Adult Fat Body Research Articles

Developmental control of Drosophila yolk protein 1 gene by cis-acting DNA elements.

In this paper we have demonstrated that at least two tissue-specific cis-acting DNA elements are necessary for the normal developmental pattern of Drosophila yp1 gene expression. One of these elements is necessary for expression in the adult female follicle cells and the other is necessary for expression in adult female fat bodies. We have localized the fat body expression element to a 125-bp fragment that lies between nucleotides -196 and -321 of the yp1 gene. This small fragment has at least one of the characteristics of enhancer sequences because it can direct a heterologous Drosophila promoter to be transcribed with the yp1 fat body expression pattern.

The effects of synthetic locust adipokinetic hormone on dispersed locust fat body cell preparations: cAMP induction, lipid mobilization, and inhibition of protein synthesis

A procedure for the preparation of functional cells from adult locust fat bodies by collagenase treatment has been developed. The high variability of replicates encountered when whole fat bodies are incubated in vitro is greatly reduced in incubation of the dispersed cells. Synthetic locust adipokinetic hormone (AKH) (40 n M) stimulated release of lipids from the dispersed fat body cells at a rate comparable to that observed using whole fat bodies in vitro. Synthetic AKH elevated cAMP levels sixfold in dispersed cells. In addition, AKH inhibited protein synthesis to a maximum of 50–70% in a concentration-dependent manner. None of these actions of AKH required the presence of locust hemolymph components. These results demonstrate the utility of the isolated locust fat body cells for investigating hormonal action in vitro.

Juvenile hormone-dependent vitellogenin synthesis in Locusta migratoria fat body: Inducibility related to sex and stage

Juvenile hormone (JH)-dependent vitellogenin (Vg) synthesis in the fat body of Locusta migratoria is normally limited to sexually mature adult females. As a step toward examining the basis of this limitation, we have tested female and male locusts in a series of stages after the third larval molt for inducibility of Vg synthesis by the synthetic JH analog, methoprene. We find that in the fourth and fifth larval instars fat body of both sexes can be induced to produce Vg, but in the adult stage females respond strongly while no more than trace amounts can be induced in males. Quantitative assays show relative responsiveness in the order: adult female > fifth instar female > fifth instar male ⪢ adult male. During the fifth instar of both sexes, maximal vitellogenic response was obtained in midinstar. After the larval-adult ecdysis, female fat body was unresponsive during the first 4 days, then responsiveness increased and by Day 8 after ecdysis fat bodies were fully as competent to produce Vg as at Day 14, the usual maximum of the first vitellogenic cycle due to endogenous JH. Larval and adult female fat bodies implanted into male larvae are competent for Vg synthesis after metamorphosis, so that the differences between adult male and female cannot be imposed by the male milieu intérieur during the larval-adult molt. In male and female precocious adults, produced by treatment of fourth instars with precocene, fat body responded to methoprene as in normal adults. We conclude that factors intrinsic to the fat body cells, determined early in development, are responsible for differential gene programing in males and females, which is partially expressed by the fifth instar but fully manifest only after a molt in the absence of JH.

Sécrétions protéiques du tissu adipeux chez Rhodnius prolixus (Stål). Étude immunochimique

The proteins present in saline extracts or culture medium of the fat body from fifth-instar larvae or adults of Rhodnius prolixus were examined. Immunochemical analyses show six or seven antigens common to both the haemolymph and the fat body of adults. Continuous gradient gel electrophoresis reveals that most of haemolymph proteins are found in the fat body extract and incubation medium. The three predominant adult haemolymph fractions (lipoprotein 5, glycoprotein 3, and vitellogenin) are identified in the fat body of both female and male adults but no vitellogenin is found in the fifth-instar larvae fat body. Vitellogenin synthesized and secreted in vitro by the adult fat body consists mostly of protein A which is a 260 000 molecular weight lipoglycoprotein. The larger complex of 500 000 (protein B) is mainly found in haemolymph and ovocytes.

