Rudimentary mutants ofDrosophila melanogaster

Summary The X-linkedrudimentary (r) mutants ofDrosophila melanogaster are pyrimidine auxotrophs and require exogenous pyrimidines (Nørby, 1970; Falk, 1976). We have established a set ofrudimentary cell lines that are derived from embryos, homozygous for eitherr ¹ orr ³⁶. The enzymatic activities of the pyrimidine synthesizing enzymes were measured in the mutant lines. We have further investigated the nutritional requirements of the mutant cells in vitro by using a pyrimidine free culture medium.Ther ¹ cell lines were found to express 3–7%dihydroorotase (DHOase) activity as compared to a wildtype cell line. Reducedaspartate transcarbamylase (ATCase) activity was measured in somer ¹ cell lines whereas wildtypecarbamylphosphate synthetase (CPSase) activity is expressed in allr ¹ cell lines. Ther ³⁶ cell line expresses wildtype activity ofDHOase andCPSase. ATCase activity was found to be reduced to 10% of the wildtype activity.The mutant cell lines do not proliferate in pyrimidine free minimal medium and cell proliferation is obtained by the addition of crude RNA. Proliferation of ther ¹ cells is restored by the supplementation of the minimal medium withdihydroorotate whereas proliferation of ther ³⁶ cells is restored by supplementation with eitherdihydroorotate orcarbamylaspartate.The results demonstrate that therudimentary phenotypesr ¹ andr ³⁶ are expressed at the cellular level and that the two mutant cell types behave as cellular pyrimidine auxotrophs in vitro.     

An ultrastructural and developmental analysis of the corpus allatum of juvenile hormone deficient mutants ofDrosophila melanogaster

Summary The ultrastructure of the corpus allatum of theapterous mutantsap ⁴ andap ^56f ofDrosophila melanogaster during larval-pupal-adult metamorphosis and adult life was correlated with the gland's ability to synthesize juvenile hormone in vitro. During the early wandering period of the third instar of both mutants, a high concentration of smooth endoplasmic reticulum, mitochondria and mitochondrion-scalariform junction complexes are typical features of an active corpus allatum cell. Juvenile hormone biosynthesis by the glands is high at that time and, in fact, only slightly lower than that of wild type glands. In contrast to the wild type gland, the cells of the pupal and pharate adult corpus allatum of both mutants contains highly electron dense mitochondria with tubular cristae but no whorls of smooth endoplasmic reticulum nor glycogen clusters. The frequency and size of the lipid droplets, putatives depots of the juvenile hormone precursors, in cells of theap ^56f gland is a function of the insect's age, but both are lower than in wild type gland cells. Juvenile hormone biosynthesis by both mutant glands remains at the basal level when compared to increased synthesis by the wild type gland. The frequency and density of lipid droplets in cells of theap ⁴ corpus allatum are much lower than in theap ^56f glands. During adult life, the ultrastructural profile of theap ^56f corpus allatum is similar to that of the wild type gland although the in vitro production of juvenile hormone by the former is much lower than that of the wild type gland. The ultrastructural features of the adult corpus allatum ofap ⁴ homozygotes reveal precocious degeneration and support the view that this non-vitellogenic mutant is a juvenile hormone deficient mutation.     

Behavioral analysis ofDrosophila mutants displaying abnormal male courtship

We describe six recessive autosomal male sterile mutations inDrosophila, generated by mobilization of single P-elements, exhibiting abnormal male courtship behavior. Detailed analysis of courtship behavior elicited by virgin wild type females indicated that five of the six mutants are affected in the early steps of courtship. The sixth mutant is blocked at the step of attempted copulation which occurs later in the courtship sequence. All of the mutants have normal olfactory responses and normal locomotor activity. No defect in the visual modality has been observed for the five mutants affected in the initiation of courtship. The mutant blocked at attempted copulation lacks the ‘on’ and ‘off’ transients, but this appears to be due to genetic background rather than the mutation itself. Abnormal spermatogenesis was observed in five of the mutants. Spermatogenic defects vary and include lesions in the proliferation of the germline, in meiosis, and in the differentiation and maturation of the spermatids into motile sperm.     

