Ribosome protein mutant cells rely on the GR64 cluster of gustatory receptors for survival and proteostasis in Drosophila

Mutations in ribosome protein (Rp) genes and ribosome biogenesis factors result in debilitating diseases, known as ribosomopathies. Recent studies in Drosophila have shown that cells heterozygous mutant for Rp genes (Rp/+) exhibit proteotoxic stress and aggregates, which drive stress pathway activation and apoptosis. Understanding how Rp/+ cells fend off proteotoxic stress could suggest mechanisms to ameliorate these and other conditions caused by proteotoxic stress. Here we find that Rp/+ epithelial cells express all six Gustatory Receptor 64 (Gr64) genes, a cluster of sugar receptors involved in taste sensation. We show that Rp/+ cells depend on Gr64 for survival and that loss of Gr64 autonomously exacerbates stress pathway activation and proteotoxic stress by negatively effecting autophagy and proteasome function in Rp/+ cells. This work identifies a non-canonical role in proteostasis maintenance for a family of gustatory receptors known for their function in neuronal sensation.

Drosophila local search emerges from iterative odometry of consecutive run lengths

SUMMARYThe ability to keep track of one’s location in space is a critical behavior for animals navigating to and from a salient location, but its computational basis remains unknown. Here, we tracked flies in a ring-shaped channel as they executed bouts of search, triggered by optogenetic activation of sugar receptors. Flies centered their back-and-forth local search excursions near fictive food locations by closely matching the length of consecutive runs. We tested a set of agent-based models that incorporate iterative odometry to store and retrieve the distance walked between consecutive events, such as reversals in walking direction. In contrast to memoryless models such as Lévy flight, simulations employing reversal-to-reversal integration recapitulated flies’ centered search behavior, even during epochs when the food stimulus was withheld or in experiments with multiple food sites. However, experiments in which flies reinitiated local search after circumnavigating the arena suggest that flies can also integrate azimuthal heading to perform path integration. Together, this work provides a concrete theoretical framework and experimental system to advance investigations of the neural basis of path integration.

Smelling in the dark: phylogenomic insights on the chemosensory system of a subterranean beetle

ABSTRACTThe chemosensory system has experienced relevant changes in subterranean animals, facilitating the orientation into darkness via the perception of specific chemical signals critical to survive in this particular environment. However, the genomic basis of chemoreception in cave-dwelling fauna is largely unexplored. We generated de novo transcriptomes for antennae and body samples of the troglobitic beetle Speonomus longicornis (whose characters suggest an extreme adaptation to the deep subterranean) in order to interrogate the evolutionary origin and diversification of the chemosensory gene repertoire across coleopterans through a phylogenomic approach. Our results suggested a diminished diversity of odorant and gustatory gene repertoires compared to polyphagous epigean beetles. Moreover, S. longicornis showed a large diversity of odorant-binding proteins, suggesting an important role of these proteins in capturing airborne chemical cues. We identified a gene duplication in the ionotropic co-receptor I R25a, a highly conserved single-copy gene in protostomes involved in thermal and humidity sensing. In addition, no homologous genes to sugar receptors or the ionotropic receptor IR41a were detected. Our findings suggest that the chemosensory gene repertoire of this cave beetle may have been reshaped by the low complexity of chemical signals of this particular environment, and that gene duplication and loss may have played an important role in the evolution of genes involved in chemoreception. Altogether, our results shed light on the genomic basis of chemoreception in a cave-dwelling invertebrate and pave the road towards understanding the genomic underpinnings of adaptation to the subterranean lifestyle at a deeper level.

Identification and expression profiles of candidate chemosensory receptors in Histia rhodope (Lepidoptera: Zygaenidae)

Insect olfaction and vision play important roles in survival and reproduction. Diurnal butterflies mainly rely on visual cues whereas nocturnal moths rely on olfactory signals to locate external resources. Histia rhodope Cramer (Lepidoptera: Zygaenidae) is an important pest of the landscape tree Bischofia polycarpa in China and other Southeast Asian regions. As a diurnal moth, H. rhodope represents a suitable model for studying the evolutionary shift from olfactory to visual communication. However, only a few chemosensory soluble proteins have been characterized and information on H. rhodope chemoreceptor genes is currently lacking. In this study, we identified 45 odorant receptors (ORs), nine ionotropic receptors (IRs), eight gustatory receptors (GRs) and two sensory neuron membrane proteins (SNMPs) from our previously acquired H. rhodope antennal transcriptomic data. The number of chemoreceptors of H. rhodope was less compared with that found in many nocturnal moths. Some specific chemoreceptors such as OR co-receptor (ORco), ionotropic receptors co-receptor, CO2 receptors, sugar receptors and bitter receptors were predicted by phylogenetic analysis. Notably, two candidate pheromone receptors (PRs) were identified within a novel PR lineage. qRT-PCR results showed that almost all tested genes (22/24) were predominantly expressed in antennae, indicating that they may be important in olfactory function. Among these antennae-enriched genes, six ORs, five IRs and two GRs displayed female-biased expression, while two ORs displayed male-biased expression. Additionally, HrhoIR75q.2 and HrhoGR67 were more highly expressed in heads and legs. This study enriches the olfactory gene inventory of H. rhodope and provides the foundation for further research of the chemoreception mechanism in diurnal moths.

