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4. An Expressed Sequence Tag collection from the male antennae of the Noctuid moth Spodoptera littoralis: a resource for olfactory and pheromone detection research

Author

Christelle Monsempes, Fabrice Legeai, Julie Poulain, Marie-Christine François, Christine Merlin, Nicolas Montagné, Frédérick Gavory, Sébastien Malpel, Martine Maïbèche-Coisne, Emmanuelle Jacquin-Joly, François Cousserans, Biologie des organismes et des populations appliquées à la protection des plantes (BIO3P), Institut National de la Recherche Agronomique (INRA)-Université de Rennes (UR)-AGROCAMPUS OUEST, Biological systems and models, bioinformatics and sequences (SYMBIOSE), Institut de Recherche en Informatique et Systèmes Aléatoires (IRISA), Université de Rennes (UR)-Institut National des Sciences Appliquées - Rennes (INSA Rennes), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de Recherche en Informatique et en Automatique (Inria)-Centre National de la Recherche Scientifique (CNRS)-Université de Rennes (UR)-Institut National des Sciences Appliquées - Rennes (INSA Rennes), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de Recherche en Informatique et en Automatique (Inria)-Centre National de la Recherche Scientifique (CNRS)-Inria Rennes – Bretagne Atlantique, Institut National de Recherche en Informatique et en Automatique (Inria), Développement et Communication Chimique chez les Insectes (DCCI), Université de Bourgogne (UB)-Centre National de la Recherche Scientifique (CNRS), Physiologie de l'Insecte : Signalisation et Communication (PISC), Institut National de la Recherche Agronomique (INRA)-Université Pierre et Marie Curie - Paris 6 (UPMC)-AgroParisTech, Biologie Intégrative et Virologie des Insectes [Univ. de Montpellier II] (BIVI), Institut National de la Recherche Agronomique (INRA)-Université Montpellier 2 - Sciences et Techniques (UM2), Genoscope - Centre national de séquençage [Evry] (GENOSCOPE), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Universite Pierre et Marie Curie (UPMC) ANR-07-NEURO-037-01 ANR-09-BLAN0239-01, Jacquin Joly, Emmanuelle, Institut National de la Recherche Agronomique (INRA)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-AGROCAMPUS OUEST, Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Institut National des Sciences Appliquées - Rennes (INSA Rennes), Institut National des Sciences Appliquées (INSA)-Université de Rennes (UNIV-RENNES)-Institut National des Sciences Appliquées (INSA)-Institut National de Recherche en Informatique et en Automatique (Inria)-Centre National de la Recherche Scientifique (CNRS)-Université de Rennes 1 (UR1), Institut National des Sciences Appliquées (INSA)-Université de Rennes (UNIV-RENNES)-Institut National des Sciences Appliquées (INSA)-Institut National de Recherche en Informatique et en Automatique (Inria)-Centre National de la Recherche Scientifique (CNRS)-Inria Rennes – Bretagne Atlantique, Centre des Sciences du Goût et de l'Alimentation [Dijon] (CSGA), Institut National de la Recherche Agronomique (INRA)-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Biologie des organismes et des populations appliquées à la protection des plantes ( BIO3P ), Institut National de la Recherche Agronomique ( INRA ) -Université de Rennes 1 ( UR1 ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -AGROCAMPUS OUEST, Biological systems and models, bioinformatics and sequences ( SYMBIOSE ), Institut de Recherche en Informatique et Systèmes Aléatoires ( IRISA ), Université de Rennes 1 ( UR1 ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Institut National des Sciences Appliquées - Rennes ( INSA Rennes ) -Institut National de Recherche en Informatique et en Automatique ( Inria ) -Centre National de la Recherche Scientifique ( CNRS ) -Université de Rennes 1 ( UR1 ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Institut National des Sciences Appliquées - Rennes ( INSA Rennes ) -Institut National de Recherche en Informatique et en Automatique ( Inria ) -Centre National de la Recherche Scientifique ( CNRS ) -Inria Rennes – Bretagne Atlantique, Institut National de Recherche en Informatique et en Automatique ( Inria ), Centre des Sciences du Goût et de l'Alimentation [Dijon] ( CSGA ), Institut National de la Recherche Agronomique ( INRA ) -Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Centre National de la Recherche Scientifique ( CNRS ), Développement et Communication Chimique chez les Insectes ( DCCI ), Centre National de la Recherche Scientifique ( CNRS ) -Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Physiologie de l'Insecte : Signalisation et Communication ( PISC ), Institut National de la Recherche Agronomique ( INRA ) -Université Pierre et Marie Curie - Paris 6 ( UPMC ) -AgroParisTech, Biologie Intégrative et Virologie des Insectes [Univ. de Montpellier II] ( BIVI ), Institut National de la Recherche Agronomique ( INRA ) -Université Montpellier 2 - Sciences et Techniques ( UM2 ), Genoscope - Centre national de séquençage [Evry] ( GENOSCOPE ), Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), and Université Montpellier 2 - Sciences et Techniques (UM2)-Institut National de la Recherche Agronomique (INRA)

