Induction Of Vitellogenin Synthesis Research Articles

Estrone and estradiol participation during exogenous vitellogenesis in the female rainbow trout, Salmo gairdneri

Ovarian steroids were treated for their ability to induce vitellogenin synthesis in the liver of the rainbow trout, Salmo gairdneri. These steroids were chosen because they had been reported to induce vitellogenin in the plasma of teleosts, i.e., estrone, estradiol, and testosterone, or because they were synthesized during vitellogenesis, as was the case with the above steroids and androstenedione and 17α-hydroxyprogesterone. Immature trout were ovariectomized and injected daily for 7 days. The administered doses of steroid were 100, 250, and 500 ng/g body wt. Only estradiol and estrone induced vitellogenin in the plasma; estrone had about 5% of the potency of estradiol. A dose-effect curve was determined for estradiol in the range from 25 to 1000 ng/g body wt. A maximal amount of 7 mg vitellogenin/ml plasma was found following the administration of a dose of 250 ng/g estradiol. Vitellogenin was not present in the plasma of animals treated with saline, nor could it be detected in the liver, not even after the administration of estradiol. Estradiol administration increased the total plasma protein concentration from 35 to maximally 51 mg/ml and decreased the total piver protein concentration from 163 to minimally 118 mg/g. The relative weight of the liver increased from 12.9% to maximally 22.4%. Vitellogenin was not detected in the liver of any of the experimental animals, indicating a low storage and rapid secretion of vitellogenin. The other steroids influenced some of the variables, but never was the total pattern of effect comparable to that of estradiol. Estradiol is found to be the ovarian steroid that physiologically regulates the synthesis of vitellogenin in the liver of the rainbow trout; estrone is less active. Experiments undertaken to determine the effects of the combined presence of estrone and estradiol revealed that estrone was capable of boosting the vitellogenic effect of estradiol when compared to the induced vitellogenin levels following treatment with a combination of testosterone and estradiol. The vitellogenin concentrations induced by a certain dose of one of the combinations of estrone and estradiol approximated the concentrations induced by the same dose of estradiol alone. This effect was independent of the ratio in which estrone and estradiol were administered. These findings could not be explained by conversions of estrone into estradiol or by the vitellogenic activity of estrone alone. The estrone, estradiol, and vitellogenin concentrations in the plasma of the experimental animals were in the same range as determined previously in untreated mature vitellogenic females. Vitellogenin and estradiol levels were found to correlate in experimental animals treated with estradiol ( r = 0.627; N = 20). This was not the case in animals treated with combinations of estrone and estradiol. In these animals, however, the sum of estrone and estradiol levels in the plasma correlated with vitellogenin levels ( r = 0.724; N = 42). Vitellogenin was not detected in the liver of any of the experimental animals, which indicates a low storage and rapid secretion of vitellogenin. The importance of viewing the sum of estrone and estradiol plasma levels in connection to physiological studies of the regulation of exogenous vitellogenesis in the rainbow trout is discussed.

Vitellogenin synthesis induced in locust fat body by juvenile hormone analog in vitro

Fat bodies from adult females ofLocusta migratoria continue to synthesize vitellogenin and other proteins when cultured in vitro. A strong secondary induction of vitellogenin synthesis was obtained in fat bodies cultured in the presence of methoprene, and a weaker but significant primary induction was also obtained using higher doses (>100 μg) of methoprene.

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.

The role of thyroxine in the transition of vitellogenin synthesis from noninducibility to inducibility during metamorphosis in Xenopus laevis

Vitellogenin synthesis can be induced in vivo by oestrogen in the livers of Xenopus laevis larvae during metamorphic climax. The timing of the appearance of vitellogenin inducibility has been precisely defined in relation to changing morphological characteristics, and occurs during normal development at stage 62. This suggests that thyroid hormones may be involved in the transition from the uninducible to the inducible state of the vitellogenin gene. Chemical inhibition of the larval thyroid by propyl thiouracil or thiourea was used to produce developmentally static larvae. Tests on these animals for response to oestrogen showed that the transition depends on metamorphic stage, not on chronological age. Immersion of normal tadpoles or developmentally static tadpoles in high concentrations of thyroxine induces rapid metamorphosis, but does not alter the developmental stage at which vitellogenin inducibility is acquired in either case. Induction of vitellogenin synthesis in animals before stage 62 was only achieved by permitting thyrostatic animals to continue a delayed metamorphosis. It is apparent that thyroxine does not directly confer inducibility on the vitellogenin gene.

