Background Krüppel homolog 1 (Kr-h1) is a critical transcription factor for juvenile hormone (JH) signaling, known to play a key role in regulating metamorphosis and adult reproduction in insects. Kr-h1 can also be induced by molting hormone 20-hydroxyecdysone (20E), however, the underlying mechanism of 20E-induced Kr-h1 expression remains unclear. In the present study, we investigated the molecular mechanism of Kr-h1 induction by 20E in the reproductive system of a model lepidopteran insect, Bombyx mori. Results Developmental and tissue-specific expression analysis revealed that BmKr-h1 was highly expressed in ovaries during the late pupal and adult stages and the expression was induced by 20E. RNA interference (RNAi)-mediated depletion of BmKr-h1 in female pupae severely repressed the transcription of vitellogenin receptor (VgR), resulting in the reduction in vitellogenin (Vg) deposition in oocytes. BmKr-h1 specifically bound the Kr-h1 binding site (KBS) between − 2818 and − 2805 nt upstream of BmVgR and enhanced the transcription of BmVgR. A 20E cis-regulatory element (CRE) was identified in the promoter of BmKr-h1 and functionally verified using luciferase reporter assay, EMSA and DNA-ChIP. Using pull-down assays, we identified a novel transcription factor B. mori Kr-h1 regulatory protein (BmKRP) that specifically bound the BmKr-h1 CRE and activated its transcription. CRISPR/Cas9-mediated knockout of BmKRP in female pupae suppressed the transcription of BmKr-h1 and BmVgR, resulting in arrested oogenesis. Conclusion We identified BmKRP as a new transcription factor mediating 20E regulation of B. mori oogenesis. Our data suggests that induction of BmKRP by 20E regulates BmKr-h1 expression, which in turn induces BmVgR expression to facilitate Vg uptake and oogenesis.

Spodoptera litura is a destructive agricultural pest in tropical and subtropical areas. Understanding the molecular mechanisms of S. litura adaptation to its preferred host plants may help identify target genes useful for pest control. We used high-throughput sequencing to characterize the expression patterns of messenger RNAs (mRNAs) and microRNAs (miRNAs) in the midgut of S. litura fed on Brassica juncea for 6 h and 48 h. A total of 108 known and 134 novel miRNAs were identified, 29 miRNAs and 237 mRNAs were differentially expressed at 6 h of B. juncea feeding, 26 miRNAs and 433 mRNAs were differentially expressed at 48 h. For the mRNAs, the up-regulated genes were mostly enriched in detoxification enzymes (cytochrome P450, esterase, glutathione S-transferase, uridine diphosphate-glucuronosyl transferase), while the down-regulated genes were mostly enriched in proteinases and immune-related genes. Furthermore, most detoxification enzymes begin to up-regulate at 6 h, while most digestion and immunerelated genes begin to up-or down-regulate at 48 h. Eighteen and 37 differently expressed transcription factors were identified at 6 h and 48 h, which may regulate the functional genes. We acquired 136 and 41 miRNA versus mRNA pairs at 6 h and 48 h, respectively. Some down-regulated and up-regulated miRNAs were predicted to target detoxification enzymes and proteinases, respectively. Real-time quantitative polymerase chain reaction of nine randomly selected miRNAs and 28 genes confirmed the results of RNA-seq. This analyses of miRNA and mRNA transcriptomes provides useful information about the molecular mechanisms of S. litura response to B. juncea.

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