Enzymes of ecdysteroid transformation and inactivation in the midgut of the cotton leafworm, Spodoptera littoralis: properties and developmental profiles

Abstract

In the midgut cytosol of Lepidoptera, ecdysteroids undergo inactivation by transformation via the 3-dehydro derivative to the corresponding 3-epiecdysteroid (3 alpha-hydroxy) and by phosphate conjugation. The oxygen-dependent oxidase catalyses formation of 3-dehydroecdysteroid, which can be reduced either irreversibly by 3-dehydroecdysone 3 alpha-reductase to 3-epiecdysteroid, or by 3-dehydroecdysone 3 beta-reductase back to the initial ecdysteroid. Furthermore, these ecdysteroids undergo further inactivation by phosphorylation. These ecdysteroid transformations have been investigated in last instar larvae of the cotton leafworm, Spodoptera littoralis. The products of the phosphorylation have been characterized as predominantly ecdysteroid 2-phosphate accompanied by smaller amounts of the corresponding 22-phosphate. The phosphotransferases require Mg2+ and ATP. Whereas the 3-dehydroecdysone 3 alpha-reductase has a clear preference for NADPH rather than NADH, the corresponding 3 beta-reductase markedly favours NADH. The physiological significance of the latter enzyme is unclear. The profiles of the various enzymic activities in dialysed midgut cytosol supplemented with appropriate cofactors were determined throughout the last larval instar. All activities were detectable throughout the instar, but the respective enzymes exhibited maxima at different times. Ecdysone oxidase showed a peak early in the instar, with 3-dehydroecdysone 3 alpha-reductase increasing to a peak as the former activity declined. The 3-dehydroecdysone 3 beta-reductase exhibited peak activity late in the instar, a profile similar to that observed for the corresponding haemolymph enzyme involved in reduction of the 3-dehydroecdysone product of the prothoracic glands to ecdysone. Thus, the significance of the midgut 3 beta-reductase may be related to production of active hormone. Both ecydsteroid 22- and 2-phosphotransferases showed high activities early in the instar and then declined. The physiological significance of the profiles for the ecdysone oxidase, the 3-dehydroecdysone 3 alpha-reductase and phosphotransferases is unclear.

Induction of Enzymes Involved in Molting Hormone (Ecdysteroid) Inactivation by Ecdysteroids and an Agonist, 1,2-Dibenzoyl-1-tert-butylhydrazine (RH-5849)

“…These enzymatic activities in the midgut cytosol ( Fig. 1) also exhibit developmental changes (8,9).…”

3-dehydroecdysone secretion by the molting gland of the cockroach,Periplaneta americana (L.)

“…The formation of this endproduct is irreversible. The conversion from 3DE to ecdysone is catalyzed by 3β-ketoreductase (Webb et al, 1995).…”

“…In this insect, however, 2-deoxyecdysone is secreted by the prothoracic glands and seems to serve as a prohormone. Recently, Webb et al (1995) and Chen et al (1996) provided evidence that in last instar larvae of the cotton leafworm, Spodoptera littoralis, 3β-ketoreductase exhibits peak activity late in the instar. The significance of the 3β-ketoreductase activity late in the instar may be related to the production peak of the molting hormone.…”

“…It has been documented that 3DE is rapidly converted to ecdysone in the hemolymph by a ketoreductase (3-oxoecdysteroid-3β-reductase) (Blais and Lafont, 1984) present in the hemolymph (Warren et al, 1988;Roussel, 1992;Chen et al, 1996;Nomura et al, 1996) and in other tissues such as the midgut (Webb et al, 1995) and eggs . The ratio of 3DE to ecdysone secreted by the prothoracic glands varies greatly among the investigated lepidopteran species between 2.5:1 (Galleria mellonella) and 36:1 (Papilio xuthus) (Kiriishi et al, 1990).…”

Ectopic expression of ecdysone oxidase impairs tissue degeneration in Bombyx mori

“…By contrast, less is known about 20E metabolism. Although several genes that function in 20E inactivation have been identified, the biological functions of genes involved in this pathway are not well understood [3,5].…”