Ecdysterone responsive functions in the mutantl(1)su(f) ts67g ofDrosophila melanogaster

Summary Transferring the temperature sensitive mutantl(1)su(f) ^ts67g from 25° C to 30° C before or early in the third larval instar blocks the increase in the ecdysterone titer that normally occurs at the end of the larval period. Feeding exogenous ecdysterone to these hormone-deficient larvae results in the formation of pseudopupae. The mutant was used to study ecdysterone-inducible functions in late larval salivary glands by preparing three animal samples with different hormone titers: the titer was low in one sample because of an earlier temperature shift, high in a second sample because the larvae were subsequently transferred to ecdysterone-supplemented food, and also high in a third sample that was kept at 25°C, providing a control for normal development. The effect of the different hormone conditions was studied by³⁵S-methionine labeling of the salivary gland proteins during the larval to prepupal transition and the prepupal period. The results indicate that synthesis of several of the proteins normally appearing during the transition and prepupal period is induced by exogenous ecdysterone.     

Autonomous yolk protein synthesis in ovaries ofDrosophila cultured in vivo

Summary Immature ovaries ofDrosophila mercatorum were injected into young larvae and into adult males ofD. mercatorum, D. melanogaster, D. hydei, D. virilis, andZaprionius vittiger. These homo- and heteroplastic transplantations allow normal vitellogenesis to occur in the donor ovary. By SDS gel electrophoresis, we identified the major species-specific yolk proteins of mature eggs (stage 14) which were exclusively of donor-specific origin. Other experiments withD. hydei andZ. vittiger showed that, when females were used as hosts, the host-specific yolk proteins became incorporated into the donor eggs. When two immature ovaries, one ofD. mercatorum and one ofD. hydei, were co-cultured in males, again only the donor-specific yolk proteins were found in the mature eggs implying that these yolk proteins were not released into the host hemolymph.A parthenogenetic strain ofD. mercatorum was used to demonstrate the ability of transplanted immature ovaries to produce viable eggs which can give rise to fertile adults.The role of the species-specific yolk proteins is discussed with respect to the dual origin of these proteins during normal vitellogenesis, i.e., an autonomous synthesis within the ovary itself in addition to the well-known production by the fat body. Further experiments with pupae as hosts indicate that even in the absence of juvenile hormone and in the presence of high doses of ecdysone, vitellogenesis can proceed within the donor ovary.Based on these experiments, a new hyopthesis on the hormonal control of vitellogenesis inDrosophila is presented. We propose that yolk proteins derived from the fat body are controlled by juvenile hormone, whereas the independent and autonomous vitellogenesis within the ovary itself is controlled by endogenously synthesized ecdysone.     

The effect of temperature onshibire ts cell clones in the compound eye ofDrosophila melanogaster

Summary We have analysed the effect of temperature on both developing and adult eye cell clones homozygous forshi ^ST139, a temperature-sensitive mutant ofDrosophila melanogaster. The mutant gene, autonomous in its cellular expression, causes structural modifications of ommatidial cells when adult clones of cells are exposed to the restrictive temperature (29°C) for several days. However, the mutant phenotype reverses to normal within 4 days at the permissive temperature (20°C). The results of pulse, shift-up and shift-down experiments show that the temperaturesensitive period for developing compound eye cells is from the late second instar up to the early pupa. Cytodifferentiation of compound eye cells is blocked by restrictive temperature treatment during this period, whereas cell proliferation does not seem to be directly affected. These results are discussed with regard to the other known aspects of the phenotype observed in mutant individuals.     

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.     

A maternal effect mutation leading to deficiencies of organs and homeotic transformations in the adults ofDrosophila

Summary The temperature sensitive mutationfs(l)h is characterized at the restrictive temperature of 29°C by both a maternal effect responsible for the early embryonic lethality and pupal zygotic lethality. The two phenotypes are inseparable and map at a short deletion in the X chromosome (7Dl, 7D5-6). At semipermissive temperatures, hemizygous mutant females produce adults with morphological defects, such as organ deficiencies and homeotic transformations of haltere to wing and third leg to second leg. These defects depend on the maternal genotype and are governed by an early temperature sensitive period, which covers the end of oogenesis and the first hours of embryogenesis. Furthermore, this maternal effect mutation interacts with some dominant mutations of the bithorax system. These properties suggest thatfs(l)h is somehow involved in segmental determination.     