Construction and partial characterization of a recombinant DNA probe for locust vitellogenin messenger RNA.

Double-stranded DNA complementary to poly(A)-containing RNA from the fat body of adult female locusts, Locusta migratoria, was synthesized. Hybrid molecules containing this cDNA was constructed in the PstI site of the plasmid pAT 153 by the technique of dC . dG tailing and amplified in Escherichia coli K-12 strain HB 101. Ten colonies of bacteria were identified as carrying recombinant plasmids containing DNA complementary to locust vitellogenin mRNA by (a) 'Northern' blot hybridization analysis and (b) hybrid selection of vitellogenin mRNA and immunological detection of the products of translation of the mRNA. Of the ten recombinant plasmids, one, termed plasmid 4E, containing a cDNA insert of about 650 nucleotides, was characterized in greater detail and a partial restriction map obtained. Using this hybrid plasmid it was possible to derive a value for the average content of vitellogenin mRNA in the adult female locust fat body as 1.5 x 10(5) molecules/cell, and to establish that the haploid genome of L. migratoria contains only one or two genes coding for vitellogenin.

Quantitative relationship between ingested blood and follicular growth in the stable fly, Stomoxys calcitrant

Female stable flies, Stomoxys calcitrans (L.), fed blood at 24-h intervals, required five blood meals to produce their first batch of eggs. The first three blood meals mainly promoted growth of the adult fat body but the latter two meals resulted in rapid ovarian growth accompanied by a decline in fat body weight. Thus, the lack of gonotrophic concordance in these flies can probably be attributed to a shortage of reserves carried over from the larval stage. Ovariectomy at emergence resulted in hypertrophy of the adult fat body in blood-fed females, indicating that the ovary regulates the flow of nutrients from the fat body during vitellogenesis.

Vitellogenins and other haemolymph proteins involved in the oogenesis of Rhynchosciara americana

The electrophoretical analysis of haemolymph proteins from Rhynchosciara americana, in the presence of sodium dodecyl sulphate revealed the occurrence of two vitellogenic polypeptides, V 1 and V 2, with molecular weights of 240,000 and 220,000 respectively. These polypeptides are synthesized by the adult female fat bodies and are found in the yolk granules. The vitellin molecule is formed at least by the polypeptides V 1, V 2 and two others, V 4 and V 5 with molecular weights of 67,000 and 55,000 respectively. All these polypeptides can be stained by PAS and by methyl green, suggesting that they are phosphoglycopeptides. Haemolymph proteins, other than vitellogenin, participate in egg formation, the most prominent being a monomeric larval protein of molecular weight 47,000. These larval proteins are found in the soluble fraction of the eggs and make a massive contribution to the total egg protein.

Tissue-specific effects of the juvenile hormone analogue ZR 515 during metamorphosis in Drosophila cell cultures

The juvenile hormone analogue ZR 515 has specific effects on ecdysone-induced metamorphic differentation of Drosophila cells cultured in vitro. The number of vesicles containing imaginal cuticular structures is reduced to 10% of control levels. Similarly, the differentiation of adult fat body is partly inhibited by ZR 515. The differentiation of adult tubular and fibrillar muscles, however, is not affected. ZR 515 does not inhibit cuticle secretion by tracheal cells and larval epidermal cells.

Heterogeneity of juvenile hormone binding compounds in fat bodies of a cockroach

Fat bodies of adult vitellogenic or non-vitellogenic females and those of adult males contain 2 populations of saturable JH binding compounds. The respective K d values are 2–5 × 10 −9 M and 1–4 × 10 −8 M. Fat bodies of penultimate nymphs, hemolymph of any animal, and midgut epithelia contain one saturable JH binder with a K d of 1–5 × 10 −8 M. Unlabeled JH I and III, as well as the JH analog ZR-515 compete effectively for the binding sites in these tissues. The 2 classes of hormone binders were separated by DEAE ion-exchange column chromatography. The very high affinity molecules of the adult fat bodies are presumably the specific JH receptors which may function in translocation of the hormone to the nucleus. The compound with relatively lower hormone affinity may be the JH hemolymph carrier which is also produced by the fat bodies and then exported.

