactivity to imaginal and in vivo smoking and control cues. One hundred current smokers were assessed during 2 sessions separated by a 6-hr interval, and half of the participants were abstinent from smoking over this interval. Verbal and physiological reactivity measures were selected based on their relevance for several models of urge. Results indicated that imaginal and in vivo smoking cues were equally effective at eliciting high levels of self-reported urges. Smoking deprivation led to a general enhancement in urge report, rather than a specific increase to smoking cues. Physiological responding differed somewhat as a function of urge induction method, although autonomic responses to smoking cues were uniformly consistent with the direct effects of nicotine. There was no relationship between verbal and physiological urge indices. Implications of the findings for several contemporary models of drug urges are discussed.

Adult↗

Dorsoventral lineage restriction in wing imaginal discs requires Notch.

The formation of boundaries that prevent the intermixing of cells is an important developmental patterning mechanism. The compartmental lineage restrictions that appear in the developing imaginal discs of Drosophila are striking examples of such boundaries. However, little is known about the cellular mechanism underlying compartmental lineage restrictions. The dorsoventral (D/V) lineage restriction that arises late in the developing wing imaginal disc requires the dorsal expression of the transcription factor Apterous and it has been hypothesized that apterous (ap) maintains compartmentalization by directly regulating the expression of molecules that modify cell adhesion or affinity. However, ap expression also regulates signalling between dorsal and ventral compartments, resulting in high levels of Notch signalling at the D/V boundary. Here we show that the formation of Notch-dependent boundary cells is required for the D/V lineage restriction.

Animals↗

Over-expression of DREF in the Drosophila wing imaginal disc induces apoptosis and a notching wing phenotype.

BACKGROUND: DNA replication-related element binding factor (DREF) has been suggested to be involved in regulation of DNA replication- and proliferation-related genes in Drosophila. While the effects on the mutation in the DNA replication-related element (DRE) in cultured cells have been studied extensively, the consequences of elevating wild-type DREF activity in developing tissues have hitherto remained unclear. RESULTS: We over-expressed DREF in the wing imaginal disc using a GAL4-UAS targeted expression system in Drosophila. Over-expression of DREF induced a notching wing phenotype, which was associated with ectopic apoptosis. A half reduction of the reaper, head involution defective and grim gene dose suppressed this DREF-induced notching wing phenotype. Furthermore, this was also the case with co-expression of baculovirus P35, a caspase inhibitor. In addition, over-expression of the 32 kDa boundary element-associated factor (BEAF-32), thought to compete against DREF for common binding sites in genomic regions, rescued the DREF-induced notching wing phenotype, while a half reduction of the genomic region, including the BEAF-32 gene, exerted enhancing effects. To our knowledge, this is the first evidence for a genetic interaction between DREF and BEAF-32. CONCLUSION: The DREF-induced notching wing phenotype is caused by induction of apoptosis in the Drosophila wing imaginal disc.

Animals↗

Polypeptides differentially expressed in imaginal discs define the peroxiredoxin family of genes in Drosophila.

2D gel electrophoresis followed by microsequencing has been used to purify and identify a protein (catalogued in the database as SSP5111) from Drosophila wing imaginal discs of third instar larvae that showed significant differences in their level of expression when compared with other imaginal discs of the same age. The microsequence data showed identity with amino acids encoded by the human proliferation association gene, pag, which is a thiol-specific antioxidant. By virtue of this homology we have cloned and sequenced two cDNAs that appear to define the peroxiredoxin family of Drosophila. One of them, Jafrac1, encodes the SSP5111 protein searched, had 194 amino acids and mapped in the region 11E in the X chromosome. The other, Jafrac2, encodes a protein of 242 amino acids and mapped in the region 62F in the 3 L chromosome. Both new peroxidases contain two conserved cysteines and share homology with other peroxidases that extends over the entire sequence and ranges between 47% and 76%. An antiserum raised against the SSP5111 protein showed significant changes in the amount of protein in different stages of Drosophila development, being a major product in early embryos. In 2D gels the antibody not only recognizes the SSP5111 polypeptide but also a related one (catalogued in the database as SSP6107) that exhibits identical amino-acid sequence over at least 85% of its sequence. The data also suggest that the SSP5111 polypeptide could be a maternal-effect product.

Alternative Splicing↗

Establishment of a cell line from lepidopteran wing imaginal discs: Induction of newly synthesized proteins by 20-hydroxyecdysone.

A cell line has been developed from the imaginal wing discs of the lepidopteran insect Trichoplusia ni (Hübner). The cells grow in suspension as fluid-filled, multicellular vesicles. The cell line has been designated IAL-TND1. Two-dimensional electrophoresis (isoelectric focusing and sodium dodecyl sulfate/polyacrylamide gels) of [(35)S]methionine-labeled proteins revealed similar patterns for IAL-TND1 cells and whole T. ni wing imaginal discs. Additionally, exposure to the insect steroid hormone 20-hydroxyecdysone stimulated the synthesis of the same new proteins in both whole wing discs and IAL-TND1 cell vesicles.

Journal Article↗

Dynamic posturography in humans imagining a fixed spatial reference.

OBJECTIVE: To investigate the effect of imagining a fixed spatial reference on balance control. MATERIAL AND METHODS: Twenty-one healthy subjects were asked to remain as stable as possible while standing on a sway-referenced platform (NeuroCom Equitest posturography system). Subjects were instructed to keep their eyes open in the dark and to either look far into the distance without fixating on any particular point in space or to direct their gaze towards a remembered earth-fixed target (25-cm distant). Room lights were switched off immediately before each 20-s sway recording. Postural stability measures included equilibrium scores and root-mean-square sway amplitudes in the anterior-posterior plane. RESULTS: Postural sway was not influenced by the instruction to gaze towards a remembered target. CONCLUSION: Imagining a fixed spatial reference does not influence postural control, at least during sway-referenced dynamic posturography.

Adolescent↗