The defective dorsal discs gene of Drosophila is required for the growth of specific imaginal discs

The wild-type allele of the gene defective dorsal discs (ddd) is required for the normal development of the dorsal thoracic discs in Drosophila melanogaster. In ddd mutant larvae the dorsal discs (wing, haltere, and humeral) are greatly reduced in size or absent while the ventral discs (leg) are unaffected. We have examined the function of the ddd+ gene in wing development. The ddd+ product is not involved in the initial determination of wing cells but rather is required for their subsequent proliferation during the larval period. Analysis of chimaeras shows that there is a requirement for ddd+ gene expression in wing discs, but it is sufficient for normal development that only some cells in a disc express the gene. We propose that the ddd+ product is involved in the synthesis of a factor which is required for the normal growth of wing discs and which can be transferred between wing disc cells.     

Peripodial cells regulate proliferation and patterning of Drosophila imaginal discs

Cells employ a diverse array of signaling mechanisms to establish spatial patterns during development. Nowhere is this better understood than in Drosophila, where the limbs and eyes arise from discrete epithelial sacs called imaginal discs. Molecular-genetic analyses of pattern formation have generally treated discs as single epithelial sheets. Anatomically, however, discs comprise a columnar cell monolayer covered by a squamous epithelium known as the peripodial membrane. Here we demonstrate that during development, peripodial cells signal to disc columnar cells via microtubule-based apical extensions. Ablation and targeted gene misexpression experiments demonstrate that peripodial cell signaling contributes to growth control and pattern formation in the eye and wing primordia. These findings challenge the traditional view of discs as monolayers and provide foundational evidence for peripodial cell function in Drosophila appendage development.     

Continuity of the membrane systems in the cells of imaginal discs

Summary Imaginal discs from mature larvae of the fly Calliphora erythrocephala fixed in permanganate and embedded in Epon 812 were observed by electron microscopy. Irrespective of the type of the disc, the endoplasmic reticulum in the cells was found to be in unbroken contact with the outer nuclear membrane, Golgi areas, the mitochondria and the cell membrane. The imaginai discs are syncytial in that all cells are in cytoplasmic communication through secondary breaks in the cell membranes.The investigation was facilitated by grants from the Nordic Insulin Foundation.I am indebted to Mrs. Mariann Carleson and Miss Brita Nilsson for technical aid.     

Metamorphosis of imaginal discs of Drosophila melanogaster

Summary Imaginal discs ofDrosophila melanogaster larvae, 24–53 hrs after oviposition, were transplanted into mature immobile larval hosts. The transplants did not respond to the hormonal stimuli of metamorphosis, but instead completed their larval development. When reinjected into mature larval hosts, they now differentiated the full set of their presumptive imaginal structures. The process of acquiring competence for metamorphosis appears to be independent of the hormonal conditions.Supported by a credit of the Swiss National Foundation granted to Prof. Dr. E. Hadorn. I thank Dr. R. Nöthiger for his valuable criticism during this investigation.     

A screen for genes expressed in Drosophila imaginal discs

The development of Drosophila imaginal discs serves as a model system to understand how genes determine the shape and size of an organ. The identification of genes involved in this process is an important step towards this goal. Here we describe a P-element based enhancer trap screen for genes expressed in the larval imaginal discs. Our aim was to establish a large collection of enhancer trap lines each showing expression of Gal4 in imaginal discs. To this end, we improved the well established P-element vector pGawB in order to obtain higher in vivo transposition frequencies. In addition we chose an F1-screening approach using UAS-GFP as a reporter gene. This system permits the efficient screening of larval and pupal stages of living animals and the detection of imaginal gene expression patterns through the transparent cuticle. The procedure has been optimized for high-throughput. 2'000 P-element insertions have been established which exhibit expression in imaginal discs.     

Origin and proliferation of blastema cells during regeneration of Drosophila wing imaginal discs

Following a period of neglect, there has been a resurgence of interest in Drosophila imaginal discs as a model with which to analyze the relationships between growth and pattern formation during regeneration. To broaden our understanding of this process, we used cell lineage techniques to trace the origin of blastema cells and the early and late boundaries of the blastema in regenerating 3/4 wing disc fragments, examined the distribution of S-phase, mitotic and dead cells, and undertook clonal analysis to determine the topology of cell proliferation and its relationship to pattern formation. Using lineage tagging with the JNK phosphatase puckered (puc), we demonstrate that a substantial number of blastema cells arise from cells in which JNK is activated. Furthermore, we show that DNA synthesis and mitosis are activated well before wound healing is completed, in a region where the JNK pathway is activated; later, DNA synthesis and mitosis are observed in scattered cells throughout the blastema. Finally, clonal analysis shows a close relationship between the size and shape of clones and disparities in the positional values of the apposed surfaces.     

