Comparative Studies of Ecdysteroid Metabolism between Diapause Eggs and Non-diapause Eggs of the Silkworm, Bombyx mori

Sonobe, Haruyuki; Tokushige, Hideki; Makka, Toshihiro; Tsutsumi, Hibiki; Hara, Noriyuki; Fujimoto, Yoshinori

doi:10.2108/zsj.16.935

Cited by 23 publications

(24 citation statements)

References 26 publications

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“…Indeed, it is only after 6 h that the 20E-inducible cascade of early transcription factor genes E75A&B, DHR3, DHR4, DHR39, and ␤FTZ-F1 becomes apparent (April A. Sullivan and Carl S. Thummel, personal communication). Additional support comes from recent findings that E20MO activity, both in vivo and in vitro, increases dramatically after gastrulation in developing nondiapause eggs of the silk moth, Bombyx mori, relative to the very low activity measured in diapausedestined eggs, which cease further embryonic development at this time (44,45).…”

Section: Discussionmentioning

confidence: 89%

Shade is the Drosophila P450 enzyme that mediates the hydroxylation of ecdysone to the steroid insect molting hormone 20-hydroxyecdysone

Petryk

Warren

Marqués

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

410

373

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The steroid 20-hydroxyecdysone (20E) is the primary regulatory hormone that mediates developmental transitions in insects and other arthropods. 20E is produced from ecdysone (E) by the action of a P450 monooxygenase that hydroxylates E at carbon 20. The gene coding for this key enzyme of ecdysteroidogenesis has not been identified definitively in any insect. We show here that the Drosophila E-20-monooxygenase (E20MO) is the product of the shade (shd) locus (cytochrome p450, CYP314a1). When shd is transfected into Drosophila S2 cells, extensive conversion of E to 20E is observed, whereas in sorted homozygous shd embryos, no E20MO activity is apparent either in vivo or in vitro. Mutations in shd lead to severe disruptions in late embryonic morphogenesis and exhibit phenotypes identical to those seen in disembodied (dib) and shadow (sad) mutants, two other genes of the Halloween class that code for P450 enzymes that catalyze the final two steps in the synthesis of E from 2,22-dideoxyecdysone. Unlike dib and sad, shd is not expressed in the ring gland but is expressed in peripheral tissues such as the epidermis, midgut, Malpighian tubules, and fat body, i.e., tissues known to be major sites of E20MO activity in a variety of insects. However, the tissue in which shd is expressed does not appear to be important for developmental function because misexpression of shd in the embryonic mesoderm instead of the epidermis, the normal embryonic tissue in which shd is expressed, rescues embryonic lethality.

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Section: Discussionmentioning

confidence: 89%

Shade is the Drosophila P450 enzyme that mediates the hydroxylation of ecdysone to the steroid insect molting hormone 20-hydroxyecdysone

Petryk

Warren

Marqués

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

410

373

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“…In the diapause eggs, the bulk of the ecdysteroids are conjugated forms (phosphoric esters), but in the non-diapause eggs free forms co-exist with the conjugated forms (Ohnishi et al, 1971(Ohnishi et al, 1977Mizuno et al, 1981;Mamiya et al, 1995;Sonobe et al, 1997). In the course of quantitative analysis of egg ecdysteroids in the silkworm, we found that the 20-hydroxyecdysone titer, which is thought to be a physiologically active molecule in the silkworm egg (Gharib et al, 1981a), in-creased sharply in the non-diapause eggs as embryonic development proceeded, but remained at a low level in the diapause eggs .…”

Section: Introductionmentioning

confidence: 99%

Ecdysone 20-monooxygenase in eggs of the silkworm,Bombyx mori: Enzymatic properties and developmental changes

Horike

Sonobe

1999

Arch. Insect Biochem. Physiol.

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Ecdysone 20‐monooxygenase in eggs of the silkworm Bombyx mori was characterized in relation to embryonic development. First, subcellular fractions were prepared by means of differential centrifugation, and analyzed using marker enzymes and antibodies against NADPH‐cytochrome P450 reductase. It was demonstrated that most ecdysone 20‐monooxygenase activity was associated with microsomes, and that there was little or no intrinsic mitochondrial ecdysone 20‐monooxygenase. Next, conditions for the measurement of ecdysone 20‐monooxygenase activity were established for the microsomal fraction, and changes in the enzyme activity were measured in diapause eggs and non‐diapause eggs during early embryogenesis. It was demonstrated that enzyme activity in diapause eggs remained at a low level, while that in the non‐diapause eggs increased from the gastrula stage. The increase in egg ecdysone 20‐monooxygenase activity was prevented by actinomycin D and α‐amanitin, suggesting that gene transcription is required for eliciting an increase in ecdysone 20‐monooxygenase activity. Arch. Insect Biochem. Physiol. 41:9–17, 1999. © 1999 Wiley‐Liss, Inc.

