The P450 enzymes (mixed function oxidases, cytochrome P450 monooxygenases), a diverse class of enzymes found in virtually all insect tissues, fulfill many important tasks, from the synthesis and degradation of ecdysteroids and juvenile hormones to the metabolism of foreign chemicals of natural or synthetic origin. This diversity in function is achieved by a diversity in structure, as insect genomes probably carry about 100 P450 genes, sometimes arranged in clusters, and each coding for a different P450 enzyme. Both microsomal and mitochondrial P450s are present in insects and are best studied by heterologous expression of their cDNA and reconstitution of purified enzymes. P450 genes are under complex regulation, with induction playing a central role in the adaptation to plant chemicals and regulatory mutations playing a central role in insecticide resistance. Polymorphisms in induction or constitutive expression allow insects to scan their P450 gene repertoire for the appropriate response to chemical insults, and these evolutionary pressures in turn maintain P450 diversity.

Insect P450 enzymes.

Ecdysone metabolism and distribution during the pupal-adult development of Manduca sexta

Periodic pulses of the insect steroid molting hormone 20-hydroxyecdysone (20E), acting via its nuclear receptor complex (EcR/USP), control gene expression at many stages throughout Drosophila development. However, during the last larval instar of some lepidopteran insects, subtle changes in titers of ecdysteroids have been documented, including the so-called "commitment peak." This small elevation of 20E reprograms the larva for metamorphosis to the pupa. Similar periods of ecdysteroid immunoreactivity have been observed during the last larval instar of Drosophila. However, due to low amplitude and short duration, along with small body size and staging difficulties, their timing and ecdysteroid composition have remained uncertain. Employing a rigorous regimen of Drosophila culture and a salivary gland reporter gene, Sgs3-GFP, we used RP-HPLC and differential ecdysteroid RIA analysis to determine whole body titers of 20E during the last larval instar. Three small peaks of 20E were observed at 8, 20, and 28 hr following ecdysis, prior to the well-characterized large peak around the time of pupariation. The possible regulation of 20E levels by biosynthetic P450 enzymes and the roles of these early peaks in coordinating gene expression and late larval development are discussed.

https://onlinelibrary.wiley.com/doi/pdf/10.1002/dvdy.20626

Discrete Pulses of Molting Hormone, 20-Hydroxyecdysone, During Late Larval Development of Drosophila melanogaster: Correlations With Changes in Gene Activity

CYP18A1, a key enzyme of Drosophila steroid hormone inactivation, is essential for metamorphosis.

Publisher Summary This chapter focuses on the techniques used for ecdysteroid analysis and the biosynthesis and metabolism of these hormones. Ecdysteroids could be defined either by their biological activity (molting hormones) or their chemical structure. The huge chemical diversity of ecdysteroids results from variations in the number and position of OH groups, which can be either free or conjugated to various polar or apolar moieties, giving rise to the 70 different molecules. The greatest diversity has been found in the eggs/embryos of Orthoptera and Lepidoptera, probably because they contain very large amounts of these molecules. The nomenclature of insect ecdysteroids is generally derived from the ecdysone name, but this is not always the case, and the trivial name of many ecdysteroids is related to the name of the species from which they have been isolated; standardized abbreviations have been proposed for insect ecdysteroids and are used.

Ecdysteroid Chemistry and Biochemistry

Sample processing for high-performance liquid chromatography of ecdysteroids

Insects convert ecdysone and 20-hydroxyecdysone into their corresponding 26-oic derivatives, named ecdysonoic acid and 20-hydroxyecdysonoic acid respectively The conversion takes piace in several tissues and can either be the only pathway for converting ecdysone into highly polar ecdysteroids, or coexist with various conjugating mechanisms



20-Hydroxyecdysonoic acid was isolated from Pieris brassicae pupae as its methyl ester derivative Its chemical structure was identified by Cl/D mass spectrometry and compared with a synthetic compound (20-hydroxy-25-deoxyecdysonoic acid) chemically prepared by oxidation of inokosterone (20,26-dihydroxy-25-deoxyecdysone) Natural ecdysonoic acids appear to exist as a mixture of 25R and 25S isomers



The significance of this pathway is discussed in comparison with similar reactions occuring in the metabolism of steroid hormones in vertebrates

Conversion of ecdysone and 20-Hydroxyecdysone into 26-OIC derivatives is a major pathway in larvae and pupae of species from three insect orders

Summary The metabolism of ecdysone and 20-hydroxyecdysone has been investigated in fifth instar larvae and male adults of Locusta migratoria (Insecta, Orthoptera). The major pathways have been elucidated by labelling experiments, followed by the isolation and the characterization of the different metabolites. These ecdysteroids are converted by locusts through three major reactions: (1) C-26 hydroxylation followed by conversion to ecdysonoic acids, (2) C-3 acetylation, and (3) C-2 phosphate ester formation. The combination of (2) and (3) gives rise to acetylphosphate derivatives, which represent the major conjugates in insect faeces. Minor metabolites are also formed, corresponding to 3-dehydroecdysteroids and phosphate conjugates of various metabolites which have not yet been fully characterized. The reactions are identical in larvae and adults and are similar to those described for embryos of the same species and the related Schistocerca gregaria. In vitro organ incubations have allowed us to determine ...

Jean-François Modde

Papers

Sample processing for high-performance liquid chromatography of ecdysteroids

Conversion of ecdysone and 20-Hydroxyecdysone into 26-OIC derivatives is a major pathway in larvae and pupae of species from three insect orders

Ecdysone metabolism in Locusta migratoria larvae and adults