Subjects
Male, MESH: Sequence Analysis, DNA, [ SDV.AEN ] Life Sciences [q-bio]/Food and Nutrition, MESH : Arthropod Antennae, détection olfactive, MESH : Molecular Sequence Annotation, Genes, Insect, Insect, MESH: Genes, Insect, MESH : Pheromones, Pheromones, 0302 clinical medicine, Arthropod Antennae, Databases, Genetic, MESH: Smell, MESH: Animals, MESH: Phylogeny, Phylogeny, MESH: Databases, Genetic, media_common, Expressed Sequence Tags, 0303 health sciences, Expressed sequence tag, MESH: Pheromones, noctuelle, MESH: Spodoptera, MESH : Expressed Sequence Tags, MESH : Genes, Insect, food and beverages, Smell, Sex pheromone, Pheromone, Biotechnology, Research Article, papillon de nuit, lcsh:QH426-470, MESH : Male, lcsh:Biotechnology, media_common.quotation_subject, Gene prediction, Olfaction, Computational biology, MESH: Molecular Sequence Annotation, Biology, Spodoptera, MESH : Spodoptera, MESH: Expressed Sequence Tags, 03 medical and health sciences, MESH: Gene Expression Profiling, lcsh:TP248.13-248.65, Botany, MESH: Gene Library, Genetics, Animals, MESH : Databases, Genetic, Spodoptera littoralis, 030304 developmental biology, Gene Library, MESH: Arthropod Antennae, MESH : Gene Expression Profiling, Gene Expression Profiling, séquence génomique, fungi, MESH : Phylogeny, Molecular Sequence Annotation, Sequence Analysis, DNA, MESH : Gene Library, biology.organism_classification, MESH: Male, lcsh:Genetics, MESH : Smell, MESH : Animals, [SDV.AEN]Life Sciences [q-bio]/Food and Nutrition, 030217 neurology & neurosurgery, MESH : Sequence Analysis, DNA
Abstract

Background Nocturnal insects such as moths are ideal models to study the molecular bases of olfaction that they use, among examples, for the detection of mating partners and host plants. Knowing how an odour generates a neuronal signal in insect antennae is crucial for understanding the physiological bases of olfaction, and also could lead to the identification of original targets for the development of olfactory-based control strategies against herbivorous moth pests. Here, we describe an Expressed Sequence Tag (EST) project to characterize the antennal transcriptome of the noctuid pest model, Spodoptera littoralis, and to identify candidate genes involved in odour/pheromone detection. Results By targeting cDNAs from male antennae, we biased gene discovery towards genes potentially involved in male olfaction, including pheromone reception. A total of 20760 ESTs were obtained from a normalized library and were assembled in 9033 unigenes. 6530 were annotated based on BLAST analyses and gene prediction software identified 6738 ORFs. The unigenes were compared to the Bombyx mori proteome and to ESTs derived from Lepidoptera transcriptome projects. We identified a large number of candidate genes involved in odour and pheromone detection and turnover, including 31 candidate chemosensory receptor genes, but also genes potentially involved in olfactory modulation. Conclusions Our project has generated a large collection of antennal transcripts from a Lepidoptera. The normalization process, allowing enrichment in low abundant genes, proved to be particularly relevant to identify chemosensory receptors in a species for which no genomic data are available. Our results also suggest that olfactory modulation can take place at the level of the antennae itself. These EST resources will be invaluable for exploring the mechanisms of olfaction and pheromone detection in S. littoralis, and for ultimately identifying original targets to fight against moth herbivorous pests.