CRISPR/Cas 9-Mediated Mutations as a New Tool for Studying Taste in Honeybees

Honeybees rely on nectar as their main source of carbohydrates. Sucrose, glucose, and fructose are the main components of plant nectars. Intriguingly, honeybees express only 3 putative sugar receptors (AmGr1, AmGr2, and AmGr3), which is in stark contrast to many other insects and vertebrates. The sugar receptors are only partially characterized. AmGr1 detects different sugars including sucrose and glucose. AmGr2 is assumed to act as a co-receptor only, while AmGr3 is assumedly a fructose receptor. We show that honeybee gustatory receptor AmGr3 is highly specialized for fructose perception when expressed in Xenopus oocytes. When we introduced nonsense mutations to the respective AmGr3 gene using CRISPR/Cas9 in eggs of female workers, the resulting mutants displayed almost a complete loss of responsiveness to fructose. In contrast, responses to sucrose were normal. Nonsense mutations introduced by CRISPR/Cas9 in honeybees can thus induce a measurable behavioral change and serve to characterize the function of taste receptors in vivo. CRISPR/Cas9 is an excellent novel tool for characterizing honeybee taste receptors in vivo. Biophysical receptor characterization in Xenopus oocytes and nonsense mutation of AmGr3 in honeybees unequivocally demonstrate that this receptor is highly specific for fructose.

CRISPR/Cas9 mediated mutations as a new tool for studying taste in honeybees

BackgroundHoneybees rely on nectar as their main source of carbohydrates [1]. Sucrose, glucose and fructose are the main components of plant nectars [2] [3]. Intriguingly, honeybees express only three putative sugar receptors (AmGr1, AmGr2 and AmGr3) [4], which is in stark contrast to many other insects and vertebrates. The sugar receptors are only partially characterized [5] [6]. AmGr1 detects different sugars including sucrose and glucose. AmGr2 is assumed to act as a co-receptor only, while AmGr3 is assumedly a fructose receptor.ResultsWe show that honeybee gustatory receptor AmGr3 is highly specialized for fructose perception when expressed in Xenopus oocytes. When we introduced nonsense mutations to the respective AmGr3 gene using CRISPR/Cas9 in eggs of female workers, the resulting mutants displayed almost a complete loss of responsiveness to fructose. In contrast, responses to sucrose were normal. Nonsense mutations introduced by CRISPR/Cas9 in honeybees can thus induce a measurable behavioural change and serve to characterize the function of taste receptors in vivo.ConclusionCRISPR/Cas9 is an excellent novel tool for characterizing honeybee taste receptors in vivo. Biophysical receptor characterisation in Xenopus oocytes and nonsense mutation of AmGr3 in honeybees unequivocally demonstrate that this receptor is highly specific for fructose.Graphical AbstractFigure 0

How altering the modular architecture affects aspects of lectin activity: case study on human galectin-1

Discoveries on involvement of glycan–protein recognition in many (patho)physiological processes are directing attention to exploring the significance of a fundamental structural aspect of sugar receptors beyond glycan specificity, i.e., occurrence of distinct types of modular architecture. In order to trace clues for defining design–functionality relationships in human lectins, a lectin's structural unit has been used as source material for engineering custom-made variants of the wild-type protein. Their availability facilitates comparative analysis toward the stated aim. With adhesion/growth-regulatory human galectin-1 as example, the strategy of evaluating how changes of its design (here, from the homodimer of non-covalently associated domains to (i) linker-connected di- and tetramers and (ii) a galectin-3-like protein) affect activity is illustrated by using three assay systems of increasing degree of glycan complexity. Whereas calorimetry with two cognate disaccharides and array testing with 647 (glyco)compounds disclosed no major changes, galectin histochemical staining profiles of tissue sections that present natural glycome complexity revealed differences between wild-type and linker-connected homo-oligomers as well as between the galectin-3-like variant and wild-type galectin-3 for cell-type positivity, level of intensity at the same site and susceptibility for inhibition by a bivalent glycocompound. These results underscore the strength of the documented approach. Moreover, they give direction to proceed to (i) extending its application to other members of this lectin family, especially galectin-3 and (ii) then analyzing impact of architectural alterations on cell surface lattice formation and ensuing biosignaling systematically, considering the variants’ potential for translational medicine.

Molecular characterization of sugar taste receptors in the cotton bollworm Helicoverpa armigera

Insects utilize sugars as their essential energy and nutrient sources; therefore, the sense of sugar detection plays a critical role in insect behaviours. Previously, using genomic and transcriptomic approaches, we identified eight putative sugar gustatory receptor (GR) genes from the cotton bollworm Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae). Here, we further validated these annotated sugar receptor genes (HarmGr4–HarmGr8 and HarmGr10–HarmGr12) and found HarmGr10 may be a pseudogene carrying a stop codon in the open reading frame. Sequence alignment revealed H. armigera sugar GR sequences are conserved at C-terminus and phylogenetic analysis showed that insect sugar GRs have evolved in a family-specific manner. Interestingly, all eight H. armigera sugar GRs are localized in a tandem array on the same scaffold of the genome. In silico gene expression and reverse transcription (RT)-PCR analysis showed that HarmGr10 is specifically expressed in male adult testes while HarmGr11 is specifically expressed in female adult ovaries, suggesting H. armigera sugar GRs may be involved in reproduction-related functions. This study improves our knowledge on insect sugar receptors and gustatory systems.