Ecdysteroids are key regulators of embryonic development as well as molting and metamorphosis in insects. Although an active form of ecdysteroids, 20-hydroxyecdysone (20E) is known to be produced through ecdysteroidogenesis from cholesterol and dephosphorylation of 20E-phosphate during embryogenesis in Lepidoptera, the importance of these production mechanisms in embryonic development has been unclear. Here, we investigated the activation timing of ecdysteroidogenesis from cholesterol and 20E-phosphate dephosphorylation during early embryogenesis in non-diapause eggs of the silkmoth Bombyx mori by observing morphological development, quantifying 20E and 20E-phosphate, measuring transcripts of enzymes involved in 20E production, and detecting activity of these enzymes using egg extracts. Stage-dependent 20E fluctuation and changes in mRNA amounts of enzymes suggest that the two 20E-producing mechanisms are activated at different stages during embryogenesis. Furthermore, knockdown of a dephosphorylation enzyme delayed development at early embryogenesis, whereas knockdown of an ecdysteroidogenic enzyme delayed development at early-middle embryogenesis. These results suggest that 20E is primarily produced initially by dephosphorylation of 20E-phosphate, and then by ecdysteroidogenesis from cholesterol to induce progression of embryonic development in B. mori.

蜕皮类固醇是昆虫胚胎发育以及蜕皮和变态的关键调节剂。尽管已知蜕皮类固醇的一种活性形式，即20-羟基蜕皮酮（20E）是通过鳞状类固醇通过胆固醇生成和在鳞翅目的胚胎发生过程中20E-磷酸盐的去磷酸化而产生的，但这些生产机制在胚胎发育中的重要性尚不清楚。在这里，我们调查了蚕蛾非滞育卵中早期胆固醇生成过程中胆固醇和20E-磷酸去磷酸化过程中蜕皮类固醇生成的激活时间。通过观察形态发展，定量20E和20E-磷酸盐，测量参与20E产生的酶的转录本以及使用蛋提取物检测这些酶的活性。阶段依赖性20E波动和酶mRNA含量的变化表明，两种20E产生机制在胚胎发生过程的不同阶段被激活。此外，去磷酸化酶的敲低延迟了胚胎早期的发育，而蜕皮类固醇生成酶的敲低则延迟了胚胎中早期的发育。这些结果表明，20E最初主要是通过20E-磷酸盐的去磷酸化产生的，然后是胆固醇中蜕皮类固醇生成的产物，以诱导桑蚕中胚胎发育的进展。