Juvenile hormone-stimulated polyploidy in adult locust fat body

The relative DNA contents of adult fat body nuclei of Locusta migratoria have been assayed by scanning microspectrophotometry after Feulgen staining, spermatid and brain nuclei being used as haploid and diploid reference standards. In the fat body, several DNA classes were found, the mean ploidy increasing during the period of reproductive maturation. The extinction frequency peaks fall at values that are less than doublings of the diploid value, which suggests incomplete replication of the genome, although this interpretation requires confirmation by independent techniques. On Day 1 the female and male fat body nuclei are chiefly tetraploids. Elevation to octaploidy or higher, especially in the female is evident by Day 15. Allatectomy, removing the source of JH, prevents the increase in ploidy, and administration of the JH mimic, ZR-515, restores it in both sexes. Incorporation of [ 3H]thymidine in female fat body shows a broad peak, maximal at Day 6. After allatectomy, incorporation of [ 3H]thymidine is strongly stimulated by ZR-515 in both sexes. It is suggested that the JH-dependent polyploidy in female fat body permits accelerated production of mRNA for vitellogenin and possibly other proteins. The male fat body responds to JH by DNA replication but not by expression of the vitellogenin gene.

Age dependent ultrastructural changes in different organs of the mecopteran fly Panorpa vulgaris

1. 1. Age related ultrastructural changes in flight muscle, fat body, malpighian tubules, salivary glands, and optic lobes of male mecopteran flies Panorpa vulgaris were studied. 2. 2. In all these organs striking degenerative changes arise with increasing age. Flight muscle degeneration is characterized by myofibril dissolution, deformed and “whorl-like” mitochondria and an extensive hypertrophy of sarcoplasmic reticulum which cover the whole muscle in 13-week old adults. In fat body of aged adults a fatty “cirrhotic degeneration” takes place and a change in fatty acid composition of lipids is assumed. Malpighian tubules in 91-day old individuals show abundant inclusions of the myeloid-type and renal stone-like spheric residual bodies. The mitochondria completely disappeared from the brush border and exhibit different stages of degeneration. The salivary glands of old males lack the protein granules lying in the lumen of the young glands. In the optic lobes (neuropile area) glial and neuronal cytoplasm become reduced with advanced age. Scattered autophagic vacuoles and residual bodies are also present. In contrast, no changes in the optic apparatus itself could be detected. 3. 3. The results were compared with those of Phormia terrae novae, where no such degenerative changes occur.

SELECTION FOR AMYLASE ALLOZYMES IN DROSOPHILA MELANOGASTER: SOME QUESTIONS.