Published
2011
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5. A Role for Blind DN2 Clock Neurons in Temperature Entrainment of the Drosophila Larval Brain

Author

François Rouyer, Sébastien Malpel, André Klarsfeld, Elisabeth Chélot, Marie Picot, Neurobiologie génétique et intégrative (NGI), Centre National de la Recherche Scientifique (CNRS), Institut de Neurobiologie Alfred Fessard (INAF), Développement et Communication Chimique chez les Insectes (DCCI), Centre National de la Recherche Scientifique (CNRS)-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Centre des Sciences du Goût et de l'Alimentation [Dijon] (CSGA), Institut National de la Recherche Agronomique (INRA)-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Centre National de la Recherche Scientifique (CNRS), Neurobiologie génétique et intégrative ( NGI ), Centre National de la Recherche Scientifique ( CNRS ), Institut de Neurobiologie Alfred Fessard ( INAF ), Centre des Sciences du Goût et de l'Alimentation [Dijon] ( CSGA ), Institut National de la Recherche Agronomique ( INRA ) -Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Centre National de la Recherche Scientifique ( CNRS ), Développement et Communication Chimique chez les Insectes ( DCCI ), and Centre National de la Recherche Scientifique ( CNRS ) -Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement

Subjects
Time Factors, Light, Green Fluorescent Proteins, Circadian clock, Neuropeptide, Biology, Receptors, G-Protein-Coupled, Animals, Genetically Modified, 03 medical and health sciences, 0302 clinical medicine, Rhythm, Cryptochrome, Biological Clocks, Biologie animale, Animals, Drosophila Proteins, Eye Proteins, 030304 developmental biology, Neurons, Animal biology, Analysis of Variance, 0303 health sciences, Larva, Communication, Behavior, Animal, business.industry, General Neuroscience, [SDV.BA]Life Sciences [q-bio]/Animal biology, Neuropeptides, fungi, Temperature, Neurosciences, Brain, Articles, Clock network, Cryptochromes, nervous system, Brain Hemisphere, Neurons and Cognition, Drosophila, [SDV.NEU]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC], Entrainment (chronobiology), business, Neuroscience, 030217 neurology & neurosurgery, Signal Transduction
Abstract

International audience; Circadian clocks synchronize to the solar day by sensing the diurnal changes in light and temperature. In adult Drosophila, the brain clock that controls rest-activity rhythms relies on neurons showing Period oscillations. Nine of these neurons are present in each larval brain hemisphere. They can receive light inputs through Cryptochrome (CRY) and the visual system, but temperature input pathways are unknown. Here, we investigate how the larval clock network responds to light and temperature. We focused on the CRY-negative dorsal neurons (DN2s), in which light-dark (LD) cycles set molecular oscillations almost in antiphase to all other clock neurons. We first showed that the phasing of the DN2s in LD depends on the pigment-dispersing factor (PDF) neuropeptide in four lateral neurons (LNs), and on the PDF receptor in the DN2s. In the absence of PDF signaling, these cells appear blind, but still synchronize to temperature cycles. Period oscillations in the DN2s were stronger in thermocycles than in LD, but with a very similar phase. Conversely, the oscillations of LNs were weaker in thermocycles than in LD, and were phase-shifted in synchrony with the DN2s, whereas the phase of the three other clock neurons was advanced by a few hours. In the absence of any other functional clock neurons, the PDF-positive LNs were entrained by LD cycles but not by temperature cycles. Our results show that the larval clock neurons respond very differently to light and temperature, and strongly suggest that the CRY-negative DN2s play a prominent role in the temperature entrainment of the network.

