The vitellogenin of Leucophaea maderae: Synthesis as a large phosphorylated precursor

Phosphorylation of vitellogenin (yolk protein precursor) and vitellin (major yolk protein) polypeptides of Leucophaea maderae was studied by [³²P]ortho phosphate labeling and subsequent sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) autoradiography. The vitellogenin molecule was isolated from the hemolymph and fat body by antibody precipitation and high-performance liquid chromatography (HPLC), and shown to consist of at least five polypeptides (“subunits”) which had apparent molecular masses of 155, 112, 95, 92 and 54 kD. Labeling studies with ³²P showed that the covalently attached phosphorus was distributed in an uneven fashion among the five polypeptides. The two heavily-phosphorylated polypeptides, 112 and 54 kD, corresponded to the large and small, mature vitellin subunits. Quantitative SDS-PAGE analysis of long-term ³²P-labeled vitellin showed that these large and small “subunits” contained 55 and 30%, respectively, of the total radioactivity.When fat body was pulse-labeled with ³²P we found a heavily-phosphorylated intracellular 215 kD polypeptide which was precipitable with anti-vitellogenin. The synthesis of this intracellular precursorform of vitellogenin (pre-Vg) was under absolute juvenile hormone control. In vitro³²P pulse-chase experiments showed that pre-Vg was proteolytically processed within the fat body into some (or possibly all) of the mature vitellogenin subnits. Furthermore, peptide mapping confirmed that all of the phosphorylated vitellogenin subunits were derived from pre-Vg. Since previous studies have shown that phosphoserine residues account for essentially all of the covalently-attached phosphorus of the native vitellogenin molecule, we speculate that the asymmetric pattern of vitellogenin and vitellin subunit-phosphorylation is due to an uneven distribution of phosphoserine residues along the initial pre-Vg polypeptide chain. Finally, we conclude that phosphorylation of vitellogenin occurred post-translationally in the fat body endoplasmic reticulum because we could identify ³²P-labeled pre-Vg in purified microsomal vesicles but not in nascent vitellogenin polypeptide chains attached to vitellogenin polyribosomes.     

Polypeptide composition and biosynthesis of the yolk proteins in the firebrat,Thermobia domestica (insecta,thysanura)

Ion-exchange chromatography of crude ovarian extracts of the primitive insect Thermobia domestica allowed the separation, in native conditions, of major and minor vitellins of molecular weights of 300,000 and 430,000, respectively. Their polypeptide subunits were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunotransfer using an antiserum prepared against major vitellin. This protein was resolved into large (M_r 166,000–212,000) and small (around M_r 50,000) polypeptides. Minor vitellin, on the other hand, exclusively contained small polypeptides that are immunologically different from those of the major vitellin. Vitellogenin polypeptides from the hemolymph of mature females exhibited electrophoretic mobilities and immunological properties similar to vitellin polypeptides. Pulse-chase experiments showed that the female fat body synthesizes radioactive and immunoprecipitable proteins, whose polypeptide pattern is close to that of the major vitellogenin. However, part of the primary vitellogenic polypeptides, at M_r 210,000 and 212,000, is rapidly processed to M_r 176,000 and 182,000 subunits. These two polypeptides, as well as the precursors, enter into the composition of the major hemolymph vitellogenin. Finally, processing of the still uncleaved 210,000–212,000 polypeptides takes place in the ovary, which performs the same step of vitellogenin maturation as the fat body.     

Relationship between feeding,mating, vitellogenin production and vitellogenesis in the tickDermacentor variabilis

Anti-vitellin IgG directed againstDermacentor variabilis egg vitellin was used in sodium dodecyl sulfate polyacrylamide (SDS-PAGE) gradient gel immunoblots to detect the presence of vitellin and its precursor, vitellogenin, in the organs of feeding adults and in the immature stages of this tick. Vitellin polypeptides were found in the egg, larvae, nymph, and in the unfed adult stages of both sexes. Vitellin polypeptides were first detected in the ovary of mated females during the rapid-engorgement feeding, period. These polypeptides were also present in the ovaries of ovipositing females, unmated females fed for extended periods, and fed unmated females that were detached from the host and held for 12 h before dissection. The same anti-vitellin antibody was used in immunoblots to monitor the appearance of vitellogenin in the organs and hemolymph of female ticks. Immunoreactive peptides of vitellogenin were found in the fat body, midgut, and hemolymph of pre-rapid-engorging mated and unmated females. These polypeptides were not found in fed males nor in Malpighian tubes of feeding or ovipositing females Our data supported the following conclusions: 1) presence of immunoreactive vitellogenin in the adult female fat body, hemolymph, and midgut was, dependent upon feeding; 2) in mated feeding females, we could not detect the uptake of vitellogenin by the ovary until rapid engorgement; 3) in unmated females, vitellogenesis did not, begin unless prolonged feeding occurred; and 4) during the early developmental stages of this tick, vitellin served as an embryonic nutrient reserve and as a reserve against starvation between feedings.     

