Segmental organisation of the tail region in the embryo of Drosophila melanogaster

Summary The segmental organisation of the tail region in the embryo of Drosophila melanogaster, which is defined here as the epidermal region posterior to the boundary between abdominal segments A7 and A8, has been investigated by means of ultraviolet (UV) laser fate-mapping and phenotypic analysis of embryonic mutants that alter the segmental pattern of the larval cuticle. Wild-type embryos were irradiated in the presumptive tail region with a UV- laser microbeam of 20 m diameter at the blastoderm stage. The ensuing defects were scored in the cuticle pattern of the tail region of the first-instar larva, which is described in detail in this paper. The spatial distribution of defect frequencies was used to construct a blastoderm fate-map of the cuticle structures of the larval tail region. The segmental origin of the larval tail structures was inferred from the phenotypic analysis of segmentation and homoeotic mutants, which revealed pattern repetition throughout the embryonic tail region corresponding to four segment anlagen, A8 to A11, and a non-segmental telson. These data enabled the transformation of the blastoderm fate-map of cuticle structures into a map of tail segment anlagen. The tail anlage occupies about 10% of the egg length (EL), bounded by segment A7 anteriorly at 20% EL and by the proctodaeum posteriorly at 10% EL, as measured from the posterior pole. The anlagen of segments A8 and A9 appear to be narrow dorso-ventral strips of blastoderm cells similar to the anlagen of the trunk segments, whereas the anlagen of A10 and A11 are smaller and produce fewer pattern elements. The telson is represented in the cuticle by the tuft which derives from a very dorsal posterior position. The antero-posterior axis of the entire tail anlage appears curved upward posteriorly. Differences in the mode of development between tail and trunk segments are discussed, as are similarities of larval and imaginal tail development in Drosophila. Comparison with tail development in other insects suggests that, during evolution, the transition from semi-long-germ to long-germ development modified the organisation of the tail region without affecting its primary subdivision into metameric units.     

Segmental organisation of the head in the embryo of Drosophila melanogaster

Summary Embryos of Drosophila melanogaster were irradiated in the presumptive head region with a UV-laser microbeam of 20 m diameter at two developmental stages, the cellular blastoderm and the extended germ band. The ensuing defects were scored in the cuticle pattern of the head of the first-instar larva, which is described in detail in this paper. The defects caused by irradiating germ band embryos when morphologically recognisable lobes appear in the head region were used to establish the segmental origin of various head structures. This information enabled us to translate the spatial distribution of blastoderm defects into a fate map of segment anlagen. The gnathal segments derive from a region of the blastoderm between 60% and 70% egg length (EL) dorsally and 60% and 80% ventrally. The area anterior to the mandibular anlage and posterior to the stomodaeum is occupied by the small anlagen of the intercalary and antennal segments ventrally and dorsally, respectively. The labrum, which originates from a paired anlage dorsally at 90% EL, is separated from the remaining head segments by an area for which we did not observe cuticle defects following blastoderm irradiation, presumably because those cells give rise to the brain. The dorsal and lateral parts of the cephalo-pharyngeal skeleton appear to be the only cuticle derivatives of the non-segmental acron. These structures derive from a dorso-lateral area just behind the putative brain anlage and may overlap the latter. In addition to the segment anlagen, the regions of the presumptive dorsal pouch, anterior lobe and post-oral epithelium, whose morphogenetic movements during head involution result in the characteristic acephalic appearance of the larva, have been projected onto the blastoderm fate map. The results suggest that initially the head of the Drosophila embryo does not differ substantially from the generalised insect head as judged by comparison of fate map and segmental organisation.     

Origin of the coelomic cavities and the heart in the polypterids (Pisces)]

In polypterus the mesodermal cavities appear quite late during embryonic life. They are generally small and they only get somewhat more voluminous in the anterior mesomeres (where they establish the pronephrie chambers) and in the ventral anterior mesoderm (where they become an embryonic pericardial cavity). The anlage of the heart appears in the anterior part of the tissue that is situated between the paired mesodermal cavities of these stages. It assumes some unawaited dispositions that are truly confusing in the case of a superficial inspection. It is only during larval life that a coelomic cavity appears all along the truncal part of the mesoderm. In the beginning this is a pericardio-peritoneal cavity. But because of the coalescence between several membranes an anterior cavity gets isolated and this one is the pericardial cavity of the adult specimens; this cavity is much more limited than its homonymic counterpart of the embryonic stages.     