Extra tarsal joints and abnormal cuticular polarities in various mutants ofDrosophila melanogaster

Summary The legs of flies from 16 different mutant strains ofDrosophila melanogaster were examined for abnormal cuticular polarities and extra joints. The strains were chosen for study because they manifest abnormal cuticular polarities in some parts of the body (10 strains) or because they have missing or defective tarsal joints (6 strains). All but three of the stocks were found to exhibit misorientations of either the bristles, hairs, or “bract-socket vectors” on the legs. The latter term denotes an imaginary vector pointing from a hairlike structure called a “bract” to the bristle socket with which it is associated. On the legs of wild-type flies nearly all such vectors point distally, as do the bristles and hairs. In the mutant flies, the most common vector misorientation is a 180° reversal. When the bract-socket vectors of adjacent bristle sites in the same bristle row point toward one another, the distance between the sites is frequently abnormally large, whereas when the vectors point in opposite directions, the interval is frequently abnormally small. This correlation is interpreted to mean that bristle cells actively repel one another via cytoplasmic extensions that are longer in the direction of the bract-socket vector than in the opposite direction. Repulsive forces of this kind may be responsible for “fine-tuning” the regularity of bristle spacing in wild-type flies. Extra tarsal joints were found in eight of the 16 strains. A ninth strain completely lacking tarsal joints appears in some cases to have an extra tibia-basitarsus joint in its tibia. Whereas the tarsi of wild-type flies contain four joints, the tarsi ofspiny legs mutant flies contain as many as eight joints. In this extreme extra-joint phenotype, four of the joints correspond to the normal wild-type joints, and there is an extra joint in every tarsal segment except the distal-most (fifth) segment. Nearly all such ectopic extra joints have inverted polarity. In other strains the extra tarsal joints are located mainly at the wild-type joint sites, and joints of this sort have wild-type polarity. The alternation of normal and inverted (extra) joints inspiny legs resembles the alternation of normal and inverted (extra) body segment boundaries in the embryonic-lethal mutantpatch, suggesting that tarsal and body segmentation may share a common patterning mechanism.     

Regulation and metamorphosis of the abdominal histoblasts ofDrosophila melanogaster

Summary The development of the adult abdomen ofDrosophila melanogaster was analyzed by histology, microcautery, and genetic strategies. Eight nests of diploid histoblasts were identified in the newly hatched larva among the polytene epidermal cells of each abdominal segment: pairs of anterior dorsal, posterior dorsal, and ventral histoblast nests and a pair of spiracular anlagen. The histoblasts do not divide during larval life but begin dividing rapidly 3 h after pupariation, doubling every 3.6 h. Initially they remain confined to their original area, but 15 h after pupariation the nests enlarge, and histoblasts replace adjacent epidermis cell by cell. The histoblasts cover half the abdomen by 28 h after pupariation and the rest by 36 h. Polytene epidermal cells of the intersegmental margin are replaced last. Cautery of the anterior dorsal nest caused deletion of the whole corresponding hemitergite, whereas cautery of the posterior dorsal nest caused the deletion of the macrochaetae of the posterior of the hemitergite. Cautery of the ventral nest deleted the hemisternite and the pleura, whereas cautery of the spiracular anlagen deleted the spiracle. Results of cautery also revealed that no macrochaetae formed on the tergite in the absence of adjacent microchaetae. Clonal analysis revealed that there were no clonal restrictions within a hemitergite at pupariation. Cautery of polytene epidermal cells other than those of the intersegmental margin failed to affect tergite development. However, cautery of polytene epidermal cells of the intersegmental margin adjacent to either dorsal histoblast nest caused mirror-image duplications of the anterior or posterior of the hemitergite in 10% of the hemitergites. Forty percent of the damaged presumptive hemitergites formed complete hemitergites, indicating extensive pattern regulation and regeneration. Pattern duplication and regeneration were accounted for in terms of intercalation and a model of epimorphic pattern regulation (French et al., 1976). Histoblasts in adjacent segments normally develop independently, but if they are enabled to interact by deleting the polytene epidermal cells of the intersegmental margin, they undergo intercalation which results in duplication or regeneration. The possible role of the intersegmental margin cells of insects in development was analyzed.     