Rhipicephalus sanguinius: localization of vitellogenin synthesis by immunological methods and electron microscopy

In ovipositing Rhipicephalus sanguinius (Latrelle), complete immunological identity existed between vitellogenin from the midgut, fat body, and hemolymph and vitellin from eggs. This supported the hypothesis that the same vitellogenin was synthesized by both the midgut and fat body, then released into the hemolymph and transported to the ovary. Vitellogenin was taken up unaltered by the oocytes during vitellogenesis to become vitellin. Antivitellogenin did not react with host (dog) hemoglobin. Transmission electron microscopy showed specialized cells with large amounts of rough endoplasmic reticulum, Golgi complexes, and secretory granules in the midgut and fat body of ovipositing females that were absent in the midgut and fat body of fed males. It is suggested that these cells synthesize vitellogenin.     

Biosynthesis,purification, and characterization of Aedes aegypti vitellin and vitellogenin

Vitellin, the major egg yolk protein, and vitellogenin, the hemolymph precursor of egg yolk protein, have been purified to apparent homogeneity from the mosquito Aedes aegypti. The purification procedure included chromatography on ion exchange, hydrophobic, and gel filtration columns. Vitellin and vitellogenin have a similar molecular weight (M_r 300,000) on gel filtration columns. However, the molecular weights of vitellin and vitellogenin, as determined from SDS electrophoresis, were 393,000 and 337,000, respectively. Vitellin in sodium dodecyl sulfate released six subunits of molecular weight 116,000, 83,000, 75,000, 54,000, 36,000, and 29,000, whereas vitellogenin released only three subunits (155,000, 120,000, and 62,000). The average molecular weights of vitellin and vitellogenin after gel filtration and SDS electrophoresis were 346,000 and 318,000, respectively. Vitellin has a high content of aspartic acid and glutamic acid, and a low content of histidine, methionine, cysteine, and tryptophan. Vitellin also contains 0.9% mol of glucosamine and no galactosamine. The isoelectric points of vitellin and vitellogenin are at pH 6.4 and 6.3, respectively. Aedes aegypti fat bodies incubated for short intervals in tissue culture medium in the presence of [³H]valine showed incorporation by radio-immunoprecipitation and SDS electrophoresis into three primary vitellogenin polypeptides of molecular weights (± SEM) 156,000 ± 4,000, 114,000 ± 5,000, and 62,000 ± 400 inside the fat body and 162,000 ± 3,000, 118,200 ± 2,000, and 63,000 ± 300 in the medium. These results suggest that the molecular weight of vitellogenin synthesized inside the fat body (M_r 332,000) remains unchanged when secreted into the hemolymph (M_r 343,000). The three vitellogenin subunits are processed by the ovary into six subunits which are then deposited in the yolk granules as vitellin.     

Purification and partial characterization of vitellin from the eggs of the hard tick,Dermacentor variabilis

The major yolk proteins were purified from the eggs of the hard tick, Dermacentor variabilis using gel filtration and ion exchange chromatography. Two vitellin proteins were identified and designated vitellin A (480 kilodaltons; kDa) and vitellin B (370 kDa). The isolectric points were pH 6.1 and 6.25, respectively. The absorption maxima for both proteins were 280 and 400 nm. The buoyant density of vitellin A was 1.281 g/ml and vitellin B 1.278 g/ml. The vitellins were hemoglycolipoproteins as indicated by selective staining of polyacrylamide gels, carbohydrate analyses and lipid analyses. Under reducing conditions (SDS-PAGE), vitellin A had eight major polypeptides at 135, 110, 98, 80, 67, 50, 45, and 35 kDa. Vitellin B was identical to vitellin A with the addition of a 93 kDa subunit. The only carbohydrate detectable in the proteins was mannose. The neutral lipids detected in both proteins were cholesteryl esters, triglycerides, free fatty acids and their methyl esters, and cholestrol. The only detectable phospholipid in both proteins was phosphatidylethanolamine. The purified vitellins were immunologically identical to female hemolymph proteins but not to host hemoglobin. Antivitellin antibodies to vitellin were used to identify possible locations of vitellogenin in the organs of ovipositing females.     

