Krüppel Homolog 1 Inhibits Insect Metamorphosis via Direct Transcriptional Repression of Broad-Complex, a Pupal Specifier Gene

• Published in: The Journal of biological chemistry 2016-01, Vol.291 (4), p.1751-1762
• Main Authors: Kayukawa, Takumi, Nagamine, Keisuke, Ito, Yuka, Nishita, Yoshinori, Ishikawa, Yukio, Shinoda, Tetsuro
• Format: Article
• Language: English
• Subjects: Animals; Bombyx - genetics; Bombyx - growth & development; Bombyx - metabolism; development; Developmental Biology; Down-Regulation; Ecdysterone - metabolism; endocrinology; Gene Expression Regulation, Developmental; hormone; insect; Insect Proteins - genetics; Insect Proteins - metabolism; Juvenile Hormones - metabolism; Kruppel-Like Transcription Factors - genetics; Kruppel-Like Transcription Factors - metabolism; Larva - genetics; Larva - growth & development; Larva - metabolism; Metamorphosis, Biological; Promoter Regions, Genetic; Protein Binding; Pupa - genetics; Pupa - growth & development; Pupa - metabolism; Signal Transduction; signaling; Species Specificity; Transcription Factors - genetics; Transcription Factors - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M115.686121

The Broad-Complex gene (BR-C) encodes transcription factors that dictate larval-pupal metamorphosis in insects. The expression of BR-C is induced by molting hormone (20-hydroxyecdysone (20E)), and this induction is repressed by juvenile hormone (JH), which exists during the premature larval stage. Krüppel homolog 1 gene (Kr-h1) has been known as a JH-early inducible gene responsible for repression of metamorphosis; however, the functional relationship between Kr-h1 and repression of BR-C has remained unclear. To elucidate this relationship, we analyzed cis- and trans elements involved in the repression of BR-C using a Bombyx mori cell line. In the cells, as observed in larvae, JH induced the expression of Kr-h1 and concurrently suppressed 20E-induced expression of BR-C. Forced expression of Kr-h1 repressed the 20E-dependent activation of the BR-C promoter in the absence of JH, and Kr-h1 RNAi inhibited the JH-mediated repression, suggesting that Kr-h1 controlled the repression of BR-C. A survey of the upstream sequence of BR-C gene revealed a Kr-h1 binding site (KBS) in the BR-C promoter. When KBS was deleted from the promoter, the repression of BR-C was abolished. Electrophoresis mobility shift demonstrated that two Kr-h1 molecules bound to KBS in the BR-C promoter. Based on these results, we conclude that Kr-h1 protein molecules directly bind to the KBS sequence in the BR-C promoter and thereby repress 20E-dependent activation of the pupal specifier, BR-C. This study has revealed a considerable portion of the picture of JH signaling pathways from the reception of JH to the repression of metamorphosis.