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.