Juvenile hormone-induced vitellogenesis in Apterous4, a non-vitellogenic mutant in Drosophila melanogaster

D. melanogaster females homozygous for the ap⁴ mutant synthesize yolk protein and circulate this protein in the haemolymph at concentrations not different from concentrations found in normal females. However, ap⁴ females deposit little or no yolk protein into developing oöcytes. Topical application of a juvenile hormone analogue (JHA), ZR-515, stimulated sequestration of yolk protein by developing oöcytes of ap⁴ females. JHA had no detectable effects on haemolymph concentrations of yolk protein in either normal or ap⁴ females nor on the protein profiles obtained from electrophoresis of haemolymph samples.     

Ultrastructural changes induced by juvenile hormone analogue in oöcyte membranes of apterous4Drosophila melanogaster

Egg chambers from apterous⁴ (ap⁴), a female sterile mutant of Drosophila melanogaster, show none of the microvilli or pinocytotic vesicles which are a prominent feature of the membrane of the wild-type vitellogenic oöcyte. The studies reported here show that a juvenile hormone analogue (ZR515) stimulates formation of microvilli and pinocytotic vesicles in oöcytes of ap⁴ flies. Within 12 hr after topical application of ZR515 to homozygous ap⁴ females the oöcyte membranes exhibit extensive microvilli and pinocytotic activity. The follicle-cell surface adjoining the oöcyte also shows some changes. In vitro studies in which ap⁴ ovaries were incubated in Schneider's Drosophila tissue-culture medium in the presence of ZR515 with or without female haemolymph, or in the absence of ZR515, showed that the analogue acts alone directly on the ovary to cause formation of microvilli and pinocytotic vesicles on the oöcyte membrane.     

Yolk polypeptide gene expression in Minute Drosophila females

Minutes have been considered for some time to be mutant at the sites of synthesis of some components of the protein synthetic apparatus. To study the hypothetical relationship between Minutes and suboptimal translation, a group of abundant proteins, the yolk polypeptides, was assayed in outcrossed females bearing M(3)w, M(3)h ^y , or M(1)n mutations. Recently emerged Minute females contained a lower amount of yolk polypeptides, in both ovarian and nonovarian tissues, than their non-Minute sisters. This low level correlated with the lower abundance of cytoplasmic RNA in Minutes compared to control females. By 1 week of age, both M(3)w and their non-Minute sibs contained the same amount of yolk polypeptides and the corresponding mRNA. The double heterozygote, ap ⁴/+;M(3)w/+, did not differ in yolk polypeptide content from control flies. M(3)w females demonstrated reduced fecundity during the period of low yolk polypeptide content but gradually increased egg deposition as yolk polypeptide levels rose. These results suggest that the low protein levels are due to the slower maturation of M(3)w, and not to less efficient translation machinery.This work was supported by the NSERC (Canada) and a Queen's University ARC grant.     

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.     

The specificity of yolk protein uptake in cyclorrhaphan diptera is conserved through evolution

Summary Yolk proteins are transported from the hemolymph into the oocytes of insects during vitellogenesis by receptor-mediated endocytosis. Since other hemolymph proteins, both native and foreign, are not accumulated in the oocyte, the process of uptake is selective for yolk proteins. Peptide domains within the yolk proteins must therefore be involved in receptor recognition. With the longterm aim of identifying these domains and to open the possibility of understanding the molecular basis of receptor-mediated endocytosis of yolk proteins, we began investigating how well this mechanism has been conserved in evolution. We studied the uptake of yolk proteins from 13 different Drosophila species and five other dipteran species, namely, Calliphora erythrocephala, Sarcophaga argyrostoma, Musca domestica, Lucilia servicata, and Protophormia terrae-novae, into the ovaries of Drosophila melanogaster and Drosophila funebris. The results from these experiments showed that in all cases the foreign yolk proteins were taken up by the host ovaries, indicating that the mechanism and peptide domains of the yolk proteins involved in recognition of the receptor have been well conserved in dipteran evolution. Offprint requests to: M. Bownes     