Published
2009
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6. Molecular identification and characterization of two new Lepidoptera chemoreceptors belonging to the Drosophila melanogaster OR83b family

Author

Emmanuelle Jacquin-Joly, Christine Merlin, Marie-Christine François, Sébastien Malpel, Développement et Communication Chimique chez les Insectes ( DCCI ), Centre National de la Recherche Scientifique ( CNRS ) -Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Développement et Communication Chimique chez les Insectes (DCCI), and Centre National de la Recherche Scientifique (CNRS)-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement

Subjects
Male, Subfamily, olfactory receptor, [SDV]Life Sciences [q-bio], media_common.quotation_subject, Molecular Sequence Data, Olfaction, Insect, Receptors, Odorant, Lepidoptera genitalia, Botany, Genetics, medicine, Melanogaster, Animals, Drosophila Proteins, Amino Acid Sequence, Spodoptera littoralis, Molecular Biology, media_common, Olfactory receptor, biology, Sequence Homology, Amino Acid, Reverse Transcriptase Polymerase Chain Reaction, fungi, Animal Structures, Gene Expression Regulation, Developmental, biology.organism_classification, Protein Structure, Tertiary, Lepidoptera, medicine.anatomical_structure, Drosophila melanogaster, Insect Science, Female, Mamestra brassicae, Sequence Alignment, olfaction
Abstract

1365-2583 10.1111/j.1365-2583.2008.00830.x; In insect antennae, olfaction depends on olfactory receptors (ORs) that function through heterodimerization with an unusually highly conserved partner orthologue to the Drosophila melanogaster DOR83b. Here, we report the identification of two cDNAs encoding new DOR83b orthologues that represent the first members, although nonconventional, of the OR families of two noctuid crop pests, the cotton leafworm Spodoptera littoralis and the cabbage armyworm Mamestra brassicae. They both displayed high protein sequence conservation with previously identified DOR83b orthologues. Transcripts were abundantly detected in adult chemosensory organs as well as in fifth instar larvae heads. In adult antennae, the expression patterns of both genes revealed common features with other members of the OR83b subfamily: they appeared to be expressed at the bases of numerous olfactory sensilla belonging to different functional categories, suggesting that both receptors may be co-expressed with yet unidentified conventional ORs. Bioinformatic analyses predicted the occurrence of seven transmembrane domains and an unusual topology with intracellular N-termini and extracellular C-termini, extending to Lepidoptera the hypothesis of an inverted topology for DOR83b orthologues, demonstrated to date only in D.†melanogaster.

Published
2008
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7. Novel Features of Cryptochrome-Mediated Photoreception in the Brain Circadian Clock of Drosophila

Author

Sébastien Malpel, Elisabeth Chélot, François Rouyer, André Klarsfeld, Christine Michard-Vanhée, Marie Picot, Neurobiologie génétique et intégrative (NGI), Centre National de la Recherche Scientifique (CNRS), and Institut de Neurobiologie Alfred Fessard (INAF)

Subjects
Light, MESH: Drosophila, Circadian clock, circadian photoreception, MESH: Neurons, PERIOD protein, MESH: Neuropeptides, Receptors, G-Protein-Coupled, 0302 clinical medicine, MESH: Eye Proteins, Cryptochrome, Drosophila Proteins, MESH: Animals, Photopigment, Neurons, 0303 health sciences, dorsal neurons, MESH: Photoreceptors, Invertebrate, Behavior, Animal, General Neuroscience, MESH: Darkness, Brain, Nuclear Proteins, MESH: Behavior,Biological Clocks, Period Circadian Proteins, Darkness, MESH: Motor Activity, Circadian Rhythm, DNA-Binding Proteins, medicine.anatomical_structure, Larva, [SDV.NEU]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC], Drosophila, Female, Photoreceptor Cells, Invertebrate, cryptochrome, Visual phototransduction, Cellular/Molecular, Locomotor activity rhythms, endocrine system, MESH: Mutation, MESH: Drosophila Proteins, Period (gene), Photoperiod, GLASS protein, Biology, Motor Activity, MESH: Photoperiod, MESH: Brain, 03 medical and health sciences, Pigment dispersing factor, Biological Clocks, medicine, Animals, MESH: Circadian Rhythm, Circadian rhythm, Eye Proteins, 030304 developmental biology, Neuropeptides, MESH: Light, Cryptochromes, pigment-dispersing factor, Mutation, MESH: Larva, MESH: Nuclear Proteins, MESH: Female, Neuroscience, Nucleus, MESH: DNA-Binding Proteins, 030217 neurology & neurosurgery
Abstract