The Developmental Genetics of the Temperature-Sensitive Lethal Allele of the Suppressor of Forked, l(1)su(f)ts67g, in DROSOPHILA MELANOGASTER

A temperature-sensitive lethal allele of suppressor of forked, l(1)su(f)(ts67g) (ts67), has been discovered and characterized as follows: Flies which are hemizygous for ts67 live at 18 degrees and 25 degrees but die at 30 degrees primarily as larvae. The temperature-sensitive period for ts67 homozygotes or hemizygotes begins in second instar and ends at pupation. ts67 is lethal at 30 degrees when heterozygous with suppressor of forked (su(f)), a deficiency for suppressor of forked (su(f)(-)), and a non-conditional lethal allele of suppressor of forked (3DES). It is viable at 30 degrees when heterozygous with the wild-type allele of suppressor of forked. At 25 degrees but not at 18 degrees forked bristles are suppressed in flies of the following genotypes: f(s)ts67/Y, f(s)ts67/f(s)ts67, f(s)ts67/f(s)su(f), f(u)ts67/f(s)3DES, f(u)ts67/f(s)su(f)(-), f(u)ts67/f(s)su(f). There is some suppression of forked bristles at 25 degrees in the heterozygote, f(s)ts67/f(s)+(su(f)). The forked bristle phenotype is not suppressed at either temperature in flies of the genotypes f(u)ts67/Y, f(u)ts67/f(u)ts67/ (f(s) and f(u) indicating suppressible and unsuppressible alleles of forked). The temperature-sensitive period for suppression of forked bristles begins at pupation and extends through the period of bristle synthesis. The deficiency phenotype (bristles reduced in size or absent, wing wrinkled or blistered, eyes rough) typical of flies of the genotype f(s)su(f)/f(s)su(f)(-) at 18 degrees and 25 degrees , is exhibited by flies of the genotypes f(s)ts67/f(s)su(f)(-) at 25 degrees and f(u)ts67/f(s)su(f) at 29 degrees . An allele of lozenge (lz(1)) which can be suppressed by su(f) is suppressed at 25 degrees but not at 18 degrees in lz(1)ts67/Y males. ts67 homozygous females are fertile at 25 degrees but sterile at 30 degrees . The hypothesis is discussed that the su(f) locus codes for a ribosomal protein and that suppression and enhancement are affected by mutations at the locus by mutant ribosome-induced misreading. The possibility is presented that ts67 may be used to determine the translation time in development of any gene.     

Identification of a female-sterile mutation affecting yolk protein 2 in Drosophila melanogaster

Summary The three yolk proteins (YP1, YP2 and YP3) of Drosophila melanogaster are synthesised in the fat body and ovarian follicle cells and selectively accumulated in the developing oocytes to provide a nutrient source for embryogenesis. We have described the phenotype of a temperaturesensitive female-sterile mutant, fs(1) K313, and characterised its yolk proteins. This mutation affects the secretion of YP2 and is the first mutation affecting YP2 to be described. Using genetic and molecular tests we argue that the female-sterile phenotype results, at least in part, from the abnormal secretion of YP2 perturbing the follicle cell secretory pathway in general and thus causing defects in chorion protein secretion. The gene coding for YP2 in fs (1) K313 has been cloned and sequenced. Two amino acid substitutions have been found which probably cause the abnormal secretion of YP2 and the resulting female-sterile phenotype.     

cell clones and pattern formation: Studies onsevenless,a mutant ofDrosophila melanogaster

Summary sev ^LY3,the only existing allele at thesev locus (1–33,2±0,2), behaves as strongly hypomorph or even as amorph. Ommatidia in asev compound eye have only seven receptor cells, the position of the R7 pattern element being vacant. Various criteria showing that the missing cell is R7 have been verified. These include (i) anatomical characteristics ofsev ommatidia; (ii) behaviour of central R cells insev rdgB double mutants; (iii) medullary projection of central R cell axons; and (iv) mitotic pattern ofsev imaginal discs. The analysis of morphogeneticsev-sev ⁺ mosaics has shown thatsev is expressed autonomously by R7 cells, indicating that thesev phenotype is not due to asev genotype of ommatidial pattern elements other than R7. The study of third instarsev imaginal discs has not brought any direct evidence for death of clustered presumptive R7 cells; however, clonal analysis of the developingsev compound eye has given evidence of developmental parameters comparable to those ofsev ⁺, therefore favouring the hypothesis that R7 cells die insev mutants. On the other hand,sev ⁺ seems to be required for the determination of the R7 cells, since thesev phenotype cannot be uncovered during the last mitoses of heterozygous mutant cells.     