The laboratory selection experiments for amylase allozymes of Drosophila melanogaster reported by Hickey (1977) elicited word of caution from Yardley (1978) and reply to Yardley from Scharloo and de Jong (1980). Results of these experiments, in conjunction with earlier ones described by de Jong and Scharloo (1976), were taken as strong evidence for the view that environmental factors can have a specific effect on allozyme alleles. This would support the selectionist explanation for the maintenance of allozymic variation in natural populations. Without becoming embroiled in the selectionist versus neutralist controversy, I wish to raise some questions of a technical nature about the above experiments, as well as others recently published by Hoorn and Scharloo (1978a, 1978b, 1979, 1980). The questions concern the genetic backgrounds of Amy stocks and stem, in part, from work (Doane, unpubl.) on amylase allozymes in D. melanogaster and D. hydei and linked adp (adipose; Lindsley and Grell, 1968) mutants in the former species. The answers may prove critical to interpretations being placed on Hickey's data and perhaps bear on work from Scharloo's laboratory. Yardley (1978) and Scharloo and de Jong (1980) already raised an important point about Hickey's experimental populations; namely, his base population was a cross between two laboratory stocks with no attempt at randomization of the genetic background. The stocks, Amy'- and Amy4'6, came from my laboratory where they had been made isogenic for chromosomes 1, 2 and 3 and coisogenic for chromosomes 1 and 3 many years ago. An Oregon-R strain was the source of the Amylc stock and provided the coisogenic portions of the Amy4'f stock. Chromosome 2 in the Amy4'6 stock which carries the Amy region (2.77.7, see Doane, 1969), came from the Kaduna, Nigeria, wild population maintained at Edinburgh, Scotland. This chromosome also contains the recessive mutant adp6 (adipose 60 : 2-83.4) for which it is homozygous (Doane, 1963a, 1969). Clearly, adp6O was introduced into Hickey's initial replicate populations. The question is: Might the presence of adp6' in these cage populations have played a role in the outcome of his experiment? If so, his results warrant re-evaluation. Evidence follows which suggests that this, indeed, may be the case. Furthermore, there is a possibility that the adp6' mutant may have influenced results of some experiments done in Scharloo's laboratory as well, because the Amy' and Amy4'6 strains used there were also derived from a Kaduna population. Preliminary studies (Doane, 1963a, 1963b) showed that adp'; affects lipid and carbohydrate metabolism in larvae and adults in a manner similar to that of its more extensively studied allele, adpf` (adiposefeallf-st'rilf, =fs(2)adp), which came from the same Kaduna population (Doane, 1960a, 1960b, 1961; Cummings and Ganetzky, 1972). Unlike the latter allele, adp6'ladp60 females are 45-90% fertile in terms of egg hatchability, depending on their residual genetic background. Homozygotes for either allele develop hypertrophied fat bodies when reared under uncrowded conditions on standard cornmeal-molasses-yeast food medium. The mature adult fat body becomes so filled with liDid reserves at the expense of stored glycogen that it has a characteristic fatty phenotype (Fig. 1) which is visible through the body wall.

Age and endocrine effects on fat body metabolite composition in adult male Blaberus discoidalis cockroaches

AbstractEndocrine‐and age‐related changes in metabolite contents were used as a measure of fat body development and function in adult male Blaberus discoidalis cockroaches. Fat body dry weight, protein, glycogen, and lipid levels coordinately decreased at days two to three of adult life, increased at day four, then declined to lower stable levels by days six to seven. The metabolite contents remained stable from days seven through ten and were still at this level on day 30. Long‐term endocrine deficiencies resulting from corpora cardiaca‐allata extirpation did not change the amounts of metabolites present at 30 days of age. The fat body DNA content remained constant regardless of age or endocrine balance. Unlike other metabolites, the RNA content increased to twice its original level during the first six days of adult life and was unaffected by either age or endocrine balance thereafter. The findings suggest that the adult fat body changes from a storage function at the time of the nymphal‐adult molt to a biosynthetic function by day six. The transition is independent of endocrine control by the corpora cardiaca‐allata complex.

ABSENCE OF LARVAL FAT BODY IN THE BUFFALO FLY, HAEMATOBIA IRRITANS EXIGUA (DIPTERA: MUSC1DAE)

