Molecular and cellular organization of odorant binding protein genes in Drosophila.

Chemosensation is important for the survival and reproduction of animals. The odorant binding proteins (OBPs) are thought to be involved in chemosensation together with chemosensory receptors. While OBPs were initially considered to deliver hydrophobic odorants to olfactory receptors in the aqueous lymph solution, recent studies suggest more complex roles in various organs. Here, we use GAL4 transgenes to systematically analyze the expression patterns of all 52 members of the Obp gene family and 3 related chemosensory protein genes in adult Drosophila, focusing on chemosensory organs such as the antenna, maxillary palp, pharynx, and labellum, and other organs such as the brain, ventral nerve cord, leg, wing, and intestine. The OBPs were observed to express in diverse organs and in multiple cell types, suggesting that these proteins can indeed carry out diverse functional roles. Also, we constructed 10 labellar-expressing Obp mutants, and obtained behavioral evidence that these OBPs may be involved in bitter sensing. The resources we constructed should be useful for future Drosophila OBP gene family research.

Postprandial sodium sensing by enteric neurons in Drosophila.

Sodium is essential for all living organisms1. Animals including insects and mammals detect sodium primarily through peripheral taste cells2-7. It is not known, however, whether animals can detect this essential micronutrient independently of the taste system. Here, we report that Drosophila Ir76b mutants that were unable to detect sodium2 became capable of responding to sodium following a period of salt deprivation. From a screen for cells required for the deprivation-induced sodium preference, we identified a population of anterior enteric neurons, which we named internal sodium-sensing (INSO) neurons, that are essential for directing a behavioural preference for sodium. Enteric INSO neurons innervate the gut epithelia mainly through their dendritic processes and send their axonal projections along the oesophagus to the brain and to the crop duct. Through calcium imaging and CaLexA experiments, we found that INSO neurons respond immediately and specifically to sodium ions. Notably, the sodium-evoked responses were observed only after a period of sodium deprivation. Taken together, we have identified a taste-independent sodium sensor that is essential for the maintenance of sodium homeostasis.

Wg/Wnt1 and Erasp link ER stress to proapoptotic signaling in an autosomal dominant retinitis pigmentosa model.

The endoplasmic reticulum (ER) is a subcellular organelle essential for cellular homeostasis. Perturbation of ER functions due to various conditions can induce apoptosis. Chronic ER stress has been implicated in a wide range of diseases, including autosomal dominant retinitis pigmentosa (ADRP), which is characterized by age-dependent retinal degeneration caused by mutant rhodopsin alleles. However, the signaling pathways that mediate apoptosis in response to ER stress remain poorly understood. In this study, we performed an unbiased in vivo RNAi screen with a Drosophila ADRP model and found that Wg/Wnt1 mediated apoptosis. Subsequent transcriptome analysis revealed that ER stress-associated serine protease (Erasp), which has been predicted to show serine-type endopeptidase activity, was a downstream target of Wg/Wnt1 during ER stress. Furthermore, knocking down Erasp via RNAi suppressed apoptosis induced by mutant rhodopsin-1 (Rh-1P37H) toxicity, alleviating retinal degeneration in the Drosophila ADRP model. In contrast, overexpression of Erasp resulted in enhanced caspase activity in Drosophila S2 cells treated with apoptotic inducers and the stabilization of the initiator caspase Dronc (Death regulator Nedd2-like caspase) by stimulating DIAP1 (Drosophila inhibitor of apoptosis protein 1) degradation. These findings helped identify a novel cell death signaling pathway involved in retinal degeneration in an autosomal dominant retinitis pigmentosa model.

Tctp, a unique Ing5-binding partner, inhibits the chromatin binding of Enok in Drosophila.

The MOZ/MORF histone acetyltransferase complex is highly conserved in eukaryotes and controls transcription, development, and tumorigenesis. However, little is known about how its chromatin localization is regulated. Inhibitor of growth 5 (ING5) tumor suppressor is a subunit of the MOZ/MORF complex. Nevertheless, the in vivo function of ING5 remains unclear. Here, we report an antagonistic interaction between Drosophila Translationally controlled tumor protein (TCTP) (Tctp) and ING5 (Ing5) required for chromatin localization of the MOZ/MORF (Enok) complex and H3K23 acetylation. Yeast two-hybrid screening using Tctp identified Ing5 as a unique binding partner. In vivo, Ing5 controlled differentiation and down-regulated epidermal growth factor receptor signaling, whereas it is required in the Yorkie (Yki) pathway to determine organ size. Ing5 and Enok mutants promoted tumor-like tissue overgrowth when combined with uncontrolled Yki activity. Tctp depletion rescued the abnormal phenotypes of the Ing5 mutation and increased the nuclear translocation of Ing5 and chromatin binding of Enok. Nonfunctional Enok promoted the nuclear translocation of Ing5 by reducing Tctp, indicating a feedback mechanism between Tctp, Ing5, and Enok to regulate histone acetylation. Therefore, Tctp is essential for H3K23 acetylation by controlling the nuclear translocation of Ing5 and chromatin localization of Enok, providing insights into the roles of human TCTP and ING5-MOZ/MORF in tumorigenesis.

Reduced branched-chain aminotransferase activity alleviates metabolic vulnerability caused by dim light exposure at night in Drosophila.