Temperature sensitivity ofTumorous head,a homoeotic mutant ofDrosophila melanogaster

Summary A temperature-sensitive period during early embryogenesis for three stocks carrying thetuh-3 gene suggests that it is a homoeotic mutation involved in the initial determination of the eye-antennal disc rather in maintenance of the determination.     

br regulates the expression of the ecdysone biosynthesis gene npc1

The growth and metamorphosis of insects are regulated by ecdysteroid hormones produced in the ring gland. Ecdysone biosynthesis-related genes are both highly and specifically expressed in the ring gland. However, the intrinsic regulation of ecdysone biosynthesis has received little attention. Here we used the Drosophila npc1 gene to study the mechanism of ring gland-specific gene expression. npc1 is important for sterol trafficking in the ring gland during ecdysone biosynthesis. We have identified a conserved ring gland-specific cis-regulatory element (RSE) in the npc1 promoter using promoter fusion reporter analysis. Furthermore, genetic loss-of-function analysis and in vitro electrophoretic mobility shift assays revealed that the ecdysone early response gene broad complex (br) is a vital factor in the positive regulation of npc1 ring gland expression. Moreover, br also affects the ring gland expression of many other ecdysone biosynthetic genes as well as torso and InR, two key factors in the regulation of ecdysone biosynthesis. These results imply that ecdysone could potentially act through its early response gene br to achieve positive feedback regulation of ecdysone biosynthesis during development.     

Transdetermination in leg imaginal discs ofDrosophila melanogaster undDrosophila nigromelanica

Summary The transdetermination capacities of leg discs ofDrosophila melanogaster were examined by mechanically disrupting and kneading whole discs from late third instar larvae and by culturing the resulting tissue mass for 10–14 days in adult female abdomens where the cells continued to divide. The grown implants were then dissected from the abdomens and injected into third instar larvae to undergo metamorphosis.After this treatment, prothoracic leg discs ofDrosophila melanogaster transdetermined with a high frequency (59% of all implants) to wing. Mesothoracic leg discs also transdetermined to wing, but at a very low frequency (4%). Metathoracic leg discs exhibited the same low frequency of transdetermination (4%), but in this case the direction of transdetermination was to haltere (Table 1,D. melanogaster).Very similar results were obtained with leg discs ofDrosophila nigromelanica (Table 1,D. nigromelanica), showing that the peculiar behavior of the three leg discs is not unique forDrosophila melanogaster.The homeotic mutation Polycomb (Pc ³) which partially transforms meso- and metathoracic legs into prothoracic legs did not significantly increase the frequencies of transdetermination in these leg dises and had clearly no effect on the direction of transdetermination (Table 1).We dedicate this publication to the memory of our teacher and advisor, the late Professor Ernst Hadorn, whose enthusiasm and interest stimulated our work     

Pattern specification in imaginal discs ofDrosophila melanogaster

Summary l(1)su(f)^mad-ts (mad) is a new temperature-sensitive (ts) lethal mutant ofDrosophila melanogaster which produces duplicated legs after temperature pulse treatment during larval development. The ts-lethality was studied in temperature experiments and genetic mosaics. Temperature pulses given during two distinct TSPs of larval development result in two different types of leg pattern duplication. Total differ from partial duplications with respect to the affected leg compartments and the orientation of the planes of symmetry which are perpendicular to the dorso-ventral and the proximo-distal leg axes in total and partial duplications, respectively. Genetic mosaic studies indicate (i) disc autonomy of leg pattern duplication, (ii) clonal separation of the anlagen of the two pattern copies, and (iii) clonal restriction along the antero-posterior compartment border in the two pattern copies of totally duplicated legs.The results suggest thatmad leg pattern duplication is caused by a change in positional information rather than by cell death and subsequent regeneration. Our data are compatible with the assumption that during normal development the leg disc cells acquire information about their position within the disc with respect to the different leg axes independently and at different times.     