Extracellular peptidases of imaginal discs of Drosophila melanogaster

The imaginal discs of Drosophila melanogaster give rise to the adult epidermis during metamorphosis. During this developmental period several peptidase genes are expressed in disc cells, but there is a paucity of biochemical information regarding substrate specificity. We have used peptides and peptidyl 7-amino-4-methylcoumarin (AMC) substrates to detect several peptidases either positioned on the surface of wing discs or secreted by the imaginal cells. Using [Leu(5)]enkephalin as a substrate, a captopril sensitive dipeptidyl carboxypeptidase (angiotensin I-converting enzyme) and an amastatin-sensitive aminopeptidase were detected as prominent activities associated with intact discs. The formation of [Leu(5)]enkephalin-derived Phe was attributed to the concerted action of the D. melanogaster angiotensin I-converting enzyme (Ance) and a dipeptidase. The disc Ance also showed endopeptidic activity towards locust tachykinin-1 (LomTK-I) by cleaving the Gly-Val peptide bond, but this enzyme was not the sole endopeptidase activity associated with discs. Complete inhibition of the endopeptidic hydrolysis of the LomTK-1 by a disc homogenate required a combination of captopril and the neprilysin inhibitor, phosphoramidon, providing biochemical evidence for a neprilysin-like peptidase, in addition to Ance, in imaginal discs of D. melanogaster. Peptidyl AMC substrates for furin, prohormone convertase and tryptase provided evidence for trypsin-like serine endopeptidases in addition to the metalloendopeptidases. We conclude that imaginal discs are endowed with a variety of peptidases from different families that together are capable of hydrolyzing a broad range of peptides and proteins. Some of these peptidases might be responsible for the metabolic activation/inactivation of signaling peptides, as well as being involved in the production of dipeptides and free amino acids required for protein synthesis and osmotic balance during adult morphogenesis.     

Cell cycle changes during growth and differentiation of imaginal leg discs in Drosophila melanogaster

The relative DNA content of Drosophila melanogaster imaginal leg disc nuclei during larval growth and pupal and adult differentiation was measured by microspectrophotometry. During the larval proliferative phase there were twice as many nuclei in the 4C class as nuclei in the 2C class. At the end of the third larval instar, the proportion of nuclei with a 4C DNA value increased. By 3 hr after pupariation, during pupal cuticle secretion, 90% of the nuclei were in this class. After pupal apolysis which occurs at 12 hr after pupariation, the 4C to 2C ratio was reversed. The increase in the proportion of nuclei with a 2C value was observed until 24 hr after pupariation when 90% of the nuclei were in this class. We propose that most cells divide at least once between pupal and adult differentiation. All of these changes in the cell cycle were correlated temporally with changes in the ecdysteroid titers that occur during these periods.     

Parameters of growth in primary cultures and cell lines established from Drosophila imaginal discs

We have further characterised our tissue culture system for the growth in vitro of Drosophila imaginal disc cells, including the culture medium requirements for optimum growth and we have adjusted the protocol recommended for the initiation of cultures. Many imaginal disc fragments become organised into vesicles, and some of these secrete extracellular material into the lumen. Sensory axons differentiate in primary disc cultures, in the absence of bristle formation. The early stages of cell division to form a cell line are recorded.     

Both genes for EF-Tu in Salmonella typhimurium are individually dispensable for growth

Each of the two genes encoding EF-Tu in Salmonella typhimurium has been inactivated using a mini-Mu MudJ insertion. Eleven independently isolated insertions are described, six in tufA and five in tufB. Transduction analysis shows that the inserted MudJ is 100% linked to the appropriate tuf gene. A mutant strain with electrophoretically distinguishable EF-TuA and EF-TuB was used to show, on two-dimensional gels, that the MudJ insertions result in the loss of the appropriate EF-Tu protein. Southern blotting, using cloned Escherichia coli tuf sequences as probes, shows that each MudJ insertion results in the physical breakage of the appropriate tuf gene. The degree of growth-rate impairment associated with each tuf inactivation is independent of which tuf gene is inactivated. The viability of S. typhimurium strains with either tuf gene inactive contrasts strongly with data suggesting that in the closely related bacterium E. coli, an active tufA gene is essential for growth. Finally the strains described here facilitate the analysis of phenotypes associated with individual mutant or wild-type Tus both in vivo and in vitro.     

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.     

The embryonic origin of imaginal discs in Drosophila

The thoracic imaginal discs of Drosophila melanogaster can be observed during embryogenesis as clusters of cells with particular shapes, sizes and behaviours. These structures can be detected soon after germ band shortening and their development appears to be tightly linked to that of the larval epidermis.     