Ethanol and liver protein synthesis in vivo

Ethanol was administered i.p. to adult roosters during hormonally induced vitellogenin synthesis. At moderate doses, ethanol had no influence on the synthesis of vitellogenin nor did it cause alterations in the size distribution of liver polyribosomes.

Estrogen-induced ultrastructural changes in cultured chick embryo hepatocytes

Administration of estradiol-17-beta induces the synthesis of vitellogenin in primary cultures of chick embryo liver. The ultrastructural changes accompanying steroid induced vitellogenin synthesis were investigated in hepatocytes cultures incubated for 0, 12 and 24 h following hormonal treatment. Both electron microscopy and immuno-chemical techniques were used. The immuno-fluorescence data indicate that the cultures contain a cell population of 90--95% hepatocytes, both in control and estradiol-treated groups. Ultrastructurally, cultured hepatocytes are similar to in vivo ones, except for a reduced accumulation of storage materials, glycogen and lipids. In the estradiol-treated cultures hepatocytes show an accumulation of free ribosomes, enlargement of rough endoplasmic reticulum, and development of the Golgi apparatus.

Induction of vitellogenin synthesis in goldfish by massive doses of androgens

Oral administration of massive doses of methyltestosterone into goldfish caused an extensive proliferation of the rough endoplasmic reticulum, hypertrophy of the Golgi apparatus, and the production of numerous secretory granules in the liver, suggesting the induced synthesis of some secretory proteins. The electrophoretic pattern of serum proteins was prominently altered, showing three bands which were absent in normal, immature fish. All of these changes were similarly observed in estradiol-treated fish. In order to identify the hormone-induced serum proteins, they were purified from estradiol- and methyltestosterone-treated fish and were subjected to chemical and immunological analyses. The results indicated that the serum proteins induced by methyltestosterone and by estradiol are identical in the contents of lipids, phospholipids, calcium, and protein-bound phosphorus and the reactivity to antiserum elicited to the estradiol-induced serum proteins. Furthermore, it was found that the antibody forms a single, connecting precipitin line with the hormone-induced proteins, sera of vitellogenic fish, and ovary extracts of matured fish, but not with sera of normal males. In view of the above findings, it was concluded that the serum proteins induced by methyltestosterone are vitellogenin. Ethynyltestosterone and methylandrostenediol were also effective, but testosterone, dihydrotestosterone, and methyldihydrotestosterone were much less effective in inducing vitellogenin synthesis.

Thyroid hormone induces competence for oestrogen-dependent vitellogenin synthesis in developing Xenopus laevis liver.

OESTROGEN-DEPENDENT synthesis of vitellogenin, the precursor of the major yolk proteins, in the liver of egg-laying vertebrates is an attractive model for investigating regulatory mechanisms of gene expression. Induction of vitellogenin synthesis by oestrogen occurs not only in the liver of females but also in males which normally do not produce this protein, thus showing that responsiveness to oestrogen is not linked to sex (for review see ref. 1). However, Follet and Redshaw2 reported that liver of Xenopus laevis larvae is refractory to oestrogen, becoming responsive shortly after completion of metamorphosis. As amphibian metamorphosis is known to be obligatorily dependent on thyroid hormones (for review see ref. 3), the question arises as to whether acquisition of competence to respond to oestrogen by the Xenopus liver cells is part of the hormonally regulated metamorphic changes or merely reflects age-dependent maturation. Here we present evidence that during normal development of Xenopus laevis the liver cells become responsive to oestrogen during metamorphosis and before vitellogenin synthesis has been initiated by endogeneous oestrogens. Moreover, we demonstrate that responsiveness to oestrogen fails to appear when metamorphosis is suppressed, but readily appears after induction of metamorphosis by thyroxine, suggesting that acquisition of competence to respond to oestrogen by the Xenopus liver cells is controlled by thyroid hormones.

Induction of Vitellogenin Synthesis in Xenopus laevis Tadpoles

Oestradiol induces vitellogenin synthesis in vitro in liver taken from Xenopus laevis tadpoles that are in late metamorphosis. Inducibility first appears at the end of prometamorphosis, and the response to oestradiol increases during the completion of metamorphosis. Oestradiol continuously present during development does not influence the stage at which tadpole liver becomes inducible. It seems that the acquisition of inducibility is part of the normal development of the liver, and independent of both the supply of oestrogen and the sex of the tadpole.