RNA synthesised during oogenesis and early embryogenesis in an insect egg (Euscelis plebejus)

Summary RNA labelled during oogenesis or early embryogenesis was isolated from eggs of the leaf hopperEuscelis plebejus. The polyadenylated RNA fraction deposited during early oogenesis accounted for approximately 2.7% of the total RNA content of the newly laid egg. This fraction differed significantly in molecular weight (15–32 S) from poly(A)-containing RNA synthesised between early cleavage and early germ anlage stages (4–20S). Locally injected³H-uridine spread through the egg within approximately 3 h. A considerable fraction (25–35%) of label injected as³H-uridine during early cleavage was recovered in DNA at subsequent stages (10–20 h later); labelled RNA was not found prior to the cellular blastoderm stage. When the yolk-endoplasm was separated from the blastoderm cells, only the latter contained demonstrable amounts of RNA synthesised by the embryo. Of the precursor incorporated into embryonic RNA, approximately 10% was found in the polyadenylated fraction at the early blastoderm stage, but only 3% at the early germ anlage stage. No differences in size distribution of polyadenylated RNA were evident between anterior and posterior halves of the early germ anlage stage.Supported by the Deutsche Forschungsgemeinschaft, SFB 46     

The development and function of the female germ line in Drosophila melanogaster: a cell lineage study.

X-ray-induced mitotic recombination was used to follow the development and function of the female germ line in Drosophila melanogaster. Clones marked by maternal effect mutations which alter the morphology of the egg [fs(1)K10] or the phenotype of the resulting progeny (maroonlike) were produced in trans-heterozygotes irradiated during embryonic, larval, or pupal development or as 5-day-old adults. Judging from the size of clones induced at the blastoderm stage, only five to ten of the pole cells observed on the surface of the embryo contribute to the germ line. Most of the K10 clones induced during embryonic and larval development were associated with mal twin spots, indicating that both daughters of the irradiated germ cell remained in the germ line and gave rise to eggs in the adult. During larval life the number of cells increases logarithmically and reaches a maximum of 110 at 24 hr after pupation. The same value was obtained for 5-day-old adults. In contrast to the mosaic females produced as embryos and larvae, mosaics obtained after pupal and adult irradiations were of two types, those laying only one K10 egg and those laying several K10 eggs distributed over the lifespan of the adult. This result indicates that the stem cell divisions characteristic of the adult period have begun shortly after pupation. About 9 to 11 days are required for an irradiated stem cell to produce its first clonal K10 egg, and two-thirds of this time is spent in the germarium. Each ovariole possesses on the average two to three functioning stem cells. This multiplicity of stem cells was confirmed by the recovery of mosaic ovarioles when mal heterozygotes irradiated as adults or late larvae were stained for aldehyde oxidase activity.     

The longitudinal visceral musculature of Drosophila melanogaster persists through metamorphosis

The larval gut of Drosophila is coated with visceral muscles of mesodermal origin. In the midgut region this musculature comprises circular and longitudinal fibres. The complete visceral musculature is described to be removed during metamorphosis and to be replaced by a newly differentiated imaginal tissue resembling the morphology of the larval musculature. However, progenitors of this imaginal visceral musculature have never been detected prior to differentiation. Here I present results indicating that the longitudinal visceral musculature of the midgut completely persists through metamorphosis. Single cells expressing green fluorescent protein (GFP) as a marker were transplanted at the blastoderm stage. All clones contributing to the longitudinal visceral musculature detected in third instar larvae were recovered after metamorphosis in adult flies. Further evidence for the persistence of the larval visceral musculature was obtained from the P[Gal4] insertion line 5053A. It expresses GAL4 specifically in the longitudinal visceral muscles of the midgut of all developmental stages to the adult fly beginning at the end of embryogenesis. By using GFP as a reporter, it was possible to follow these cells through the entire metamorphosis. Although the muscles undergo dramatic morphological changes including the loss of their contractile system, no evidence for a replacement of the larval visceral musculature by imaginal precursor cells was detected.     

