Changes in expressions of ecdysteroidogenic enzyme and ecdysteroid signaling genes in relation to Bombyx embryonic development.

Although the role of ecdysteroids in regulating egg diapause process in Bombyx mori is well documented, temporal changes in expression levels of genes involved in ecdysteroid biosynthesis and its downstream signaling are less well understood. In the present study, we studied changes in expression levels of genes involved in ecdysteroid biosynthesis and its downstream signaling during embryonic development of B. mori. Results showed that in diapause eggs, the expression of ecdysteroid-phosphate phosphatase (EPPase) gene and Halloween genes (Spook [Spo] and Shade [Shd]) remained at very low levels. However, in eggs whose diapause initiation was prevented by HCl, significant increases in the messenger RNA (mRNA) levels of EPPase, Spo, and Shd were detected during embryonic development. Other Halloween genes (Neverland [Nvd] and Phantom [Phm]) also showed different changes between diapause and HCl-treated eggs. However, genes of Disembodied (Dib) and Shadow (Sad) showed similar changes in both diapause and HCl-treated eggs. We further investigated changes in expression levels of ecdysone receptor genes (EcRA, EcRB1, and USP) and downstream signaling genes (E75A, E75B, E74A, E74B, Br-C, HR3, HR4, KR-H1, and FTZ-F1). Results showed that genes of EcRA and the other nuclear receptors (E75A, E75B, E74A, HR3, HR4, KR-H1, and FTZ-F1) exhibited significant differential patterns between diapause and HCl-treated eggs, with increased levels being detected during later stages of embryonic development in HCl-treated eggs. Differential temporal changes in expressions of genes involved ecdysteroid biosynthesis and its downstream signaling found between diapause and HCl-treated eggs were further confirmed using nondiapause eggs. Our results showed that nondiapause eggs exhibited the same changing patterns as those in HCl-treated eggs, thus clearly indicating potential correlations between expressions of these genes and embryonic development in B. mori. To our knowledge, this is the first comprehensive report to study the transcriptional regulation of ecdysteroidogenic and ecdysteroid signaling genes, thus providing useful information for a clearer understanding of insect egg diapause mechanisms.