Reproduction and feeding in insects - the Allatotropin receptor in Tribolium castaneum and the takeout gene in Schistocerca gregaria.

Abstract

The red flour beetle, Tribolium castaneum and the desert locust, Schistocerca gregaria are important pest insect species. They both can cause a lot of damage to human food supplies; especially locusts, that have the ability to undergo phase transition and can change into voracious swarms. Two of the possible methods to control pests, are to intervene with either feeding or reproduction. In the presented doctoral research, we explored several candidate genes involved in both reproduction and feeding in insects. We characterized a G- protein coupled receptor in T. castaneum that could be activated by the allatotropin-like peptide (ATL) of T. castaneum, as well as by allatotropins (ATs) of other insects, in a dose-dependent and reversible way. AT is particularly known to stimulate the juvenile hormone (JH) synthesis and release by the corpora allata (CA), to inhibit ion transport over larval gut epithelium and to be myostimulatory. The tissue distribution of the allatotropin receptor (ATR) in T. castaneum showed a remarkable male biased transcription pattern and highest levels were present in tissues related to reproduction. By means of an RNAi mediated knockdown of ATR, we confirmed these links to male reproduction, as ATR knockdown in males caused a decrease in progeny and a diminished male accessory gland (MAG) size. As these effects were also seen in a JH acid O-methyl-transferase (JHAMT, enzyme in JH synthesis) knockdown experiment in T. castaneum, this points to a link of ATR with JH synthesis. Recently, more and more studies point to JH production and a function of JH in the adult MAGs. Our study reinforces this theory and suggests an involvement of Trica-ATL in the regulation of JH in male reproduction. When we compared recently characterized allatotropin receptors, Aedes aegypti ATR and Tc-ATR were more alike in tissue distribution and activating ligands than the characterized Manduca sexta ATR. We suggest there are possibly two or more groups of ATRs in insects involved in the different functions linked to AT.To study the link between feeding and reproduction, we performed starvation and re-feeding experiments in the desert locust and compared relative gene transcription of several reproduction linked genes: Jhamt, JH epoxide hydrolase (jheh), at, allatostatin (ast), insulin related peptide (irp), ovary maturating parsin (omp) and adipokinetic hormone II (akh II). By starting starvation before onset of sexual maturation, the later was stopped and postponed to the period following re-feeding. Transcription of JH linked genes Jhamt, jheh, at and ast in control animals all showed a trend that is consistent with JH levels. JH production was already shown to decrease upon starvation. However, starvation had a decreasing effect on transcription of jheh (JH degradation function). We suggest that the lower JH titer during starvation is not due to JH degradation by JHEH. We showed that the effect of starvation onSg at and ast transcript levels was different in male and female, with at levels being influenced in males and ast in females. Previously in S. gregaria, no influence of these peptides was shown on the JH production by the CAs. However, as stated above, we also found several indications of JH involvement in the regulation of the MAGs reproductive functions. We propose an AT and AST influence on JH production but in organs other as the CA, probably MAGs in male and ovaries in females. The reduced jhamt and omp transcription in starved individuals was not restored after re-feeding. It can be suggested that both are responsible for the persistent effect of starvation on reproduction even under re-feeding conditions. Akh II transcription showed a peak upon starvation in females and a peak upon re-feeding in males. By BLAST search in a neural EST database with Drosophila Takeout (TO), we found 3 takeout (to) genes in S. gregaria. One form, Sg-to2, was characterized in detail. Sg-to2 transcript levels displayed a circadian rhythm similar to period and were decreased upon starvation. Transcription of Sg-to2 was highest in cuticular structures with epidermis, with highest levels in antennae and labial palps. This suggests a function in interaction with the environment by intervening in odorant perception. These structures are also involved in male courtship behavior and transcription was mainly male biased. In abdominal cuticle with epidermis a role for Sg-to2 in reproduction was suggested but this only in males. Knockdown of Sg-to2 in males caused a delay in mating. We showed that Sg-to2 is down regulated by knockdown of the sex determination gene fruitless (fru) but not vice versa, suggesting that Sg-to2 is downstream of fru. In non cuticular tissues, transcript level was highest in the AKH producing corpora cardiaca. Female Sg-to2 knockdown locusts ate more compared to control and male Sg-to2 knockdown locusts. We suggest, in similarity with results in Drosophila, a role for TO in energy homeostasis. We further propose TO interacts with AKH in energy mobilization during reproduction. Difference in Sg-to2 transcript levels between solitary and gregarious phases were only observed during starvation, suggesting a role in differentially coping with limited food availability of both phases. In conclusion our study gave more insight in the physiology of insects. These insights can hopefully contribute to improve future pest management strategies.nrpages: 213status: publishe