AbstractLarval fat body persists, as spherical particles, in most Diptera at the time of adult emergence. The present study and reports in the literature indicate its presence in 25 species of muscoid flies but not in the buffalo fly. the horn fly. or tsetse.In an investigation involving the dissection of female buffalo flies. Haematobia irritans e.xigua de Meijere. for age determination based on the state of the ovaries, it was found that even newly‐emerged flies lacked the larval fat body characteristic of recently emerged specimens of other higher Diptera. This was taken as a possible indication of a special physiological feature in these economically important flies, and so further observations were made on the occurrence at adult emergence of larval fat body in other higher Diptera.Larval fat body, described and illustrated (as “‘pupal fat body’”) by Anderson (1964). is quite different from adult fat body. It consists of a large number of tiny, solid spheres, usually somewhat opalescent, lying loose in the abdominal haemocoel in both sexes. Following eclosion, decreasing amounts may be found for several days in the body of the adult fly. Once it has disappeared, similar fat body is not found again during the lifetime of the fly.The species known to have larval fat bodies persisting in the adult (Table) cover a wide taxonomic range in the Muscoidea. No remnants of the larval fat body are observed in the abdomen of newly‐emerged H. i. exigua, though there may be a variable amount of normal adult fat body. The closely related horn fly, H. irritans irritans (L.) likewise lacks larval fat body at eclosion (R. L. Harris pers. comm.), nor is there any mention of larval fat body in the literature on tsetsef (Glossina Wiedemann), and P. A. Langley (pers. comm.) has confirmed from his own experience that it is not found in adult flies of this genus.Persistent larval fat body carries over various food reserves from larva to adult (Thomson 1975), and there is no clear indication as to why it should not persist in adult Haemalobia and Glossina. When found it occurs equally in both sexes, and its presence or absence is thus not likely to be connected with reproductive physiology. It is unlikely to be a reflection of any special features of larval nutrition, for although the Glossina larva is nourished entirely by the parent female, which has access to ample blood meals, the Haematobia larva lives in the dung of bovines. as do several other Muscidae in which larval fat bodies persist.

Juvenile hormone-controlled vitellogenin synthesis in Locusta migratoria fat body : Hormonal induction in vivo

Biosynthetic processes related to the production of vitellogenin (yolk precursor protein) have been examined in the fat body of adult female Locusta migratoria. Vitellogenin-producing capacity was assayed by incubation of fat body with [ 3H]leucine, followed by precipitation from the medium with specific antiserum. In normal development, vitellogenin synthesis began at about Day 7 after emergence and became maximal at about Day 13, when this protein accounted for 60% of the total fat body protein output. The production of other proteins increased to a lesser extent, becoming maximal at about Day 6. The incorporation of uridine into fat body RNA rose to a maximum at Day 8, which coincided with a marked increase in tissue RNA content. The DNA content in adult female fat body approximately doubled between Days 3 and 8. Vitellogenin synthesis, and the increases in RNA and DNA, were prevented by removal of the corpora allata (the source of juvenile hormone). In allatectomized locusts, vitellogenin synthesis was induced by JH or an analog, ZR-515. Applied topically in acetone, these gave steep dose-response curves, half-maximal at 75 and 150 μg, respectively. After a single treatment with ZR-515, fat body vitellogenin production rose slowly during 48 hr, then steeply to a maximum at 72 hr, but after decay of this effect during 10 days, a second application of ZR-515 induced renewed synthesis with little initial lag. Hormone treatment produced a smaller increase in the output of other proteins, and an increase in incorporation into RNA which preceded the major rise in vitellogenin synthesis. Male fat body produced little or no vitellogenin. These results are consistent with action of JH at the gene level.

Properties and activation of phosphorylase kinase from silkmoth fat body

Phosphorylase kinase from pupal fat body of the silkmoth, Hyalophora cecropia, has been characterized with rabbit muscle phosphorylase b as substrate. The enzyme is converted from a non-activated to an activated form by incubation with ATP, Mg 2+ and Ca 2+ in an apparently autocatalytic process which was not stimulated by addition of cyclic AMP. It was not activated by inorganic phosphate (as reported by blowfly flight muscle phosphorylase kinase). The activated enzyme showed greater activity at pH 6.8, relative to that at pH 8.2, than the non-activated enzyme. Activity was stimulated by Ca 2+, half-maximally at about 10 −6 M and maximally at 10 −4 M, but partial activity persisted with EGTA and no added Ca 2+. The Arrhenius plot for the effect of temperature on activity was linear indicating an activation energy of 11,000 cal/mole. The K m for phosphorylase b was about 1 mg/ml both at 0°C and at 30°C. Activation of phosphorylase kinase may underlie the activation of fat body phosphorylase by shock stimuli, but the data do not explain the mechanism of phosphorylase activation induced by cold. Fat body of tobacco hornworm adults ( Manduca sexta) exhibits phosphorylase phosphatase activity, catalyzing the conversion of rabbit muscle phosphorylase a to phosphorylase b.