The Drosophila melanogaster LEM-domain protein MAN1

Here we describe the Drosophila melanogaster LEM-domain protein encoded by the annotated gene CG3167 which is the putative ortholog to vertebrate MAN1. MAN1 of Drosophila (dMAN1) and vertebrates have the following properties in common. Firstly, both molecules are integral membrane proteins of the inner nuclear membrane (INM) and share the same structural organization comprising an N-terminally located LEM motif, two transmembrane domains in the middle of the molecule, and a conserved RNA recognition motif in the C-terminal region. Secondly, dMAN1 has similar targeting domains as it has been reported for the human protein. Thirdly, immunoprecipitations with dMAN1-specific antibodies revealed that this Drosophila LEM-domain protein is contained in protein complexes together with lamins Dm0 and C. It has been previously shown that human MAN1 binds to A- and B-type lamins in vitro. During embryogenesis and early larval development LEM-domain proteins dMAN1 and otefin show the same expression pattern and are much more abundant in eggs and the first larval instar than in later larval stages and young pupae whereas the LEM-domain protein Bocksbeutel is uniformly expressed in all developmental stages. dMAN1 is detectable in the nuclear envelope of embryonic cells including the pole cells. In mitotic cells of embryos at metaphase and anaphase, LEM-domain proteins dMAN1, otefin and Bocksbeutel were predominantly localized in the region of the two spindle poles whereas the lamin B receptor and lamin Dm0 were more homogeneously distributed. Downregulation of dMAN1 by RNA interference (RNAi) in Drosophila cultured Kc167 cells has no obvious effect on nuclear architecture, viability of RNAi-treated cells and the intracellular distribution of the LEM-domain proteins Bocksbeutel and otefin. In contrast, the localization of dMAN1, Bocksbeutel and otefin at the INM is supported by lamin Dm0. We conclude that the dMAN1 protein is not a limiting component of the nuclear architecture in Drosophila cultured cells.     

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.     

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.)     

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.     

Genetic,molecular and developmental analysis of the glutamine synthetase isozymes ofDrosophila melanogaster

The glutamine synthetase isozymes ofDrosophila melanogaster offer an attractive model for the study of the molecular genetics and evolution of a small gene family encoding enzymatic isoforms that evolved to assume a variety of specific and sometimes essential biological functions. InDrosophila melanogaster two GS. isozymes have been described which exhibit different cellular localisation and are coded by a two-member gene family. The mitochondrial GS structural gene resides at the 21B region of the second chromosome, the structural gene for the cytosolic isoform at the 10B region of the X chromosome. cDNA clones corresponding to the two genes have been isolated and sequenced. Evolutionary analysis data are in accord with the hypothesis that the twoDrosophila glutamine synthetase genes are derived from a duplication event that occurred near the time of divergence between Insecta and Vertebrata. Both isoforms catalyse all reactions catalysed by other glutamine synthetases, but the different kinetic parameters and the different cellular compartmentalisation suggest strong functional specialisation. In fact, mutations of the mitochondrial GS gene produce embryo-lethal female sterility, defining a function of the gene product essential for the early stages of embryonic development. Preliminary results show strikingly distinct spatial and temporal patterns of expression of the two isoforms at later stages of development.     

Developmental studies on two ecdysone deficient mutants ofDrosophila melanogaster

Summary This paper describes two ecdysone-deficient, recessive-lethal mutants,lethal(1)giant ring gland (grg) andlethal(1)suppressor of forked ^mad-ts (mad-ts: Jürgens and Gateff 1979) and compares their ecdysteroid titers with that of the wild-type. Mutant larvae show a much reduced ecdysteroid content, amounting to 1/10 to 1/30 of the wild-type values, but never a true titer peak. They fail to pupate and die after 1–3 weeks. Ecdysteroid feeding elicits different responses in the larvae of the two mutants.mad-ts larvae pupate within 24 h, thus showing that their low ecdysteroid titer is directly connected to their inability to pupate.mad-ts resembles the mutantlethal (3)ecdysone-1 ^ts (Garen et al. 1977). Thegrg mutant larvae, on the other hand, fail to pupate after 20-hydroxyecdysone feeding as well as injection. The primary defect of thegrg mutant is not entirely clear. Thegrg larval salivary gland cells appear to possess normal ecdysteroid receptors. Furthermore, the low ecdysteroid titer ingrg is not the result of an increased ecdysteroid catabolism. The primary defect in the mutant may lie in the malfunctioning neurosecretory cells which do not show neurosecretion in histological preparations. Further support for this notion comes from electronmicrographs of the enlargedgrg ring glands which, in contrast to the wild-type, do not possess nerve endings.In the wild-type three ecdysteroid peaks were found: one shortly before puparium formation, the second at approximately 12 h and the third at about 30 h after pupation. The ecdysteroid titer peak in late third instar, wild-type larvae is mainly due to the presence of 20-dydroxyecdysone as shown by radioimmunoassays after thin layer chromatography and derivatization followed by gas liquid chromatography and mass spectroscopy. In addition, a number of unidentified polar and apolar metabolites were also present.     

