DIFFERENTIAL INCORPORATION OF CHOLESTEROL AND CHOLESTEROL DERIVATIVESINTO ECDYSTEROIDS BY THE LARVAL RING GLANDS AND ADULT OVARIES OF DROSOPHILA-MELANOGASTER - A PUTATIVE EXPLANATION FOR THE L(3)ECD(1) MUTATION

Studies in vitro revealed that intact ring glands of Drosophila melano gaster convert tritiated cholesterol (C) and 25-hydroxycholesterol (25 C) via 7-dehydrocholesterol (7dC) and 7-dehydro-25-hydroxycholesterol (7d25C), respectively, to ecdysone (E) and 2-deoxyecdysone (2dE), whil e both intact and homogenized ovaries synthesize only 2dE from these p recursors. Emulsified 7d25C was incorporated directly into ecdysteroid s by these tissue preparations at a much greater rate than was 7d25C m ade in situ from 25C. To probe the basis of the biochemical defect in the ecdysteroid deficient conditional mutant ecdysoneless (ecd(1)), th e differential incorporation into ecdysteroids of C (via 7dC), and par ticularly of 25C (via 7d25C), was measured relative to that observed a fter the incubation of 7d25C directly with both wild type and mutant t issues in vitro at 30 degrees C, the restrictive temperature. Both C a nd 25C mere equally 7,8-dehydrogenated in situ to 7dC or 7d25C, respec tively, by both wild type and mutant tissues at 30 degrees C. However, the rate of subsequent conversion of either of these Delta(5,7)-stero l intermediates synthesized in situ to ecdysteroids was reduced an ave rage of 50% in the mutant tissues relative to the wild type. Yet, when emulsified 7d25C was incubated directly with either the wild type or mutant tissues at the restrictive temperature, the amplified rate of c onversion of the freely available 7d25C to ecdysteroid by these tissue s was identical. These data suggest that the defect in ecd(1) tissue-m ediated ecdysteroidogenesis does not involve a ''hit'' on any of the e nzymes involved in either the 7,8-dehydrogenation of C or 25C or in th e subsequent oxidation of 7d25C or 7dC to ecdysteroid. Rather, the mut ation appears to affect the expression of a gene governing the translo cation of Delta(5,7)-sterol intermediates from the subcellular compart ment where they are synthesized and/or stored to the site of subsequen t oxidation to ecdysteroid. Copyright (C) 1996 Elsevier Science Ltd