The rhythmic pattern of biological processes controlled by light over 24 h is termed the circadian rhythm. Disturbance of circadian rhythm due to exposure to light at night (LAN) disrupts the sleep-wake cycle and can promote cardiovascular disease, diabetes, cancer, and metabolic disorders in humans. We studied how dim LAN affects the circadian rhythm and metabolism using male Drosophila. Wild-type flies exposed to the dim light of 10 lux at night displayed altered 24 h sleep-wake behavior and expression patterns of circadian rhythm genes. In addition, the flies became more vulnerable to metabolic stress, such as starvation. Whole-body metabolite analysis revealed decreased amounts of branched-chain amino acids (BCAAs), such as isoleucine and valine. The dim light exposure also increased the expression of branched-chain amino acid aminotransferase (BCAT) and branched-chain α-keto acid dehydrogenase (BCKDC) enzyme complexes that regulate the metabolism of BCAAs. Flies with the Bcat heterozygous mutation were not vulnerable to starvation stress, even when exposed to dim LAN, and hemolymph BCAA levels did not decrease in these flies. Furthermore, the vulnerability to starvation stress was also suppressed when the Bcat expression level was reduced in the whole body, neurons, or fat body during adulthood using conditional GAL4 and RNA interference. Finally, the metabolic vulnerability was reversed when BCAAs were fed to wild-type flies exposed to LAN. Thus, short-term dim light exposure at night affects the expression of circadian genes and BCAA metabolism in Drosophila, implying a novel function of BCAAs in suppressing metabolic stress caused by disrupted circadian rhythm.

Metabolic control of daily locomotor activity mediated by tachykinin in Drosophila.

Metabolism influences locomotor behaviors, but the understanding of neural curcuit control for that is limited. Under standard light-dark cycles, Drosophila exhibits bimodal morning (M) and evening (E) locomotor activities that are controlled by clock neurons. Here, we showed that a high-nutrient diet progressively extended M activity but not E activity. Drosophila tachykinin (DTk) and Tachykinin-like receptor at 86C (TkR86C)-mediated signaling was required for the extension of M activity. DTk neurons were anatomically and functionally connected to the posterior dorsal neuron 1s (DN1ps) in the clock neuronal network. The activation of DTk neurons reduced intracellular Ca2+ levels in DN1ps suggesting an inhibitory connection. The contacts between DN1ps and DTk neurons increased gradually over time in flies fed a high-sucrose diet, consistent with the locomotor behavior. DN1ps have been implicated in integrating environmental sensory inputs (e.g., light and temperature) to control daily locomotor behavior. This study revealed that DN1ps also coordinated nutrient information through DTk signaling to shape daily locomotor behavior.

Identification and characterization of GAL4 drivers that mark distinct cell types and regions in the Drosophila adult gut.

The gastrointestinal tract in the adult Drosophila serves as a model system for exploring the mechanisms underlying digestion, absorption and excretion, stem cell plasticity, and inter-organ communication, particularly through the gut-brain axis. It is also useful for studying the cellular and adaptive responses to dietary changes, alterations in microbiota and immunity, and systematic and endocrine signals. Despite the various cell types and distinct regions in the gastrointestinal tract, few tools are available to target and manipulate the activity of each cell type and region, and their gene expression. Here, we report 353 GAL4 lines and several split-GAL4 lines that are expressed in enteric neurons (ENs), progenitors (ISCs and EBs), enterocytes (ECs), enteroendocrine cells (EEs), or/and other cell types that are yet to be identified in distinct regions of the gut. We had initially collected approximately 600 GAL4 lines that may be expressed in the gut based on RNA sequencing data, and then crossed them to UAS-GFP to perform immunohistochemistry to identify those that are expressed selectively in the gut. The cell types and regional expression patterns that are associated with the entire set of GAL4 drivers and split-GAL4 combinations are annotated online at http://kdrc.kr/index.php (K-Gut Project). This GAL4 resource can be used to target specific populations of distinct cell types in the fly gut, and therefore, should permit a more precise investigation of gut cells that regulate important biological processes.

A homeostatic sleep-stabilizing pathway in Drosophila composed of the sex peptide receptor and its ligand, the myoinhibitory peptide.

Sleep, a reversible quiescent state found in both invertebrate and vertebrate animals, disconnects animals from their environment and is highly regulated for coordination with wakeful activities, such as reproduction. The fruit fly, Drosophila melanogaster, has proven to be a valuable model for studying the regulation of sleep by circadian clock and homeostatic mechanisms. Here, we demonstrate that the sex peptide receptor (SPR) of Drosophila, known for its role in female reproduction, is also important in stabilizing sleep in both males and females. Mutants lacking either the SPR or its central ligand, myoinhibitory peptide (MIP), fall asleep normally, but have difficulty in maintaining a sleep-like state. Our analyses have mapped the SPR sleep function to pigment dispersing factor (pdf) neurons, an arousal center in the insect brain. MIP downregulates intracellular cAMP levels in pdf neurons through the SPR. MIP is released centrally before and during night-time sleep, when the sleep drive is elevated. Sleep deprivation during the night facilitates MIP secretion from specific brain neurons innervating pdf neurons. Moreover, flies lacking either SPR or MIP cannot recover sleep after the night-time sleep deprivation. These results delineate a central neuropeptide circuit that stabilizes the sleep state by feeding a slow-acting inhibitory input into the arousal system and plays an important role in sleep homeostasis.