An estrogen receptor from xenopus laevis liver possibly connected with vitellogenin synthesis

This paper describes an estrogen receptor which is found in both the nucleus and cytoplasm of liver cells from male Xenopus laevis, and which seems to be involved in the induction of vitellogenin synthesis. It has a high affinity for estradiol (K d = 0.5 × 10 −9 M), and the affinities of various steroids for the receptor correlate well with their ability to induce vitellogenin synthesis. It sediments at 3.5S at 0°C in 0.5 M KCI. The rate of sedimentation is unaffected by incubation at 20°C prior to centrifugation, but increases if the salt concentration is lowered to 0.1 M KCI or to zero. It has a Stokes radius of 2.6 nm and a molecular weight of approximately 40, 000. The receptor is present at very low levels compared to other steroid target tissues (50–100 fold less than chick oviduct). The cytoplasm of a single hepatocyte contains 92 ± 18 binding sites for estradiol, while each nucleus contains 99 ± 19 sites.

Coordination of ribosome content and polysome formation during estradiol stimulation of vitellogenin synthesis in immature male chick livers.

To elucidate the mechanisms by which protein synthesis is affected by estradiol, we characterized cockerel liver polysomal profiles during hormone induction and withdrawal. We describe a method for isolating intact polysomes which results in preparations that are stable even after storage in solution at 10 degrees for 16 hr. In addition, our procedure eliminates the necessity for starving animals prior to experiments. Recovery of radioactive polysomes indicated that yield is about 90% and that our polysomal preparations appear to represent polysome distribution in vivo. Using this approach we show that estradiol injection stimulates ribosome content 6-fold and that formation of polysomes is coincident with the induction of vitellogenin synthesis. We also demonstrate that the size and number of polysomes increase and decrease in a coordinated fashion with the rate of vitellogenin synthesis. The kinetics of ribosome synthesis and the fact that at least 80% of the newly synthesized ribosomes are directly recruited into polysomes indicate that ribosomes might be limiting the rate of protein synthesis during the stimulatory phase of the hormone cycle.

Genetic and Hormonal Regulation of Vitellogenesis inDrosophila

In Drosophila the female-specific yolk protein, or vitellogenin, is synthesized in the fat body. In D. melanogaster , vitellogenin, is first detected in the female hemolymph at the time of adult eclosion and in the ovaries 20 hours later, suggesting different regulatory mechanisms for the processes of synthesis and uptake. Transplantations of pupal or immature adult ovaries into D. melanogaster adult males induce vitellogenin synthesis, implicating an ovarian agent in the control of synthesis. Larval ovaries fail to stimulate synthesis. Female-sterile mutants with rudimentary or previtellogenic ovaries synthesize and accumulate large quantities of vitellogenin in the hemolymph, but not in the ovaries. Transplantation of these rudimentary ovaries into males induces vitellogenin synthesis, suggesting that the ovarian inducing agent does not originate from the germ cells. Treatment of the homozygous female-sterile mutants with juvenile hormone stimulates the uptake of vitellogenin by the ovary in some strains. This shows that juvenile hormone plays a role in vitellogenin uptake. The potential importance of Drosophila vitellogenesis for studies of gene regulation is discussed.

Induction of vitellogenin synthesis in male Sarcophaga bullata by ecdysterone

Vitellogenin synthesis can be induced very quickly in adult male Sarcophaga bullata starting from 3 days after emergence by injection of a dose ≥50 nanogram of ecdysterone. Vitellogenin has been identified by electrophoresis and immunologic methods. Implantation of ovaries in males does not induce vitellogenin synthesis. Normal vitellogenesis cannot be obtained by implantation of ovaries combined with injection of ecdysterone. Apparently, ecdysterone derepresses in some way the vitellogenin gene or m-RNA in males.