龟纹瓢虫卵黄蛋白的分子特性及发生动态

研究了龟纹瓢虫Propylea japonica (Thunberg) 卵黄蛋白的基本特性以及卵黄发生过程中卵黄蛋白的动态变化。PAGE和SDS-PAGE实验表明,龟纹瓢虫卵黄蛋白分子量为294.81±40.70 kD,并由分子量分别为144.68±0.03 kD和51.23±0.27 kD的两种亚基组成。对卵黄蛋白的氨基酸组成和含量分析发现,其必需氨基酸总量占57.48％,略高于非必需氨基酸,其中谷氨酸（Glu）含量最高,为15.26％;色氨酸（Trp）和蛋氨酸（Met）含量较低,分别为0.50％和0.11％。采用间接竞争ELISA法,系统测定了龟纹瓢虫成虫期脂肪体、血淋巴和卵巢中卵黄蛋白的动态变化,结果表明：脂肪体是卵黄原蛋白合成的场所,卵黄原蛋白的合成始于羽化后第2天;脂肪体、血淋巴和卵巢中卵黄原蛋白的滴度在羽化后第4天开始迅速上升,至成虫期的第8天左右达到高峰期。     

Vitellogenin fluctuations in haemolymph and fat body and dynamics of uptake into oöcytes during the reproductive cycle of Diploptera punctata

Haemolymph and fat body soluble protein titres have been examined during the reproductive cycle of Diploptera punctata, with particular emphasis on the occurrence of vitellogenin and its uptake into the developing oöcytes. Vitellogenin was first detected in the haemolymph of mated females 2 days after adult eclosion at about the same time that vitellin deposition in basal oöcytes began. Peak haemolymph titres of vitellogenin occurred on day 6, correlated with the completion of yolk uptake. Thereafter vitellogenin levels declined and were generally undetectable throughout most of gestation, rising again shortly before parturition in association with the second gonotrophic cycle. Total haemolymph protein levels were not correlated with vitellogenesis.Soluble fat body vitellogenin titres of mated females remained low during the first oöcyte growth period but then rose several-fold at its completion and remained high throughout pregnancy and the second gonotrophic cycle. Total fat body soluble proteins decline after adult eclosion in association with oöcyte growth.Vitellin accumulation in basal oöcytes was related linearly to increase in volume until the onset of chorion formation. Thus no post-vitellogenic growth period was detected.     

Controlled accumulation of vitellin in Bombyx eggs

Summary

Implantation of female fat body and ovary discs into the young male pupae brought about vitellin accumulation in the mature eggs developed in male hosts. The amounts of vitellin increased according to the increasing amounts of implanted fat body, and the vitellin synthesis activity of female fat body in male hosts was similar to that found in female hosts. When implanted into male pupae, larval fat body having no ability to produce vitellogenin in situ could bring about vitellin accumulation in the eggs. No accumulation of vitellin was induced by implantation of male fat body.     

Fat body is the site of vitellogenin synthesis in the soft tick,Ornithodoros moubata

Summary Vitellogenin (Vg) synthesis in cultured tissues was analysed biochemically in a soft tick,Ornithodoros moubata. Nine tissue fractions dissected from reproductive females were incubated in vitro in a specially designed Ringer containing³⁵S-methionine. The synthesis of total protein and Vg was assayed by the radioactivity incorporated into precipitates with trichloroacetic acid and antivitellin (Vn)-serum, respectively. Fat body was the most active tissue in Vg synthesis, which comprised 46% of the Vg synthesis by all tissues and 42% of total protein synthesis by fat body. Protein synthesized by the fat body and precipitated with anti-Vn-serum was shown by electrophoresis and fluorography, to consist of six radioactive polypeptides corresponding to the components of Vg. Vg synthesized in cultured fat body was first accumulated in the tissue and secreted into the medium during incubation. Some tissues other than fat body showed low Vg synthesis (in each, less than 12% of total protein synthesis) which, however, may be due to contamination by fat body cells as seen with the scanning electron microscope (SEM). SEM also showed that fat body cells in the active stage of Vg synthesis expanded about 10-fold in length. Immunohistochemical analysis showed a very strong reaction with anti-Vn-IgG in the cytoplasm of fat body from reproductive females. Fat body from unfed females and other tissues including midgut, did not show any specific fluorescence. A positive reaction was obtained with developing oocytes. These results indicate that the fat body is the only site of Vg synthesis in this tick.Abbreviations Vg vitellogenin - Vn vitellin - SDS sodium dodecyl sulfate - PAGE polyacrylamide gel electrophoresis - SEM scanning electron microscopy - TCA trichloroacetic acid     

