Mediation of ecdysone synthesis in Manduca sexta by a hemolymph enzyme

Abstract

The prothoracic glands of Manduca sexta synthesize dehydroecdysone, which is rapidly converted to ecdysone through the mediation of a hemolymph enzyme, a 3 P-forming-3-ketosteroid reductase. The hemolymph protein fraction (HPF) containing this enzyme was obtained from diapausing and nondiapausing pupae, isolated abdomens, surgically manipulated pupae, etc., and in all cases had the capacity to affect the conversion of dehydroecdysone to ecdysone. The enzyme is heat labile, is inactivated by trypsin, and has a molecular weight of between 20,000 and 30,000. The data indicate that the conversion of dehydroecdysone to ecdysone exhibits linear kinetics and may be dependent on both the enzyme concentration and the concentration of NADPH at the beginning of the reaction but may be limited by the absolute amount of reducing equivalents after 10 miri, under the experimental conditions utilized. The capacity of the enzyme to reduce dehydroecdysone was titered in the hemolymph during the last larval instar and during prepupal and pupal lifewith maximum capacityexhibited at the beginning of the instar, on day 8 of larval life and at day 1 of pupal life. Even at its lowest point at day 5 , l ml of hemolymph was able to convert 77 pmol (-35 ng) dehydroecdysone to ecdysone in 1 min. These results require a new interpretation of the control of molting in the Lepidoptera.

Ecdysone Titer Determined by 3DE-3β-Reductase Enhances the Immune Response in the Silkworm

“…In insects, the biosynthesis of the 20E is mainly from the cholesterol pathway in the prothoracic glands (8). Nevertheless, ecdysone can also be directly synthesized from 3-dehydroecdysone (3DE) in other tissues (22)(23)(24). In immature stages of most lepidopteran species, 3DE is the major product of the prothoracic glands.…”

“…In immature stages of most lepidopteran species, 3DE is the major product of the prothoracic glands. After release from the glands, 3DE is rapidly reduced to ecdysone by 3DE-3b-reductase in hemolymph, and then the ecdysone is converted into 20E by 20-hydroxylase in the target tissues (22)(23)(24)(25)(26). 3DE-3b-reductase is considered to be an important enzyme in the biosynthesis of the ecdysone (27).…”

Purification and Characterisation of Haemolymph 3‐Dehydroecdysone 3β‐Reductase in Relation to Ecdysteroid Biosynthesis in the Cotton Leafworm Spodoptera littoralis

“…Within a single order, therefore, there are marked differences in the ratio of ecdysteroids from the prothoracic glands. Physiologically, these differences are not easily explained, although there is some evidence to support a developmental significance, since it has been noted that the 3-dehydroecdysone : ecdysone ratio increased as the last larval instar of M. sexta progressed [3]. Of relevance is the observation in the current work that no discernible interconversion of ecdysone and 3-dehydroecdysone occurred upon incubation of these steroids in turn with S. littoralis prothoracic glands.…”

“…NADPH was also a more effective cofactor than NADH for the enzyme from M. sexta [3]. Thus, the cofactor may be an important factor contributing to regulation of the enzyme activity.…”

Molecular Cloning and Characterization of Hemolymph 3-Dehydroecdysone 3β-Reductase from the Cotton Leafworm,Spodoptera littoralis

“…After secretion from the prothoracic glands, the 3DE is reduced to ecdysone by an NAD(P)H-linked 3DE 3␤-reductase in the hemolymph (3,4,6,7). Ecdysone undergoes C-20 hydroxylation in certain peripheral tissues yielding 20-hydroxyecdysone, which is considered to be the true molting hormone in most insects (8).…”