Ovarian yolk-protein synthesis in Drosophila melanogaster

The ovaries and fat bodies of Drosophila melanogaster adult females both synthesize yolk polypeptides. In a series of experiments it has been shown that the ovaries become competent to mature in an adult male host, where only ovarian synthesis occurs, very early in metamorphosis and synthesis of yolk polypeptides begins in a time-dependent sequence related to the age of the ovary when transplanted. Maturation of ovaries occurs prior to eclosion when they are transplanted to an earlier developmental stage showing that neither the event of eclosion not the adult environment is essential in triggering yolk-polypeptide synthesis by the ovary. When metamorphosing ovaries are transplanted to a female host they take up host yolk polypeptides from the haemolymph, but this does not lead to the implanted ovary developing substantially better than in a male host where only synthesis by the ovary can occur. The regulation of ovarian yolk-polypeptide synthesis therefore appears to be autonomous to the ovary itself. There may be a trigger early in metamorphosis which induces competence in the ovary so that it subsequently initiates yolk-polypeptide gene expression at eclosion.     

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.     

Establishment ofNeltumius arizonensis(Coleoptera: Bruchidae) on Mesquite (ProsopisSpecies: Mimosaceae) in South Africa

Species ofProsopis(Mimosaceae), or mesquites, are invasive rangeland weeds in South Africa's Western Cape and Northern Cape Provinces. Two bruchid seed-weevil species,Algarobius prosopis(Le Conte) andA. bottimeriKingsolver, were released for biological control in 1987 and 1990, respectively. Seed-feeding biocontrol agents were selected because mesquite pods are valued as livestock fodder. Livestock grazing of bruchid larvae developing in mesquite seeds, however, limits the effectiveness of these agents. Livestock grazing also exacerbates mesquite infestations because scarified seeds are dispersed widely in vertebrate dung. In response to the livestock grazing problem,Neltumius arizonensis(Schaeffer), a bruchid reputed to be capable of ovipositing on immature, tree-borne pods, was released at three sites in Western Cape Province in 1993 and 1994. Small populations ofN. arizonensishave become established at the release sites. Overall,N. arizonensiswas 18 times less abundant thanA. prosopis.In some monthsN. arizonensiseggs were heavily parasitized byUscanasp. (Trichogrammatidae), but the effect of this onN. arizonensispopulation dynamics is uncertain. Western CapeN. arizonensispopulations need more time to increase in size. The introduction of other, more injurious biocontrol agents such as the cecidomyiid bud feederAsphondylia prosopidisCockerell should be considered.     

Nonvitellogenic female sterile mutants and the regulation of vitellogenesis in Drosophila melanogaster.

The genetic and endocrine regulation of vitellogenesis was investigated by studying 18 female sterile mutations that disrupt the development of normal vitellogenic follicles. Applications of exogenous juvenile hormone analog and reciprocal ovarian transplants between flies of different genotypes were employed to accomplish our first two objectives: to find (1) whether the mutation blocked development of the ovary directly, and (2) whether the mutation altered the hormonal milieu. In 15 of the mutants the developmental defect was localized to the ovary, but in the other 3 the ovary was competent to respond to a permissive environment. The internal milieu of these three mutants (ap⁴, fs(3)A1, fs(2)A18) was unable to provoke normal development in wild-type ovaries, suggesting that these mutations cause endocrine defects. Our third objective was to find whether an endocrine organ was itself defective in any of these mutants. The corpus allatum from two of the mutants was unable to provoke vitellogenesis in isolated wild-type abdomens, but corpora allata from wild-type females or from other mutants were able to promote maturation of ovarian follicles in isolated abdomens. Our fourth objective was to find whether any of the mutants were able to produce yolk proteins. Immunoelectrophoresis of fly hemolymph demonstrated that in all mutants tested vitellogenins were found in the blood. These experiments permit four main conclusions. First, they identify the first Drosophila mutants in which an endocrine gland is shown to be intrinsically defective during adulthood. Second, they show that the production of morphologically normal late previtellogenic follicles is not required for the induction of vitellogenin synthesis and secretion. Third, they show that juvenile hormone can cause ovarian follicles to sequester yolk in mutant flies. And finally, they show that mutants with defective corpora allata still synthesize and secrete vitellogenin. Taken together, these conclusions suggest that in Drosophila melanogaster the uptake of vitellogenin into follicles depends upon the availability of juvenile hormone, but that the synthesis and secretion of vitellogenin are independent of both normal ovaries and totally normal corpora allata.     