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“…In B. mori (21,49), Lepidosaphes ulmi (50), and Locusta migratoria (51), 20-hydroxyecdysone plays an indispensable role in the developmental difference between diapause eggs and nondiapause eggs. In B. mori eggs, 20-hydroxyecdysone has been demonstrated to originate from both de novo biosynthesis (23,38,52,53) and dephosphorylation of ovary-derived ecdysteroid phosphates (27)(28)(29)53). Furthermore, in L. migratoria (54), S. gregaria (55), Blaberus craniifer (56), and Manduca sexta (14,15), free ecdysteroids liberated from ovary-derived ecdysteroid phosphates are suggested to participate in controlling the early events of embryonic development, such as embryonic cuticulogenesis and germ band elongation, which occur before the prothoracic glands differentiate during embryonic development.…”

Section: Discussionmentioning

confidence: 99%

“…Chemicals- [23, H]Ecdysone (2,112 GBq/mmol) was obtained from PerkinElmer Life Sciences. Ecdysone and 20-hydroxyecdysone were purchased from Sigma.…”

Section: Methodsmentioning

confidence: 99%

“…The ovaries of most insect species have the capacity to accumulate ecdysteroid phosphates, which are physiologically inactive (21), in addition to the capacity to synthesize free ecdysteroids de novo (22,23). It has been suggested that the ecdystreoid phosphates that are accumulated in the ovaries are transferred to the eggs and function as a source of active free ecdysteroid before the prothoracic glands differentiate during embryonic development (24 -26).…”

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confidence: 99%

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Purification, Kinetic Characterization, and Molecular Cloning of a Novel Enzyme, Ecdysteroid 22-Kinase

Sonobe

Ohira

Ieki

et al. 2006

Journal of Biological Chemistry

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This is the first report succeeding in the isolation and characterization of an enzyme and its gene involved in the phosphorylation of a steroid hormone. It has been demonstrated that ecdysteroid 22-phosphates in insect ovaries, which are physiologically inactive, serve as a "reservoir" that supplies active free ecdysteroids during early embryonic development and that their dephosphorylation is catalyzed by a specific enzyme, ecdysteroid-phosphate phosphatase (Yamada, R., and Sonobe, H. In steroid metabolism, it is generally accepted that cytochrome P450-mediated reaction, referred to as phase I metabolism, converts hydrophobic compounds into more polar metabolites, whereas phase II metabolism involves an adduct formation via a conjugation reaction. Phase II metabolism, such as sulfation, glucuronidation, and fatty acyl esterification, leads to a change of polarity or to a charge modification of the given metabolite, thereby modulating biological activity or facilitating elimination. In mammals, the sulfation pathway can be thought of as a reversible process, comprising two enzyme systems: the sulfotransferases, which catalyze the sulfation reaction, and the sulfatases, which catalyze the hydrolysis of sulfate esters formed by the action of sulfotransferases. Much is now known concerning the function of these enzyme systems as well as the molecular structure of the relevant cDNA and genes comprising these systems (1-4). However, to our knowledge, there exists very little information regarding the phosphorylation of steroid hormones in mammals.Ecdysteroids, which include the arthropod molting hormone, mediate a wide variety of developmental and reproductive events in insects (5). During the larval and pupal stages, ecdysteroids are synthesized in steroidogenic glands, known as the prothoracic glands (6). The detailed ecdysteroid biosynthetic pathway constituted by phase I metabolism is not fully clarified, but cytochrome P450 enzymes catalyzing the last four steps of the biosynthesis of 20-hydroxyecdysone (the principal molting hormone of arthropods) (i.e. CYP306a1 (25-hydroxylase), CYP302a1 (22-hydroxylase), CYP315a1 (2-hydroxylase), and CYP314a1 (20-hydroxylase)), have been identified and characterized in Drosophila melanogaster, Bombyx mori, and Aedes aeypti (7-13).Although glucosides (14), fatty acyl esters (15), and sulfate esters (16) of ecdysteroids are detected as minor products of phase II reaction in several insect species, the major products of the phase II reaction in most insect species are phosphate esters (17-20). The ovaries of most insect species have the capacity to accumulate ecdysteroid phosphates, which are physiologically inactive (21), in addition to the capacity to synthesize free ecdysteroids de novo (22,23). It has been suggested that the ecdystreoid phosphates that are accumulated in the ovaries are transferred to the eggs and function as a source of active free ecdysteroid before the prothoracic glands differentiate during embryonic development (24 -26). Recently, in B. mori eggs, a no...

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Comparative Studies of Ecdysteroid Metabolism between Diapause Eggs and Non-diapause Eggs of the Silkworm, Bombyx mori

Abstract: BioOne Complete (complete.BioOne.org) is a full-text database of 200 subscribed and open-access titles in the biological, ecological, and environmental sciences published by nonprofit societies, associations, museums, institutions, and presses.

Cited by 23 publications

References 26 publications

Shade is the Drosophila P450 enzyme that mediates the hydroxylation of ecdysone to the steroid insect molting hormone 20-hydroxyecdysone

Shade is the Drosophila P450 enzyme that mediates the hydroxylation of ecdysone to the steroid insect molting hormone 20-hydroxyecdysone

Ecdysone 20-monooxygenase in eggs of the silkworm,Bombyx mori: Enzymatic properties and developmental changes

Purification, Kinetic Characterization, and Molecular Cloning of a Novel Enzyme, Ecdysteroid 22-Kinase

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