Protein synthesis and secretion by the locust fat body after ovariectomy

The synthesis and secretion of proteins, including vitellogenin, by the locust fat body were investigated in vivo and in vitro at various times after ovariectomy. The rate of overall protein synthesis and secretion by the fat body in vivo is significantly less in ovariectomized animals than in the controls two to three weeks after the operation, the difference in the rates of secretion of vitellin antibody-precipitable protein being even more pronounced and detectable earlier. The significantly greater amount of newly synthesized vitellogenin retained in the fat body of ovariectomized animals is insufficient to account for this difference. The secretion of newly synthesized protein by the fat bodies of ovariectomized locusts in vitro is significantly less than that of control fat bodies, the difference being particularly marked in the case of vitellogenin. The polysome populations of fat bodies of ovariectomized and control females are quantitatively similar and the amounts of total protein and vitellin antibody-precipitable protein synthesized by these polysomes in a cell-free system do not differ significantly.     

Multiple vitellogenins from the Haemaphysalis longicornis tick are crucial for ovarian development

Ovarian development and egg maturation are crucial processes for the success of reproduction in ticks. Three full-length cDNAs encoding the precursor of major yolk protein, vitellogenin, were obtained from cDNA libraries of the Haemaphysalis longicornis tick and designated as HlVg-1, HlVg-2 and HlVg-3. The HlVg mRNAs were found in fed females with major expression sites in the midgut, fat body and ovary. Native PAGE and Western blot demonstrated that HlVgs in the hemolymph, fat body and ovary of fed females consisted of four major polypeptides. RNAi results showed that HlVg dsRNA-injected ticks obtained lower body weight, egg weight and showed higher mortality of engorged females after blood sucking than control groups. Our results indicate that all HlVgs are essential for egg development and oviposition.     

Vitellogenin in locusts (Locusta migratoria): Translation of vitellogenin mRNA in Xenopus oocytes and analysis of the polypeptide products

Cytoplasmic poly(A)⁺-RNA, prepared from fat bodies of reproductively active locust females, directed the synthesis of two large polypeptides in Xenopus oocytes. Occasionally two smaller polypeptides (X₁ and X₂) were also detectable. The two larger polypeptides were immunologically and electrophoretically indistinguishable from the unprocessed Vitellogenin polypeptides (Vg₁ and Vg₂) of locust fat bodies. Peptide patterns generated from these two translation products by Staphylococcus aureus V8 protease digestion were identical to those of Vg₁ and Vg₂. RNA isolated from allatectomized female locusts treated with the juvenile hormone analog (ZR-515) was also able to direct the synthesis of vitellogenin in Xenopus oocytes, whereas RNA isolated from mature males or allatectomized females did not. The molecular weights of fat-body Vg₁ and Vg₂ were 235,000 and 225,000, respectively and the processed vitellogenin polypeptides were found to range from 126,000 to 52,000. Electrophoretic and Chromatographic analysis of [³⁵S]methionine-containing tryptic peptides of Vg₁ and Vg₂ showed two different tryptic peptide fingerprints. Distinctly different peptide patterns were also observed when Vg₁ and Vg₂ were partially digested with V8 protease or papain. However, tryptic peptide mapping and V8 protease limited digestion mapping of fat-body X₁ and X₂ revealed that these two polypeptides were derived from Vg₁ and Vg₂. This suggests that Vg₁ and Vg₂ are products of two different vitellogenin structural genes.     