Contribution of ventral blood island mesoderm to hematopoiesis in postmetamorphic and metamorphosis-inhibited Xenopus laevis

In an effort to label very early erythrocyte and lymphocyte populations and to follow their fate in normally developing postmetamorphic frogs and goitrogen-treated permanent larvae, diploid (2N) and triploid (3N) ventral blood island (VBI) mesoderm was exchanged between neurula stage embryos (about 16-22 hr old). Beginning at 15 days of age, half of the 2N or 3N hosts were treated with sodium perchlorate to prevent thyroxine-induced developmental changes. At larval stages 55-59 (41-48 days) and at 1-2 months postmetamorphosis (110-120 days), the untreated control chimeras and age-matched perchlorate-treated chimeras were killed for analysis of the VBI contribution to blood, spleen, and thymus populations by flow cytometry. The data suggest that grafting of ventral blood island mesoderm is an effective way to label an early larval erythrocyte population that declines after metamorphosis. In perchlorate-blocked permanent larvae this early VBI-derived erythrocyte population persists. In contrast, grafting of VBI mesoderm was less useful as a method to label a larvally distinct lymphocyte population in the thymus and spleen. At the late larval stages that we examined, the proportion of VBI-derived cells in thymus and spleen was not different from that observed after metamorphosis. Inhibition of metamorphosis interfered with the thymocyte expansion that normally occurs after metamorphosis, but the proportion of VBI-derived cells in thymus and spleen was not affected. This suggests that lymphopoiesis occurring in late larval life and after metamorphosis uses a stable persisting population of VBI-derived stem cells as well as dorsally derived stem cells.     

Ontogenesis of the ovary in a moth midge,Tinearia alternata Say (Diptera: Psychodidae)

In a psychodid, Tinearia alternata, the initial differentiation of the polytrophic ovary occurs during the early larval stages. Early in development, each ovary anlage is a solid organ subdivided into three distinct zones: the cortex houses germ cells and somatic interstitial tissue, while two other somatic regions will give rise to the oviduct calyx and anterior part of the lateral oviduct. Germ cell cluster formation precedes the development of ovarioles. Each ovariole houses only one functional egg chamber. All ovarioles within paired ovaries are developmentally synchronized. In the larval ovaries, the newly formed egg chambers and then the ovarioles are intermingeled with and surrounded by the somatic interstitial tissue of the ovary cortex. The interstitial cells give rise to all the somatic elements of the ovarioles. In the pupal ovaries, the remaining interstitial tissue degenerates; thus, the ovarioles protrude into the body cavity. The ovaries in psychodids develop relatively large and swollen oviduct calyxes that are equivalent to receptaculum seminis (spermatheca). The morphological differentiation of germ cells within the egg chambers starts during late larval/early pupal stages. Nurse cell nuclei contain prominent nucleoli and polytene chromosomes. Oocyte growth results from accumulation of yolk and then, in the final stages of oogenesis, from an inflow of cytoplasm from the nurse cells. J. Morphol. 236:167–177, 1998. © 1998 Wiley-Liss, Inc.     

The development of the imaginal abdomen of Drosophila melanogaster

The development of the imaginal Drosophila melanogaster was investigated by an analysis of clones of cells marked by somatic crossing-over. The results showed that each abdominal half tergite is initiated by about 11 cells which fail to divide during late embryonic and larval stages. Cell division commences only at the time of pupariation, and continues for the next 32 hr. The spatial pattern of marked clones indicated that each tergite consists of a left and right side—marked cells do not cross the midline. Twin spots were examined and indicated that two daughter cells are aligned preferentially in the transverse orientation. These results are contrasted to data obtained for other imaginal anlage, particularly with regard to the time of cessation of cell division.     

Analysis of morphogenetic movements in the development of the notum anlage of Drosophila melanogaster

Summary A comparison of the morphogenetic maps of the notum anlage of Drosophila melanogaster derived from the gynandromorph data and mosaics induced by somatic crossing-over during the first instar larval stage revealed that practically no major morphogenetic movements occur in the development of the anlage between the blastoderm and first instar larval stages and the adult stage. By comparing the morphogenetic map derived from gynandromorphs and the fate map derived from data on the transplantation of fragments of the mature wing imaginal disc, it was observed that no major morphogenetic movements occur in the notum anlage between the stages of the allocation of the disc and the mature disc. The results are consistent with the observations of other authors concerning the larval development of eye-antenna, wing and leg discs.     

Cytoskeletal F-actin patterns in whole-mounted larval and adult salivary glands of the fleshfly, Sarcophaga bullata.

