No description available.

Drosophila insulin like peptides (DILPs) and their human homolog insulin act as messengers to control many physiological processes in the body. Fields in which insulin signaling is crucial are e.g. growth, stress responses and aging. Consequently, many diseases are caused by disturbed insulin signaling, of which diabetes is the most prominent. During the last decades the functions of insulins and their signaling pathways have been studied in detail; what remains less well understood is how the production of insulin and insulin like peptides is regulated. The family of Neprilysins (Neps) belongs to the M13-zinc ion binding metallopeptidases. Neprilysins cleave peptides that regulate a wide range of cellular processes and are therefore linked to a variety of diseases like cancer, analgesia, hypertension or Alzheimer’s disease. In the fruit fly Drosophila melanogaster, five Neprilysins are expressed; but their in vivo substrates have not yet been identified. One of the Drosophila Neprilysins, Nep4, is expressed in the CNS, in muscle tissue, in cardiac tissue and in male reproductive organs. Nep4 is expressed in two isoforms, Nep4A and Nep4B. Isoform A is composed of a short intracellular domain, a transmembrane domain and a large extracellular domain containing the catalytically active center, whereas soluble Nep4B only consists of the extracellular domain. This thesis reveals that overexpression of catalytically active Nep4A in muscle tissue leads to animals with impaired insulin expression, decreased size and weight, affected feeding behavior and reduced locomotion speed. Further phenotypes are an impaired energy metabolism and larval lethality. Knockdown of the whole enzyme or knockout of its catalytic activity also interferes with feeding and locomotion speed and, in addition, causes pupal lethality. As an explanation for the phenotypes, Nep4 mediated hydrolysis of different short neuropeptide F (sNPF) species, which were identified as novel substrates of the peptidase, is proposed. sNPF is known to regulate insulin signaling and knockdown of sNPF phenocopies the Nep4 overexpression phenotypes, which suggests that Nep4 mediated hydrolysis of sNPF regulates insulin expression in the fly. Based on these results additional regulatory peptides were identified as novel Nep4 substrates. Among them are peptides that do not only regulate insulin signaling, but also feeding behavior (Hallier et al., 2016). These findings represent good evidence that muscle bound Nep4 is key to regulate homeostasis of distinct hemolymph circulating peptide hormones. Nep4 localizing to the surface of the central nervous system is likely necessary to ensure effective ligand clearance and thus proper regulation of corresponding peptide receptors.

No description available.

No description available.

No description available.

No description available.

No description available.

No description available.

No description available.

The present study focuses on the cardiac function of Drosophila melanogaster. Drosophila heart parameters are evolutionarily conserved, making Drosophila a useful human heart disease model. This model enables the in vivo investigation of physiological and genetic methods. This thesis is subdivided into four parts: parts 1-3 comprise the introductions of three publications, and part 4 presents unpublished data. The first publication is about the heart physiology of Drosophila. It explains how intracardiac valve cells work and proves their participation in blood flow directionality. A databased model shows the orientation of myofibrils within the valve cell. The myofibrils allow the valve cells to oscillate between a roundish and elongated cell shape. A toll-GFP enhancer line was shown to mediate strong reporter gene activity in the intracardiac valve of third instar larvae, pupae and adults. Transmission electron microscopy (TEM) analyses and immunohistochemical studies showed the differentiation of larval valve cells for the first time. The second publication focuses on the cardiac extracellular matrix (ECM), which contains two unique proteins - Lonely heart (Loh) and Pericardin (Prc). The study demonstrated that Loh is crucial for Prc recruitment to the developing matrix. Loh is anchored to the ECM by its thrombospondin type 1 repeat (TSR1-1) with its embedded putative glycosaminoglycan (GAG)-binding side. The N-terminus of Loh is proposed to face the plasma membrane. Prc is presumably recruited by two Loh TSR1 domains (TSR1-2 and TSR1-4). Nearly all Drosophila tissues, except salivary glands, create Prc networks through ectopic Loh expression. The study also found that the amount of Prc and Loh in the cardiac ECM influences heart function. The third publication investigated a set of neuropeptides and their ability to modulate cardiac function in third instar larvae. The results showed that 11 of the 19 tested peptides significantly affected the heart function in semi-intact larvae. Furthermore, the peptides’ in vivo relevance was tested through the knockdown of chronotropic peptide precursors. The study found that a RNAi mediated knockdown of all respective peptide precursors affected the heart rate. By combining semi-intact heart preparations and in vivo analyses, we identified several heartbeat-modulatory peptides in Drosophila. The unpublished data introduces a new software program called HIRO. It is written in Java, platform-independent and can easily detect the heart rhythm. Only mild anaesthesia and basic equipment are needed to record the Drosophila heartbeat. HIRO was used to show the influence of the RNAi-mediated downregulation of critical ECM proteins in Drosophila third instar larvae. The screen revealed Myospheroid and Laminin A as promising candidates that can significantly affect the heart parameters. HIRO is optimised for future applications and can be used as a high-throughput screening software with a simple setup. Taken together, this thesis provides new insights into the physiology and function of the Drosophila heart. The developed software HIRO comes with a user-friendly interface and a step-by-step introduction to easily conduct heart parameter measurements. HIRO will help to expand our knowledge of the fundamental processes in the model organism Drosophila melanogaster.