Feedback regulation of vitellogenin synthesis in Aedes aegypti and Aedes atropalpus

Blood-fed hemiovariectomized Aedes aegypti synthesized vitellogenin at the same rate as intact females. Removal of both developing ovaries from blood-fed females diminished vitellogenin synthesis gradually. To determine the concentration of circulating vitellogenin, vitellin was purified using hydrophobic affinity chromatography. A titration curve of antibody-antigen complex was developed and the concentration of vitellogenin circulating in ovariectomized females was found to be five-fold higher than that in intact females.Ovaries removed from vitellogenic donors did not induce vitellogenin synthesis in non-vitellogenic (blood-fed, decapitated) recipients. Implanting a pinocytotic ovary into blood-fed females in which vitellogenesis was arrested by ovariectomy, restored vitellogenin synthesis.These results indicate that vitellogenin synthesis in Aedes aegypti is controlled by a feedback-inhibition mechanism, rather than an ovarian factor, and the ovary acts as a sink.     

Vitellogenin synthesis in the lady beetle Coccinella septempunctata

To elucidate the hormonal mechanisms which regulate reproduction in a beneficial insect, we have begun to investigate the process of vitellogenesis in Coccinella septempunctata, the seven-spotted lady beetle. Vitellin (Vn) constitutes 60–70% of the total protein in C. septempunctata eggs and is composed of four polypeptides with molecular weights determined by electrophoresis in denaturing gels of 133,000, 130,000, 46,000 and 43,000. In the egg these polypeptides occur in a ratio of approx. 1:1:1:2. The two larger Vn polypeptides yielded similar peptide fragments upon partial proteolytic digestion which suggests that they are structurally related. The two smaller Vn polypeptides appear structurally distinct because they yielded unique proteolytic fragments. The Vn precursor, vitellogenin (Vg), was observed in the haemolymph of mature females, but was not detected in the haemolymph of immature females or males. The electrophoretic mobilities, antigenicity, and proteolytic fragmentation patterns of the Vg polypeptides were indistinguishable from those of their Vn counterparts. Thus the major processing of the Vn polypeptides appears to occur prior to their secretion into the haemolymph.The synthesis of Vg was examined in whole animals and in organ cultures. Vg synthesis was observed in the fat body and to a smaller extent in the ovaries of mature females. The newly synthesized Vg was rapidly secreted. Vg synthesis was not detectable in brain or thoracic muscle of mature females or in the fat body of males or immature females. Very little vitellogenin synthesis occurred in female insects reared on artificial diets. The topical application of a juvenile hormone analogue induced Vg synthesis in non-fecund females but not in males.     

Onset of vitellogenin production and vitellogenesis,and their relationship to changes in the midgut epithelium and oocytes in the tickDermacentor variabilis

InDermacentor variabilis (Say), the onset of vitellogenin production and vitellogenesis (up-take of vitellogenin into oocytes) began during the rapid-engorgement feeding period. Mating was required for both vitellogenin production and vitellogenesis to complete the tick's life cycle. Complete immunological identity, as measured by Ouchterlony's double diffusion test, existed between vitellogenin from the fat body, midgut and hemolymph, and vitellin from the ovaries and eggs. Antivitellin antibody did not react with host hemoglobin nor with fat body, midgut, and ovary extracts from feeding females prior to rapid engorgement, feeding unmated females, or unfed or fed males. Some unmated females fed for 13 days and then hand-detached from the host eventually began oviposition after going through a preoviposition period. In these ticks, organ extracts from the midgut, fat body and ovary reacted with antivitellin antibody. The presence or absence of presumed vitellogenic cells in the midgut and yolk bodies in oocytes corresponded with the presence or absence of vitellogenin and vitellogenesis as measured by Ouchterlony's test. Presumed vitellogenic cells increased in size during the preoviposition period. These cells reached their greatest size during the time when the most eggs were being produced, and then declined in size toward the end of oviposition. Vitellogenin was deposited directly into developing yolk bodies in oocytes and was not processed through lysosomes. Feeding was the process that initiated the formation of eggshell cuticle. Detachment from the host was required for the initiation of oviposition.     