Influence of diet on synthesis and utilisation of lipids for reproduction by the tsetse fly Glossina morsitans

Fatty acid composition of lipids from adult Glossina morsitans was unaffected by an in vitro fed diet of cow blood which induces production of under-sized offspring compared to that of flies fed on a superior diet of pig blood. The commonest fatty acids were C 16:0, C 16:1 and C 18:1 and only small differences in proportions were detected between virgin and pregnant females. Rate of lipid accumulation by males and females was the same and was unaffected by diet, but males achieved a maximum of 2.5 mg on day 9 while both virgin and fertilised females reached a maximum of 5,0 mg on day 14 of adult life. Lipid content of pregnant flies then fell to 3.0 mg on the day of larviposition and accumulation began again. A cow blood diet reduced the extent to which the lipids were utilised for larval growth and this was reflected in an altered secretory activity cycle in the female uterine gland. However, no effect on the growth of adult fat body was detectable in such flies. Mating and fertilisation, which influence reproductive events through activity of the endrocine system do not seem to affect the acquisition of lipid reserves by female Glossina. However, they clearly exert considerable influence over distribution of such reserves between fat body and uterine gland, which distribution is also affected by diet.

Factors conducive to increased protein feeding by the blowfly Phormia regina

ABSTRACT. Each period of elevated protein consumption by Phormia regina was associated with some recognizable physiological event that requires protein synthesis (e.g. tanning, adult fat body development, vitellogenesis). Injecting cycloheximide to inhibit protein synthesis prior to the onset of such events delayed the events and also the development of the associated periods of protein hunger. Injecting cycloheximide after a known period of protein synthesis had occurred, however, did not postpone the normally associated protein hunger even while the inhibitor was fully active. The findings conform to a hypothesis that an unidentified deficit, created by protein synthesis, underlies the tendency by Phormia to increase its ingestion of proteinaceous materials. The effects which different hormones have on protein hunger are evidently indirect results of their control of different periods of protein synthesis.

Cyclic nucleotide-dependent protein kinase activity of adult female locust fat body

Locust fat body protein kinase is dependent on, and maximally stimulated by 5 × 10 −6 M cyclic GMP and to a lesser extent by cyclic AMP. It exhibits a pH optimum of 6.9, an unusual requirement for high Mg 2+ (maximum at 100 mM) or an alternative requirement for Mn 2+ (maximum at 5 mM). Maximal stimulation by Mg 2+ is twice that by Mn 2+. Rabbit muscle protein kinase inhibitor uncharacteristically inhibits both cyclic AMP and cyclic GMP stimulated protein kinase activity of locust fat body, the former 5-fold more than the latter, with histone IIA as acceptor. The degree of phosphorylation differs for different acceptor proteins: histone IIA > locust vitellin > arginine-rich histone. Casein is barely phosphorylated and bovine serum albumin (fraction V) not at all. The high threshold for Mg 2+ activation may be related to the high levels of Mg 2+ in locust haemolymph, to the preferential phosphorylation of vitellin, and to the fact that the fat body is the site of vitellogenin synthesis.

Molecular forms of four aldolases from larval and adult stages of the blowfly Phormia regina (Meigen)

Fructose diphosphate ldolases in crude muscle extracts of larval and adult Phormia regina are indistinguishable by polyacrylamide gel electrophoresis (PAGE) in several buffer systems. By contrast, larval and adult fat body aldolases differ in electrophoretic mobility, a difference shown to be due to the inequality of their net charge. Phormia regina larval muscle and fat body isozymes are clearly distinguishable by PAGE, and by pH optima and Michaelis constants.