Ultrastructural studies of male-incubated ovaries of the gypsy moth,Lymantria dispar

Ovaries from Lymantria dispar females were transplanted into an environment lacking the vitellogenin ligand; i.e., the male milieu. Transmission electron micrographs comparing the terminal oocytes of male-grown ovaries and normal ovaries showed that yolk sphere diameters were reduced in the male-grown oocytes. However, there were larger numbers of these small yolk spheres per unit area of cytoplasm, indicating that the coalescence of endosomes into yolk spheres is reduced in the absence of vitellogenin. Although there are larger numbers of yolk spheres in male-grown oocytes, the smaller diameter of yolk spheres resulted in less area being taken up by yolk spheres per unit area of cytoplasm in male-grown oocytes, yielding lowered yolk production. This lowered yolk production is a result at least in part of the lowered number of coated vesicles per unit area of submembrane space and in part of the reduced interfollicular spaces seen in male-grown ovaries.     

Stage specific synthesis of some follicle cell proteins inDrosophila melanogaster

Summary Ovarian protein synthesis in the temperature-sensitive mutantl(1)su(f) ^ts67g was analysed at the permissive and non-permissive temperature by SDS-polyacrylamide gel electrophoresis of³⁵S-methionine labelled ovaries. The synthesis of yolk and three other ovarian proteins of approximative molecular weights of 92K, 82K and 76K, respectively, were affected by the shift to the restrictive temperature. Examination of protein synthesis pattern in staged egg chambers revealed that these three proteins were synthesized at stage 10. Analysis of separated cell types present at stage 10 demonstrate that the three proteins were follicle cell products. We have been unable to identify these proteins as any previously described follicle cell proteins.     

Mutant fs(1) 1163 of Drosophila melanogaster alters yolk protein secretion from the fat body

Summary The mutant fs(1) 1163 of Drosophila melanogaster, which was isolated by Gans et al. (1975) is a recessive homozygous female sterile at 18°C and a dominant female — sterile at 29°C. We reported previously that there are reduced quantities of the largest of the three yolk polypeptides in Drosophila melanogaster in the haemolymph and eggs of this mutant at 29°C (Bownes and Hames 1978 a). In this paper we show that the yolk protein defect maps within approximately 2.5 recombination units of the female sterility at 21±2.5 map units on the X-chromosome. The temperature-sensitive period of the yolk protein defect is after emergence. In vitro labelling of fs(1) 1163 ovaries and fat bodies showed that they were able to synthesise yolk polypeptide 1. Interestingly, studies on the proteins present in the various tissues indicate that the fat body tends to accumulate all three yolk polypeptides in the mutant. This phenotype is partially co-dominant in that an effect is seen in heterozygotes as well as homozygotes and is enhanced by increased temperature. This mutant could therefore have a defect (a) in the structural gene for yolk polypeptide 1, (b) in the processing and secretion enzyme systems; (c) in the fat body or all tissues leading to altered secretion properties.Mutants like fs(1) 1163 which alter specific steps in vitellogenesis should be of value for analysing the genetic and biochemical control of the synthesis, transport and sequestering of the yolk polypeptides during oogenesis.     