The patterns of filamentous actin were analysed in different larval, pupal and adult stages in the salivary glands of the fleshfly Sarcophaga bullata. Using the rhodamine labelled phalloidin staining method in combination with detergent extraction specific actin filament distribution was detected. The salivary glands which are histolysed during the process of metamorphosis show distinct cellular morphology and actin filament patterns in larvae and adults. The large third instar larval salivary gland cells contain a well developed apicolateral microvillar zone. In third instar larvae this microvillar zone invaginates and expands in the basal part of the lateral membranes. Larval salivary gland cells also contain numerous parallel basal actin bundles. The larval glands are histolysed during metamorphosis and adult glands are formed out of the imaginal cell group. At the onset of metamorphosis these basal actin bundles form a network of crossing bundles. The filamentous actin patterns of the proximal part of adult gland cells is confined to the apicolateral microvillar membranes. The cells in the distal, tubular part of the adult salivary glands show intense staining of their folded lateral membranes.     

Sequential predator effects across three life stages of the African tree frog, <Emphasis Type="Italic">Hyperolius spinigularis</Emphasis>

While theoretical studies of the timing of key switch points in complex life cycles such as hatching and metamorphosis have stressed the importance of considering multiple stages, most empirical work has focused on a single life stage. However, the relationship between the fitness components of different life stages may be complex. Ontogenetic switch points such as hatching and metamorphosis do not represent new beginnings—carryover effects across stages can arise when environmental effects on the density and/or traits of early ontogenetic stages subsequently alter mortality or growth in later stages. In this study, I examine the effects of egg- and larval-stage predators on larval performance, size at metamorphosis, and post-metamorphic predation in the African tree frog Hyperolius spinigularis. I monitored the density and survival of arboreal H. spinigularis clutches in the field to estimate how much egg-stage predation reduced the input of tadpoles into the pond. I then conducted experiments to determine: (1) how reductions in initial larval density due to egg predators affect larval survival and mass and age at metamorphosis in the presence and absence of aquatic larval predators, dragonfly larvae, and (2) how differences in mass or age at metamorphosis arising from predation in the embryonic and larval environments affect encounters with post-metamorphic predators, fishing spiders. Reduction in larval densities due to egg predation tended to increase per capita larval survival, decrease larval duration and increase mass at metamorphosis. Larval predators decreased larval survival and had density-dependent effects on larval duration and mass at metamorphosis. The combined effects of embryonic and larval-stage predators increased mass at metamorphosis of survivors by 91%. Larger mass at metamorphosis may have immediate fitness benefits, as larger metamorphs had higher survival in encounters with fishing spiders. Thus, the effects of predators early in ontogeny can alter predation risk even two life stages later.     

Genital system anatomy and development of Ovatella myosotis by three‐dimensional computer visualization

Adult anatomy as well as organogenesis of the genital system of the ellobiid pulmonate Ovatella myosotis is investigated in detail by means of serial sectioning and three‐dimensional computer reconstruction and visualization. From the middle portion of the adult, which has four nidamental glands, a spermoviduct leads to a common genital aperture. From here two separate structures, the vas deferens and a groove on the body surface, lead anteriorly. The latter is termed the egg groove because it carries the egg ribbon anteriorly, a function that is recognized here for the first time in the Ellobiidae. The evolution of this structure is discussed. In development, the organ system arises from four separate anlagen: (1) the ovotestis anlage, (2) the pallial anlage giving rise to the hermaphrodite duct, fertilization pouch–spermatheca complex, nidamental glandular complex and spermoviduct, (3) the bursa copulatrix anlage and (4) the anlage of the copulatory organ, vas deferens and egg groove. This development mode strongly resembles that of the siphonariid Williamia radiata, supporting its interpretation as a plesiomorphy in Pulmonata. Similarities in development of primitive pulmonates and evolution in gastropods lead to the assumption that ontogenesis of this organ system reflects evolution to some degree.     

Sequence of the twist gene and nuclear localization of its protein in endomesodermal cells of early Drosophila embryos.

The twist gene is involved in the establishment of germ layers in Drosophila embryos: twist homozygous mutant embryos fail to form the ventral furrow at gastrulation and lack mesoderm and all internal organs. We have determined the sequence of the twist gene, that contains 'CAX' repeats in its 5' moiety, and codes for a protein of 490 amino acids. We have raised anti-twist antibodies that were used to study the distribution of the twist protein in whole mounts and tissue sections of wild-type embryos. Twist protein appears to be a nuclear protein at all developmental stages. It is present over both poles and in the midventral region (endoderm and mesoderm anlagen) at cellular blastoderm stage; later in development, it is detected within the mesodermal layer until its differentiation into somatopleura and splanchnopleura in which some cells are still labelled by anti-twist antibodies.     

