Abstract
The juvenile hormone (JH) signalling and ecdysone signalling pathways are crucial endocrine signalling pathways that orchestrate the metamorphosis of insects. The metamorphic process, the morphological change from the immature to adult forms, is orchestrated by the dramatic reduction of JH and downstream transcription factors. The Krüppel-homologue 1 (Kr-h1), a downstream transcription factor of the JH signalling pathway, represses E93 expression with an anti-metamorphic effect. However, the biochemical interaction between Kr-h1 and E93 and how the interaction regulates ovary development, a sensitive readout for endocrine regulation, remain unknown. In brown planthopper, Nilaparvata lugens, we found that the downregulation of Kr-h1 partially recovered the deteriorating effect of E93 knock-down on metamorphosis. Dual knock down of E93 and Kr-h1 increased ovary development and the number of eggs laid when compared to the effects of the knock down of E93 alone, indicating that the knock down of Kr-h1 partially recovered the deteriorating effect of the E93 knock-down on ovary development. In summary, our results indicated that E93 and Kr-h1 have antagonistic effects on regulating metamorphosis and ovary development. We tested the biochemical interaction between these two proteins and found that these molecules interact directly. Kr-h1 V and E93 II undergo strong and specific interactions, indicating that the potential interacting domain may be located in these two regions. We inferred that the nuclear receptor interaction motif (NR-box) and helix-turn-helix DNA binding motifs of the pipsqueak family (RHF1) are candidate domains responsible for the protein–protein interaction between E93 and Kr-h1. Moreover, the HA-tagged E93 and FLAG-tagged Kr-h1 were co-localized in the nucleus, and the expression of E93 was increased when Kr-h1 was downregulated, supporting that these two proteins may interact antagonistically. JH and ecdysone signalling are critical for the control of ovary development and pest populations. Our result is important for understanding the interactions between E93 and related proteins, which makes it possible to identify potential targets and develop new pesticides for pest management.
Highlights
The juvenile hormone (JH) signalling and ecdysone signalling pathways are two crucial endocrine signalling pathways that regulate the metamorphosis of insects, which is the morphological change from immature to adult forms and is orchestrated by the dramatic reduction in the JH level and the downstream transcription factor of the JH signalling pathway [1,2,3,4,5]
We down-regulated the expression of E93 and Krüppel-homologue 1 (Kr-h1) by the injection of dsRNA into 4th instar nymphs
The phenotype is reminiscent of a defect in specifying adult metamorphosis (Figure 1A), especially the external female genitalia, which is either undeveloped or underdeveloped compared to that of the control injected with dsGFP (Figure 1A), and the defect is less intense in the dual knock-down (Figure 1A), suggesting the normal morphology is partially recovered
Summary
The juvenile hormone (JH) signalling and ecdysone signalling pathways are two crucial endocrine signalling pathways that regulate the metamorphosis of insects, which is the morphological change from immature to adult forms and is orchestrated by the dramatic reduction in the JH level and the downstream transcription factor of the JH signalling pathway [1,2,3,4,5]. The binding of JH to the Methoprene-tolerant (Met) and Taiman (Tai) receptor complex induced the expression of a C2H2 down-stream zinc-finger transcription factor Krüppel-homologue 1 (Kr-h1) [6,7,8,9]. The transcription factor E93 is induced by ecdysone and broadly expressed in pupae and adult insects [10,11,12,13,14]. The molecular function of E93 includes determining programmed cell death in larval salivary glands during development and transducing ecdysone signalling to induce autophagy and caspase activity in the remodelling fat body of D. melanogaster [14,19]. The downregulation of Kr-h1 leads to a premature phenotype, and even bypasses the pupae stage in holometabolous insects [20]; both E93 and Kr-h1 regulate metamorphosis [18,20]
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