Correlation between adult transformation and aldehyde oxidase staining pattern in wing discs ofApterous genotypes inDrosophila melanogaster

Summary The aldehyde oxidase staining pattern in wing discs ofDrosophila melanogaster bearing the genotypesap ^blt /ap ^blt andap ^blt andap ^blt /ap ⁷³ⁿ showns changes from the wild-type pattern. Extensive areas of the presumptive dorsal posterior wing blade, which are normally unstained, have enzyme activity in these mutants. In wings of these genotypes, dorsal posterior structures are replaced by dorsal anterior wing structures. A strong correlation has been found between the frequencies of various staining patterns in the discs and the extent of transformation in the cuticular structures of the wing, which is consistent with the idea that aldehyde oxidase activity can be used as an indicator in the wing disc of this transformation. Unlike the homoeotic mutationengrailed, apterous has not been interpreted as a selector gene yet the work reported here shows thatapterous alleles can cause changes resembling those of theengrailed phenotype both in aldehyde oxidase staining behaviour and in the cuticular transformation.     

Courtship and Mating in the Giant Hairy Desert Scorpion, Hadrurus arizonensis (Scorpionida, Iuridae)

Hadrurus arizonensis is a large, long-lived species of North American desert scorpion with lengthy, stereotyped courtship behaviors that lead to sperm transfer via an external spermatophore. Courtship and mating behaviors in H. arizonensis and other members of the Iuridae family have not been described. H. arizonensis has reproductive behavior similar to that of other scorpions, including the promenade a deux, but with some unique components described here for the first time. Courtship and mating behaviors of H. arizonensis are presented in a flowchart to emphasize its stereotypical nature and suitability for experimental manipulation in field and laboratory studies.     

Effect of the apterous 56f mutation on N -acetyltransferase and alkaline phosphatase activities in Drosophila melanogaster females

Alkaline phosphatase (AP) and N-acetyltransferase (NAT) activities were studied in 1-day-old Drosophila melanogaster females of the apterous ^56f (ap ^56f ) strain, having an decreased level of the juvenile hormone (JH) and a increased level of dopamine as a result of the mutation, and in the Canton S ancestral wild-type strain in the normal conditions and upon an experimental increase in JH titer. The AP and NAT activities in ap ^56f females were significantly lower than in Canton S females in the norm. JH application increased the AP activity of mutant females to the level characteristic to JH-treated wild-type females. Original Russian Text ? E.V. Bogomolova, N.V. Adonyeva, N.E. Gruntenko, I.Yu. Rauschenbach, 2008, published in Genetika, 2008, Vol. 44, No. 5, pp. 710–712.     

Yolk polypeptide secretion and vitelline membrane deposition in a female sterile Drosophila mutant

Ovarian follicle cells of wild type Drosophila melanogaster simultaneously secrete yolk polypeptides (YP1, YP2 and YP3) and vitelline membrane proteins. In order to understand the relationship between these two secretory activities, we have investigated the ultrastructure of a female sterile mutation that alters YP1 secretion and vitelline membrane deposition. Homozygous fs(1)1163 females lay eggs that collapse and contain reduced quantities of YP1. Secretory granules in follicle cells contain an electron-translucent component that is assembled into the developing vitelline membrane in both mutant and wild-type ovaries, and an electron-dense component that disperses after secretion in wild-type ovaries. Mutant ovaries differ from wild-type by (1) having larger secretory granules (2) forming clumps of the dense secretory component within the developing vitelline membrane (3) accumulating more tubules in the cortical ooplasm of vitellogenic oocytes, and (4) possessing altered yolk spheres. Mutant ovaries implanted into wild-type hosts showed no improvement in the secretory granules and slight improvement in the vitelline membrane clumps but amelioration of the oocyte phenotypes. Since genetic evidence suggests that the fs(1)1163 mutation resides in or near the Yp1 gene and biochemical data show that the mutation alters YP1 structure, we conclude that the ultrastructural phenotypes are due to a structurally abnormal YP1 in the mutant. The alteration in vitelline membrane structure caused by the dense clumps could account for collapsed eggs and, hence, the female sterility of the mutant.     