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.     

The role of the peripodial membrane in the morphogenesis of the eye-antennal disc ofDrosophila melanogaster

Summary The early morphogenesis of the eye-antennal disc ofDrosophila in response to 20-hydroxy ecdysone involves the curling of the eye anlagen dorsally over the antenna. During this process, the area of the peripodial membrane is substantially reduced. The peripodial membrane is taut at this stage, and if it is cut the curling of the disc cannot continue, and the eye anlagen returns to its original position within one minute of the operation. In contrast, cutting the columnar epithelium between the eye and antennal anlagen does not disrupt curling, but actually facilitates it. During curling, the cells of the peripodial membrane appear healthy, and exhibit basal extensions. We suggest that the curling of the eye is mediated by the conversion of cuboidal peripodial membrane cells into pseudostratified columnar epithelium at the edges of the peripodial membrane. Subsequently, cells of the peripodial membrane secrete first a pupal cuticle, and then an imaginal cuticle.     

Polymorphisms in the genomic distribution of copia-like elements in related laboratory stocks ofDrosophila melanogaster

Summary The genomic distributions of the copia, 297, 412, mdg 1, and B 104 transposable elements have been compared by the Southern technique among two Oregon R and four Canton SDrosophila laboratory lines that have been maintained separately for defined periods of a few years. The heterogeneity of the autoradiographic patterns suggests that multiple transposition events have occurred during the time of separation. The hypothesis that transposition could be induced by, variations of environmental parameters is discussed.Presented at the FEBS Symposium on Genome Organization and Evolution, held in Crete, Greece, September 1–5, 1986     

fs(1)K10, a germline-dependent female sterile mutation causing abnormal chorion morphology inDrosophila melanogaster

Summary Females homozygous for a newly isolated mutation induced by ethyl methane sulphonate,fs(1)K10, lay abnormally shaped eggs in which the dorsal appendages of the chorion are enlarged and fused ventrally. The eggs are usually not fertilized and development is never normal beyond the blastoderm stage. The mutant was mapped to the tip of the X-chromosome with a meiotic position of 1–0.5 and a cytological location between 2B17 and 3A3. Using germ line mosaics constructed by transplantation of pole cells, it was shown that the abnormal morphology and the sterility are obtained only when the germ line is homozygous for the mutant.     

Homologies of positional information in thoracic imaginal discs ofDrosophila melanogaster

Summary The regulative behavior of fragments of the imaginal discs of the wing and first leg was studied when these fragments were combined with fragments of other thoracic imaginal discs. A fragment of the wing disc which does not normally regenerate when cultured could be stimulated to regenerate by combination with certain fragments of the haltere disc. When combined with a haltere disc fragment thought to be homologous by the criteria of morphology and the pattern of homoeotic transformation, such stimulated intercalary regeneration was not observed. Combinations of first and second leg disc fragments showed that a lateral first leg fragment could be stimulated to regenerate medial structures when combined with a medial second leg disc fragment but not when combined with a lateral second leg disc fragment. Combinations of wing and second leg disc fragments showed that one fragment of the second leg disc is capable of stimulating regeneration from a wing disc fragment while another second leg disc fragment fails to stimulate such regeneration. It is suggested that absence of intercalary regeneration in combinations of fragments of different thoracic imaginal discs is a result of homology or identity of the positional information residing in the cells of the fragments. The pattern of correspondence of positional information revealed by this analysis is consistant with the pattern of homology determined by morphological observation and by analysis of the positional specificity of homoeotic transformation among serially homologous appendages. The implications of the existence of homologous positional information in wing and second leg discs which share a common cell lineage early in development are discussed.