Genetic analysis of pattern-formation in the embryo ofDrosophila melanogaster

Summary The mutationbicaudal (Bull, 1966) causes embryos to develop a longitudinal mirror image duplication of the posteriormost abdominal segments, while head and thorax are missing. These embryos occur with varying frequencies among eggs laid by mutant females, irrespective of the paternal genotype. Recombination and deletion mapping indicate thatbicaudal (bic) is a recessive, hypomorphic, maternal-effect mutation mapping at a single locus on the second chromosome ofDrosophila melanogaster close tovg (67.0±0.1). The frequency of bicaudal embryos depends on the age of the mother, her genetic constitution and the temperature at which she is raised. Best producers are very young females hemizygous forbic (bic/Df(2)vg ^B ) at 28° C. Under these conditions 80% to 90% of the eggs which differentiate can show the bicaudal embryo phenotype. Upon ageing of the mother the frequency of bicaudal embryos declines rapidly, and most of the eggs develop the normal body pattern. Temperature shift experiments suggest a temperature-sensitive period at the onset of vitellogenesis.The mutation causes several types of abnormalities in the segment pattern of theDrosophila embryo, which are interpreted as various degrees of expression of the mutant character. The most frequent abnormal phenotype is the symmetrical bicaudal embryo with one to five abdominal segments duplicated. Less frequent are asymmetrical types, in which the smaller number of segments is always in the anterior reversed part. Other phenotypes are embryos with missing or rudimentary heads, and embryos with irregular gaps in the segment pattern. In bicaudal embryos, the pole cells, formed at the posterior pole of the egg prior to blastoderm formation, are not duplicated at the anterior. The significance of thebicaudal phenotypes for embryonic pattern-formation inDrosophila is discussed.     

Differentiation in vitro of larval and adult muscles from embryonic cells ofDrosophila

Summary The differentiation of muscles in primary cultures of cells fromDrosophila melanogaster embryos was investigated. In early cultures, and in the absence of exogenous ecdysone, two main classes of muscle were found. Comparison, by light and electron microscopy, of one of these classes (the myotube class) with muscles from third instar larvae shows that this class corresponds to the muscles of the body wall of the larva. When - or -ecdysone is added to the cultures, these undergo a number of metamorphic changes. Most of the larval muscles disappear, and two new types of muscle form. Ultrastructural and light microscopic examination of these two types indicates that they correspond to the two classes of skeletal muscle (fibrillar and tubular) found in adult flies.     

Sequential gene activation by ecdysteroids in polytene chromosomes ofDrosophila melanogaster

Summary Changes in polytene chromosome 3 L puffing patterns in the fat body ofDrosophila melanogaster larvae and prepupae are compared to those in the salivary gland. While some general features are common to the two tissues, there are differences which reflect their different developmental roles. In vitro experiments with fat body chromosomes show that they have a distinct response to ecdysteroids which is different from that of salivary gland chromosomes, and which does not,in this culture system, reproduce the changes observed in normal development. In short term culture experiments, the fat body chromosomes appear more sensitive to ecdysteroids than the salivary gland chromosomes and, although 20-OH ecdysone is more active than ecdysone in these assays, the possibility is not excluded that ecdysone has a role in normal development as it appears to alter gene activity at physiological levels in these cells.     

Growth is required for cell competition in the imaginal discs ofDrosophila melanogaster

Summary Imaginal wing discs from late third-instar larvae were gammairradiated to induce clones of rapidly growingMinute ^– cells in a background of slowly growingMinute cells and culturedin vivo for periods up to 18 days. Clones in discs cultured for 16 to 18 days did not grow significantly larger than clones in uncultured controls, indicating that competition between populations of cells having potentially different mitotic rates does not occur in imaginal discs after their growth is completed.     

The evagination of the genital imaginal discs ofDrosophila melanogaster

Summary The morphology of the evaginating female genital disc ofDrosophila melanogaster was examined at different stages of metamorphosis. The observations show that the internal genital organs are derived from the anterior half of the disc and that their morphogenesis is mainly a protrusion of the different primordial areas of the disc epithelium. The external genital and anal derivatives originate from the posterior half of the disc, which undergoes complex rearrangements during metamorphosis. The disc opens along the posterior margin and the dorsal and ventral epithelia evert and thereby completely reverse their anteroposterior orientation. Dramatic elongation has been observed during the formation of the seminal receptacle. The cells of the repressed male genital primordium do not form any recognizable structures and are assumed to be eliminated during metamorphosis.     

The development of ommatidial patterning in metamorphosed eye imaginal discs implants ofDrosophila melanogaster