Inheritance patterns of new genetic markers and occurrence of spontaneous mosaicism in the monogenic blowfly Chrysomya rufifacies (Diptera: Calliphoridae)

 Four new genetic markers for Chrysomya rufifacies, a fly with maternal sex determination, were characterized. The markers include one body colour mutant, black body (bl), and three eye colour mutants, brown eye (br), apricot eye (ap), and red eye (w ^r ). Two of the latter, br and w ^r , turn out to be sex linked, the others behave as autosomal genes belonging to different linkage groups. w ^r is a hypomorphic and w an apomorphic mutation of the white gene, w/w is epistatic to br/br and to ap/ap. A preliminary genetic linkage map with the sex realizer F′/f  and the loci br and w residing in homomorphic sex chromosomes is established. Evidence is presented that crossing over is absent in the male sex. The possible causes of the spontaneous appearance of mosaics for eye colour observed among individuals heterozygous for recessive genes are discussed. Received: 18 March 1996/Accepted: 9 August 1996     

Comparison of Yolk Production in Seven Pyralid Moth Species

Summary

The yolk proteins of six pyralid moths were analyzed and compared with the yolk proteins of Plodia interpunctella (Hübner). When cross-reacted in an Ouchterlony double immunodiffusion with antiserum raised to either total yolk proteins or purified vitellin from P. tnterpunctella, the yolk proteins of Anagasta kuehniella (Zeller), Cadra cautella (Walker), C. figulilella (Gregson), and Ephestia elutella (Hübner), closely related members of the subfamily Phycitinae, showed strong precipitation lines that consisted of four major yolk polypeptides (YPs). The yolk proteins from Amyelois transitella (Walker) were only weakly reactive, whereas yolk proteins from Galleria mel-lonella (L.) were not precipitated by either antiserum. Abdominal body walls (containing primarily fat body) from late pharate adult females were incubated in vitro and they secreted two major polypeptides that had molecular masses similar to the vitellogenins (YP1 and YP3) from P. interpunctella. In addition, ovarioles from late pharate adult females were incubated in vitro, and they secreted two major polypeptides that had molecular masses similar to YP2 and YP4 from P. interpunctella. When late pharate adult females were injected with ³⁵S-Met, the hemolymph of all species contained vitellogins that were secreted by their respective body walls in vitro. Ovarioles from injected females contained many labeled polypeptides, but there were four major bands that corresponded consistently to the vitellogenins secreted from the fat body and the two major polypeptides secreted from the ovarioles. These data show that the production of the major YPs in these closely related pyralid species is very similar, and that there is considerable conservation of immunological characters of yolk proteins in the subfamily Phycitinae.     

Genetic analysis of Drosophila vitellogenesis: A molecular weight variant of yolk polypeptide-2 in Drosophila simulans

Summary To assess the likelihood of finding genetic variants for the three major yolk polypeptides (YPs) within the species Drosophila melanogaster, YPs from the five species most closely related to D. melanogaster were investigated. The relative positions of the three YPs were characteristic for each species, and in all cases the mobilities of the YP in the ovary corresponded to that of the YP in the hemolymph of the same species.Different stocks of Drosophila simulans were found to have either of two forms of yolk polypeptide-2 (YP2). The YP2^S polypeptide migrated more slowly than YP2^F by an apparent molecular weight difference of about 700 daltons. The genetic factor responsible for this difference mapped to locus 35 on the X chromosome. The Yp2 allele present specified the mobility of the YP2 polypeptide in both the hemolymph and the ovary. YP1 and YP3 were the same in both Yp2 ^S and Yp2 ^F stocks indicating that they are not affected by the Yp2 gene. Densitometric scans of gels showed that there was more than twice as much YP2^F as YP2^S in the ovaries and hemolymph of homozygous animals. Yp2 ^S/Yp2 ^F heterozygotes contained both fast and slowly migrating YP2. The amount of each YP2 in Yp2 ^F/Yp2 ^S heterozygotes was about half that found in each homozygote. These dosage results suggest that this locus is the structural gene. Peptide mapping showed that the structural element contributing to retarded mobility of YP2^S is unlikely to reside at either end of the molecule. These experiments suggest a cytogenetic location in which to concentrate further investigations on the genetic regulation of YP2 synthesis.