Vitellin and vitellogenin concentrations during oögenesis in the first gonotrophic cycle of the house fly,Musca domestica

The house fly, Musca domestica, contains at least two native vitellin and two vitellogenin proteins. Both vitellins appear to have an identical vitellogenin partner. The major native vitellin has a mol. wt of 281 K Daltons, and the major native vitellogenin has a mol. wt of 283 K Daltons. These proteins are composed of three subunits with mol. wt of 48, 45 and 40 K Daltons. The relationship of the subunits to the native proteins is not known.Haemolymph vitellogenin levels are cyclical during oögenesis with no detectable amounts in previtellogenic flies and low levels in postvitellogenic flies. The highest level of vitellogenin, 10.5 μg/μl, occurred in flies with stage-7 ovaries. The vitellogenin levels during oögenesis fit a parabolic curve and the fat body vitellogenin content during oögenesis showed this same pattern.Uptake of vitellogenin into the ovary during each stage of oögenesis also fit a parabolic curve and produced a high linear correlation with haemolymph vitellogenin levels. The greatest uptake was 37 μg/stage and occurred during stage 6.     

Role of juvenile hormone in the synthesis and sequestration of vitellogenins in the red cotton stainer, Dysdercus koenigii (Heteroptera: Pyrrhocoridae)

Investigations were carried out to determine the role of juvenile hormone (JH) and 20-hydroxy ecdysone in the synthesis and uptake of vitellogenins, which were earlier identified, purified and characterised, in Dysdercus koenigii. The concentration(s) of vitellogenin(s) in fat body, haemolymph and that of vitellin(s) in ovary were significantly lower after chemical allatectomy at eclosion. In addition, at 70 h after emergence, chemical allatectomy reduced ovarian vitellin concentration, but vitellogenin levels remained normal in the fat body and haemolymph. The haemolymph vitellogenins were not incorporated into oocytes in such insects. Administration of JH-III at 20 h after allatectomy restored vitellogenin levels in the fat body and haemolymph, but the ovary failed to incorporate the available vitellogenins from haemolymph in such insects. However, when JH-III was administered twice, one at 20 h and then at 70 h after allatectomy, vitellogenin concentrations in fat body and haemolymph and also vitellin concentrations in ovary approached control levels. It is suggested that JH has two separate roles, one in vitellogenin synthesis and the other in uptake. 20-hydroxy ecdysone had no apparent role in either vitellogenin synthesis or uptake in D. koenigii.     

The relationship between nutrition,vitellogenin, vitellin and ovarian development in the housefly, Musca domestica L.

The effect of adult nutrition on oögenesis during the first gonotropic cycle was studied in three strains of the housefly, Musca domestica. Two of the strains were anautogenous and the third was autogenous. In these strains, three subunits (51, 43 and 42 kdaltons) of vitellogenin and vitellin were electrophoretically identical using SDS-PAGE electrophoresis for haemolymph proteins of vitellogenic females and for egg extracts. Each developmental stage of the ovary in individual females flies of both autogenous and anautogenous strains fed on either sugar or protein clearly reflected the appearance of electrophoretic bands for vitellogenin and vitellin. Using immunological analysis, a very small amount of vitellogenin was detectable in the haemolymph of previtellogenic flies. The highest level of vitellogenin appeared in the haemolymph at the middle of vitellogenic phase and reached about 25% of the total haemolymph protein. There were differences in vitellogenin concentration in females with mature eggs between the two anautogenous strains: vitellogenin was not detectable in one strain, and the other showed 30% of the maximal level.     

Vitellogenin synthesis,processing and hormonal regulation in the tick,Ornithodoros parkeri (Acari:Argasidae)

This study was undertaken to determine the processing of vitellogenin (Vg) and the role of juvenile hormone (JH) in the regulation of vitellogenesis in the tick Ornithodoros parkeri. Ticks usually require a blood meal to induce vitellogenesis. However, we have shown that a pyrethroid, cypermethrin (CyM), can stimulate Vg synthesis in unfed Ornithodoros moubata females. Vg concentration and synthesis were analyzed by SDS-PAGE spotting-scanning and fluorography using [³⁵S]-methionine. Although unfed females show high titers of Vg in the hemolymph, this is not due to new synthesis. Vg synthesis stimulated by engorgement increases beginning on day 2 after engorgement and reaches a maximum level on day 8. Vg is synthesized in the fat body, secreted into the hemolymph and then processed and incorporated into the ovaries as vitellin. JH I, II and III, methoprene (JHA), and CyM were topically applied to unfed females and Vg synthesis analyzed on day 5 by fluorography. JH and JHA did not stimulate Vg synthesis. CyM stimulated Vg synthesis but not ovarian development. These preliminary results indicate that JH does not function in the regulation of vitellogenin synthesis in this species.