Clonal analysis of primordial disc cells in the early embryo of Drosophila melanogaster

Single cells were marked by X-ray-induced somatic recombination at the blastoderm stage (3 hr) and at 7 and 10 hr of development, when the embryos are relatively insensitive to radiation damage. The resulting clones of genetically marked cells were analyzed on the adult cuticle. For the three stages examined there is a decrease in clone size with developmental age indicating proliferation of the primordial disc cells which give rise to the adult cuticular structures. The extent of the clones has been used to study the specificity of determination in the primordial disc cells at the time of induction. A significant fraction of the clones induced at all three stages extend from the antenna into the head. In the thorax, some of the clones induced at 3 hr extend from the second leg into the wing, thus indicating that these cells are not yet disc-specifically determined. No clones, however, were found extending from the first leg into the second or from the second into the third, and by 7 hr all thoracic clones were restricted to a single disc. The results provide evidence that determination at the blastoderm stage is not yet disc-specific.     

Developmental potencies of nuclei from cleavage,preblastoderm, and syncytial blastoderm transplanted into unfertilized eggs ofDrosophila melanogaster

Summary Wild-type nuclei from eggs ofDrosophila melanogaster at various developmental stages and from different regions of the egg—cleavage nuclei, pole nuclei from preblastoderm, and lateral nuclei from syncytial blastoderm—were singly implanted into unfertilizedy w sn ³ lz _50e eggs to determine their developmental potencies.All three types of transplanted nuclei were almost equally effective in initiating development of unfertilized eggs. Development was arrested in one of five critieal embryonic stages or in one of the three larval instars. The frequency of individuals reaching a distinct stage was approximately the same for all three types of donor nuclei. The stage-specific pattern of defects was independent of the type of nucleus transplanted.The deviations from normal development were broadly similar to those seen in controls developing from fertilized eggs which had only been punctured or into which cytoplasm had been injected. Many defective embryos also occurred in these control experiments. These and other observations indicate that a large proportion of irregularly developed individuals found after nuclear transfer can be ascribed to loss of egg material, disturbances in the internal organization of the egg during nuclear implantation, and the difficulty the implanted nucleus has in adjusting to the autonomous processes within the egg, such as the formation and migration of cytoplasmic islands.Some of the defective embryos and larvae originating from nuclear transfer were implanted into adult hosts. After culture for 14 days the early embryonic stages had formed several larval structures, and the late embryonic and larval stages had developed all larval organs. The proliferated imaginal primordia of thesein vivo cultured embryos and larvae, as well as the imaginal disks of the third instar larva, were then implanted into larval hosts with which they passed through metamorphosis and differentiated into imaginal structures. All three types of donor nuclei were capable of producing all adult structures derivedin situ from imaginal disks. The phenotype of these structures waswild-type, thus demonstrating their origin from the transplanted nuclei.The problem as to why not all transplanted nuclei initiated development, and why development after nuclear transplantation was arrested at the third larval instar, at the latest, is discussed.This article is dedicated to Professor Friedrich Seidel on the occasion of his 75th birthday.     

The potentialities of transplanted early gastrula nuclei ofDrosophila melanogaster. Production of their imago descendants by germ-line transplantation

Summary Wild-type nuclei, taken out of cells from five regions of early gastrula embryos, were implanted singly into unfertilizedy w sn ³lz^50e eggs ofDrosophila melanogaster. The different types of nuclei initiated development with nearly equal frequencies of about 60%. 2.9% of the 1073 nuclear transfers developed as far as one of the three larval instars, and one reached the pupal stage.All individuals showed stage-specific patterns of defect. Most of these abnormalities were probably due to some inevitable damage caused by the implantation procedure such as disarrangement of the internal egg morphology and loss of peripheral egg substance. The proportions of individuals arrested at different embryonic and larval stages were similar for the five nuclear groups.Fertile imagos, descendants of all five types of donor nuclei, were produced via germ-line mosaics in two ways: (1) Pole cells of nuclear-transplant blastoderm stages were implanted into the pole cell region of host blastoderm eggs. (2) Gonads were taken from nuclear-transplant larvae and implanted into host larvae. In both cases gametes developed from the transplants as could be recognized from the genotypes of their progeny. By means of suitable crosses it was possible to get clones of flies whose large chromosomes were descended from the chromosomes of only one transplanted nucleus, that is, each clone was the descendant of one somatic nucleus. The data presented show that the nuclei remain omnipotent until the early gastrula stage.