Summary The development of the rhabdomeric pattern in the compound eye ofDrosophila has been studied using combined transplantation and electron microscope techniques. In a first series of experiments eye imaginal discs of increasing age were implanted into larvae ready to pupate, thus losing variable amounts of the normal time for development. A sequence of differentiative abilities was found in the metamorphosed test pieces. As far as the photoreceptor cells are concerned, the most prominent steps of this sequence are: ability to form groups with other similar elements, anatomical polarization of microvilli, establishment of the rhabdomeric pattern and formation of an equator line. The stability of determination of the equator line was tested in a second experimental series. Fragment of different topographical origin within the mature eye anlage were brought to metamorphosis by implantation into larvae ready to pupate. It was found that an equator line differentiates only in those pieces which according to the published anlage maps contain the prospective equator region prior to metamorphosis. The mitotic abilities of implanted eye imaginal discs were investigated by means of in vitro³H-thymidine pulse-labelling and light microscope autoradiography of the differentiated test pieces. During the third larval stage the eye anlage is traversed by two consecutive mitotic waves, each one of them producing different categories of receptor cells. The first, anterior wave predominantly produces cells oriented toward the poles of the eye within the ommatidia, while the second, posterior wave gives rise to elements exclusively in an equatorial position. The dynamics of this proliferation are discussed in relation to the findings in the implantation experiments. Silver-grain counts support the possibility that at least two successive cell divisions occur in the eye anlage between labeling with tritiated thymidine and beginning of morphological differentiation. The relevance of this finding for the understanding of the concept of acquisition of competence is discussed.     

Early development of the thoracic discs ofDrosophila

Summary We estimate the number of blastoderm cells which generate the thoracic imaginal discs ofDrosophila. At hatching the wing disc is twice the size of the haltere disc, but the results suggest that both discs develop from a similar number of blastoderm cells. Two homeotic mutations, which transform the haltere into wing, affect embryonic growth but not the primordial number. All the segmental primordia may be of similar size and each may be similarly subdivided into a larger anterior, and a smaller posterior polyclone.     

Differentiation capacities of the labial imaginal disc ofDrosophila melanogaster

Summary The mature labial disc, when implanted into a larva of the same age, undergoes metamorphosis along with the host and produces one lateral half of the medi- and distiproboscis. On the basis of results obtained from transplanted disc halves (including the separate peripodial membrane) a tentative fate map of the labial disc was constructed, which shows most of the presumptive mediproboscis to be located in the dorsal, and most of the presumptive distiproboscis in the ventral part of the disc. The distal protion of the peripodial membrane also contains imaginal anlagen, viz. part of the mediproboscis, prementum, and labellar cap anlagen. The involvement of this part of the peripodial membrane was checked by a careful histological analysis of labial disc development during the first ten hours after prepupation. The results were compared with the situation described forCalliphora imaginal discs.In addition, a detailed morphological analysis was made of the proboscis of the homoeotic mutantproboscipedia (pb). At 27°C,pb changes the distiproboscis into a telopodite (leg segments distal to the coxa); the (unchanged) prementum may therefore correspond to the coxa. At 15° C, the tarsus of this homoeotic telopodite is replaced to a greater or lesser extent by an arista. The present analysis thus confirms (a) the fundamental morphological correspondence of the medi- and distiproboscis with the labium of other insects, and (b) the fundamental developmental correspondence of the labial, antennal, and leg discs.K. K. was a member of the 8th International Research Group in Developmental Biology, and was the recipient of a UNESCO travel grant.)     

Heat shocks do not mobilize mobile elements in genomes ofDrosophila melanogaster inbred lines

Summary Males of three inbred lines ofDrosophila melanogaster were heat-shocked 90 min at 37°C. The progenies from treated and untreated males mated with untreated females of the same line were checked for their chromosomal insertion patterns of various mobile elements by either in situ hybridization or Southern blots. No modification in the pattern of insertion of the elements studied was observed after heat treatment. Hence, heating males of our inbred lines did not mobilize mobile elements, contrary to recent reports on other lines ofDrosophila melanogaster.     

Interaction between a picornavirus and a wild population ofDrosophila melanogaster

Summary Drosophila C virus (DCV) has a considerable impact on ovarian morphogenesis inDrosophila melanogaster host populations. This virus also affects the developmental time and the fresh weight of infected females. In order to investigate the hypothesis that DCV may play a role in the dynamics ofDrosophila populations, the fertility and embryonic and larvo-pupal death rates of a host population and that of five DCV-free populations were determined. A comparison of two populations, one of them DCV-free, the other infected, suggested that the fertility of the DCV-infected flies was higher than that of uninfected flies, despite a greater larvo-pupal death rate. Fertility of the infected flies was greater among the infected population than for the DCV-free populations. The DCV-free populations originated from five different localities. The virus clearly does have an impact on the biotic potential of its host population. This paper reports for the first time a positive interaction between a viral population and a host population as it increases certain parameters of host population dynamics.