Pupal forewing epidermis of the Coleoptera, Tenebrio rnolitor, was used to develop an in vitro system to study the hormonal control of metamorphosis at the cellular and molecular levels. Exposure to 1 pM 20-hydroxyecdysone for 48 h caused the formation of a typical adult cuticle. Under these conditions the expression of ACP-20, an adult-specific cuticular gene, was fivefold higher than in absence of exogenous hormone. This stimulation was also observed when a higher level of 20-hydroxyecdysone was maintained, and prevented by protein inhibitors, indicating that 20-hydroxyecdysone does not act directly on this gene. Exposure to 20-hydroxyecdysone followed by exposure in hormone-free medium caused the cessation of this stimulation, showing the requirement of the 20-hydroxyecdysone continuous presence for stimulating ACP-20 gene expression. Thus, unlike the other cuticular protein genes so far studied, its expression is not repressed by 20-hydroxyecdysone, and does not need the decline in ecdysteroids titer.

The accumulation of transcripts from two adult-specific cuticular genes (ACP-20 and ACP-22) is shown to be modified after addition of exogenous 20-hydroxyecdysone. In the continuous presence of high levels of the hormone, the expression of ACP-20 gene is significantly weaker than that of untreated controls, while ACP-22 expression is 2.5-fold increased. During active synthesis of the ACP messages, a 0.5 microg 20-hydroxyecdysone injection causes a rapid 2-fold increase in ACP-22 mRNA and is not able to repress ACP-20 mRNA accumulation. We conclude that these genes whose transcripts appear in an almost coordinated manner in epidermal cells during the moulting cycle are regulated by ecdysteroids in a different way. In order to undertake a functional dissection of the promoter regions of ACP-22 gene, we have isolated and sequenced a genomic clone. The sequence similarities with other cuticular protein genes are discussed.

In this paper we describe a novel family of miniature inverted-repeat transposable elements (MITEs), named DEC, isolated from the genome of the beetle Tenebrio molitor. These elements are highly reiterated and their number is estimated to be around 3500 per haploid genome. Two of them have been isolated and the two sequences are 84% identical. Like other MITEs, they are characterized by their small size, their A + T richness, the presence of terminal inverted repeats and the absence of open reading frames. These data suggest that MITEs are probably widely distributed in arthropods.