Direct induction by estradiol of vitellogenin synthesis in organ cultures of male xenopus laevis liver

Organ cultures of liver from untreated male Xenopus respond to 17β-estradiol in the culture medium by synthesizing and secreting the yolk protein precursor vitellogenin. Vitellogenin synthesis, as a primary response, is first detectable on the fourth day of culture, and comprises up to 12% of the protein synthesized on the eight day. Estradiol is required during the lag period of 4 days. Tissue from male Xenopus injected with 1 mg of estradiol 30 days before the start of culture responds more rapidly and to a greater extent to estradiol in the medium than tissue from uninjected males. During such a secondary response, vitellogenin is first detectable in the medium on the second day of culture, and becomes up to 24% of the protein synthesized on day 6. The rate of amino acid incorporation into total protein also increases in response to estradiol, but the rate of synthesis of albumin decreases rapidly in culture whether or not estradiol is present, in both the primary and secondary responses. A maximal response is seen with 10 −8 M estradiol. Progesterone, testosterone, dexamethasone, and insulin neither induce vitellogenin synthesis in culture nor modify the response to estradiol. DNA synthesis inhibitors do not prevent the response to estradiol in vitro, suggesting that cell division is not required for the initial response leading to vitellogenin synthesis.

Synthesis of vitellogenin in cultures of male and female frog liver regulated by estradiol treatment in vitro.

Using the frog Xenopus laevis, we show that the addition of physiological concentrations of estradiol to cultures of liver from untreated males rapidly induces the synthesis of large amounts of vitellogenin. Sustained synthesis of vitellogenin requires continuous exposure to estradiol. A nonestrogenic steroid, dexamethasone, does not induce vitellogenin synthesis but does induce increased synthesis of a different protein in liver cultures.

Activation of vitellogenin synthesis in the mosquito Aedes aegypti by ecdysone

Injected β-ecdysone was found to induce the synthesis of yolk protein (vitellogenin) in adult female Aedes aegypti without a blood meal. After injection of 5 μg ecdysone per mosquito, vitellogenin constituted 80 per cent of the total protein secreted by explanted fat body, a proportion comparable to that produced by fat body from blood-fed females. Moreover, the time course of induction of vitellogenin synthesis in ecdysone-injected mosquitoes was similar to that triggered by a blood meal. Response to ecdysone is dosedependent: 0·5 μg per female was required to stimulate synthesis to 50 per cent of the level found 18 hr after a blood meal. Ecdysone was effective in decapitated or ovariectomized mosquitoes, and also when applied directly to fat body preparations in vitro. Thus it appears that ecdysone acts directly on the fat body to induce specific protein synthesis, as does the vitellogenin stimulating hormone (VSH) from the ovary of blood-fed mosquitoes. These results suggest that ecdysone can replace VSH in inducing vitellogenin synthesis in the unfed mosquito.

Influence of a juvenile hormone analogue on vitellogenin synthesis and oögenesis in larvae of Nauphoeta cinerea

In experiments on the synthesis of the vitellogenic protein, farnesylmethylester, a juvenile hormone (JH) analogue, was injected into female Nauphoeta cinerea larvae at various stages during their development. Two and 4 days after injection, 2 μl of haemolymph were assayed in a vitellogenin immunodiffusion test. In second last and last instar larvae less than 6 days before adult ecdysis, high doses (100 μg) of farnesylmethylester are necessary to induce vitellogenin synthesis, whereas older last stage larvae and decapitated adults respond to small doses (1 μg) with the synthesis of vitellogenin. It seems that the competence to synthesize the vitellogenic protein changes at the time of induction of the moulting process. If farnesylmethylester is injected into last instar larvae with a supposedly high titre of ecdysone, the vitellogenic protein can be detected in the haemolymph of a small percentage of animals only. Oöcyte maturation can be observed in last instar larvae injected after the fifth to ninth day with farnesylmethylester. The observed volume changes of the corpora allata suggest that an absence of JH for a short time is necessary for the oöcytes to become competent to grow. Last instar larvae treated with farnesylmethylester become larval-adult intermediates with partly developed oöcytes, demonstrating a simultaneous juvenilizing and gonadotropic influence of the JH analogue. In last instar larvae injected with farnesylmethylester a partial degeneration of already maturing oöcytes is induced at the time when the ecdysone titre is supposedly high and the possible reasons for this are discussed.

Juvenile hormone induced vitellogenin synthesis in the monarch butterfly.

The vitellogenin (yolk protein) of the monarch butterfly has been identified by electrophoretic and immunochemical techniques. In freshly emerged female adults ligature of the neck prevents appearance of this protein in the hemolymph; it prevents yolk deposition and egg maturation as well. These processes are restored by injection of juvenile hormone; the restoration involves induction of vitellogenin synthesis, as shown by incorporation of [(3)H]leucine.