What are and what are not imaginal discs: reevaluation of some basic concepts (Insecta, Holometabola).

Some general aspects of the concept of imaginal discs in the Holometabola are reevaluated. Their monolayer character and continuity with the surrounding epidermis are confirmed. Studies on the imaginal discs of the silkworm (Bombyx mori) and data from the literature show that the discs and their peripodial cells produce cuticle during larval life, as well as at metamorphosis. In B. mori it is demonstrated that adult and larval antennae are produced by the same cells or their progeny. The results also suggest that segments of the typically three-segmented larval antenna of Holometabola are not scape, pedicel, and one-segmented flagellum; at least segments 2 and 3 are of flagellar origin. Based on these and some additional facts it is argued that: (1) No larval organs are "replaced" at metamorphosis, but strict "sequential homology" is always maintained. (2) Imaginal discs are not undifferentiated structures destined to form the adult after larval breakdown, cannot be unambiguously defined, and do not represent qualitatively different epidermal structures. Classical imaginal discs (invaginated and present also in pre-final larval instars) arose several times independently and were not present in the larvae of ancestral Holometabola. (3) Since the disc cells are not undifferentiated and "embryonic" (if these words have a defined meaning at all), it is unreasonable to expect that the processes taking place in discs at metamorphosis would differ fundamentally from those occurring in other diploid metamorphosing epidermal cells.     

Innate immune responses of Drosophila melanogaster are altered by spaceflight

Alterations and impairment of immune responses in humans present a health risk for space exploration missions. The molecular mechanisms underpinning innate immune defense can be confounded by the complexity of the acquired immune system of humans. Drosophila (fruit fly) innate immunity is simpler, and shares many similarities with human innate immunity at the level of molecular and genetic pathways. The goals of this study were to elucidate fundamental immune processes in Drosophila affected by spaceflight and to measure host-pathogen responses post-flight. Five containers, each containing ten female and five male fruit flies, were housed and bred on the space shuttle (average orbit altitude of 330.35 km) for 12 days and 18.5 hours. A new generation of flies was reared in microgravity. In larvae, the immune system was examined by analyzing plasmatocyte number and activity in culture. In adults, the induced immune responses were analyzed by bacterial clearance and quantitative real-time polymerase chain reaction (qPCR) of selected genes following infection with E. coli. The RNA levels of relevant immune pathway genes were determined in both larvae and adults by microarray analysis. The ability of larval plasmatocytes to phagocytose E. coli in culture was attenuated following spaceflight, and in parallel, the expression of genes involved in cell maturation was downregulated. In addition, the level of constitutive expression of pattern recognition receptors and opsonins that specifically recognize bacteria, and of lysozymes, antimicrobial peptide (AMP) pathway and immune stress genes, hallmarks of humoral immunity, were also reduced in larvae. In adults, the efficiency of bacterial clearance measured in vivo following a systemic infection with E. coli post-flight, remained robust. We show that spaceflight altered both cellular and humoral immune responses in Drosophila and that the disruption occurs at multiple interacting pathways.     

Expanded functions in the apical cell domain to regulate the growth rate of imaginal discs

The Drosophila expanded (ex) gene encodes a product (Ex) that shares homology with the Protein 4.1 family of proteins, many of which are enriched at specific lateral cell junctions and the apical cellular domain. Ex colocalizes with actin in the apical domain of imaginal disc epithelial cells, where it partially overlaps the distribution of phosphotyrosine (PY)-containing proteins. This suggests that Ex is present in or associated with adherens junctions. Genetic studies show that Ex is necessary for proper regulation of final cell number in adult wings and for the formation of eyes, distal leg, and distal antennal segments. We have generated mitotic clones that lack Ex using the twin spot technique, and demonstrated that the primary function of Ex is to regulate cell proliferation. Overexpressing Ex protein results in a decrease in final cell number in wings, suggesting a direct relationship between Ex function and proliferation rate. Dev. Genet. 20:103–110, 1997. © 1997 Wiley-Liss, Inc.     

Patterns of cell division in imaginal discs of Drosophila

A transmission electron microscopic study of cell division in serially sectioned imaginal discs of early third instar fruitfly larvae revealed that mitotic cells maintain a relationship with the basal surface of the disc through thin cytoplasmic extensions abutting on vesicular material. Two patterns of cell division were discerned. In one, cell divisions were isolated and usually found near the peripodial membrane-disc epithelium junction; in the other, cell divisions were clustered. Observations on cell death and cell division in the peripodial membrane are also reported.