Reduction of embryonic E93 expression as a hypothetical driver of the evolution of insect metamorphosis.

The early embryo of the cockroach Blattella germanica exhibits high E93 expression. In general, E93 triggers adult morphogenesis during postembryonic development. Here we show that E93 is also crucial in early embryogenesis in the cockroach, as a significant number of E93-depleted embryos are unable to develop the germ band under maternal RNAi treatment targeting E93. Moreover, transcriptomic analysis indicates that E93 depletion results in important gene expression changes in the early embryo, and many of the differentially expressed genes are involved in development. Then, using public databases, we gathered E93 expression data in embryo and preadult stages, finding that embryonic expression of E93 is high in hemimetabolan species (whose juveniles, or nymphs, are similar to the adult) and low in holometabolans (whose juveniles, or larvae, are different from the adult). E93 expression is also low in Thysanoptera and in Hemiptera Sternorrhyncha, hemimetabolans with postembryonic quiescent stages, as well as in Odonata, the nymph of which is very different from the adult. In ametabolans, such as the Zygentoma Thermobia domestica, E93 transcript levels are very high in the early embryo, whereas during postembryonic development they are medium and relatively constant. We propose the hypothesis that during evolution, a reduction of E93 expression in the embryo of hemimetabolans facilitated the larval development and the emergence of holometaboly. Independent decreases of E93 transcripts in the embryo of Odonata, Thysanoptera, and different groups of Hemiptera Sternorrhyncha would have allowed the development of modified juvenile stages adapted to specific ecophysiological conditions.

Regulation of insulin-like peptide expression in adult Blattella germanica females.

The insulin-IGF-signalling (IIS) pathway regulates key processes in metazoans. The pathway is activated through the binding of the ligands, which in insects are usually referred to as insulin-like peptides (ILPs), to a class of receptor tyrosine kinases, the insect insulin receptor. To study the pathway regulation, it is therefore essential to understand how ILPs are produced and released. In this study we analysed the factors that regulate the expression of the seven ILPs (BgILPs) expressed in adult females of the German cockroach, Blattella germanica. The results showed that the starvation-induced expression reduction of brain BgILP3, 5 and 6 and fat body BgILP7 is not due to reduced juvenile hormone (JH) or decreased TOR pathway activity. In addition, depletion of FoxO in starved females did not correct the low levels of these BgILPs, but even reduced further BgILP5 expression, indicating the need to maintain certain basal levels of BgILP5 even during starvation. Furthermore, JH promoted increased BgILP5 and decreased BgILP3 expression in the brain, an effect that required Methoprene-tolerant (Met), the JH receptor, but not Krüppel homolog 1 (Kr-h1), the main JH transducer. On the other hand, JH inhibited the expression of BgILP7 in the fat body, although in this case, the action required both Met and Kr-h1. In addition, JH reduction treatments produced a decrease in the expression of the insulin receptor in the fat body, which suggests an increase in IIS. The results show a peculiar regulation of ILP expression in adult B. germanica females, which is clearly different than that seen in other species. This is understandable given that gene duplications in recent clades have resulted in different sets of ILP genes, involving substantial changes in gene regulatory networks.

Key roles of the Broad-Complex gene in insect embryogenesis.

The Broad-Complex (BR-C) is a primary response gene in the ecdysteroid signaling pathway which encodes a family of transcription factors that play a key role in metamorphic processes. With the aim of studying the possible functions of BR-C in the embryogenesis of a short germ band insect, we cloned BR-C cDNAs in the cockroach Blattella germanica (BgBR-C) and obtained the full coding region of six BR-C isoforms (Z1-Z6) of which, Z1-Z5 have homologous sequences in other insects, whereas Z6 is new. Temporal-expression patterns indicate that BgBR-C isoforms are present throughout the embryogenesis of B. germanica, although with weak fluctuations. Silencing all BgBR-C isoforms in the embryo through parental RNAi elicited a diversity of phenotypes. These phenotypes suggest roles for BgBR-C in different embryogenesis processes of B. germanica, mainly in the formation of the germ band, in the transition from 30% to 35% of total development, when the amnion and serosa rupture and the secondary dorsal organ is formed, and in the hatching process, which involves neural, endocrine and muscular coordination.