InDrosophila, light affects circadian behavioral rhythms via at least two distinct mechanisms. One of them relies on the visual phototransduction cascade. The other involves a presumptive photopigment, cryptochrome (cry), expressed in lateral brain neurons that control behavioral rhythms. We show here thatcryis expressed in most, if not all, larval and adult neuronal groups expressing the PERIOD (PER) protein, with the notable exception of larval dorsal neurons (DN2s) in which PER cycles in antiphase to all other known cells. Forcingcryexpression in the larval DN2s gave them a normal phase of PER cycling, indicating that their unique antiphase rhythm is related to their lack ofcryexpression. We were able to directly monitor CRY protein inDrosophilabrainsin situ. It appeared highly unstable in the light, whereas in the dark, it accumulated in both the nucleus and the cytoplasm, including some neuritic projections. We also show that dorsal PER-expressing brain neurons, the adult DN1s, are the only brain neurons to coexpress the CRY protein and the photoreceptor differentiation factor GLASS. Studies of various visual system mutants and their combination with thecrybmutation indicated that the adult DN1s contribute significantly to the light sensitivity of the clock controlling activity rhythms, and that this contribution depends on CRY. Moreover, all CRY-independent light inputs into this central behavioral clock were found to require the visual system. Finally, we show that the photoreceptive DN1 neurons do not behave as autonomous oscillators, because their PER oscillations in constant darkness rapidly damp out in the absence of pigment-dispersing-factor signaling from the ventral lateral neurons.

Published
2004
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8. Larval optic nerve and adult extra-retinal photoreceptors sequentially associate with clock neurons during Drosophila brain development

Author

François Rouyer, Sébastien Malpel, André Klarsfeld, Neurobiologie génétique et intégrative (NGI), Centre National de la Recherche Scientifique (CNRS), and Institut de Neurobiologie Alfred Fessard (INAF)

Subjects
MESH: Drosophila, Circadian clock, MESH: Neurons, Phospholipase C beta, norpA, 0302 clinical medicine, Drosophila Proteins, MESH: Animals, Neurons, 0303 health sciences, Larva, MESH: Photoreceptors, Invertebrate, MESH: Rhodopsin, Brain, Cell Differentiation, Anatomy, Bolwig organ, Isoenzymes, MESH: Isoenzymes, MESH: Phospholipase C, Optic nerve, [SDV.NEU]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC], Drosophila, Photoreceptor Cells, Invertebrate, MESH: Cell Differentiation, Rhodopsin, Brain development, MESH: Drosophila Proteins, Biology, MESH: Brain, 03 medical and health sciences, Animals, Rhodopsins, Circadian rhythm, Drosophila (subgenus), Molecular Biology, 030304 developmental biology, Nerve activity, Optic Nerve, MESH: Optic Nerve, biology.organism_classification, Retinal Photoreceptors, Type C Phospholipases, Dendritic tree, Hofbauer-Buchner eyelet, MESH: Larva, Neuroscience, 030217 neurology & neurosurgery, Developmental Biology
Abstract

The visual system is one of the input pathways for light into the circadian clock of the Drosophila brain. In particular, extra-retinal visual structures have been proposed to play a role in both larval and adult circadian photoreception. We have analyzed the interactions between extra-retinal structures of the visual system and the clock neurons during brain development. We first show that the larval optic nerve, or Bolwig nerve, already contacts clock cells (the lateral neurons) in the embryonic brain. Analysis of visual system-defective genotypes showed that the absence of the afferent Bolwig nerve resulted in a severe reduction of the lateral neurons dendritic arborization, and that the inhibition of nerve activity induced alterations of the dendritic morphology. During wild-type development, the loss of a functional Bolwig nerve in the early pupa was also accompanied by remodeling of the arborization of the lateral neurons. Approximately 1.5 days later, visual fibers that came from the Hofbauer-Buchner eyelet, a putative photoreceptive organ for the adult circadian clock, were seen contacting the lateral neurons. Both types of extra-retinal photoreceptors expressed rhodopsins RH5 and RH6, as well as the norpA-encoded phospholipase C. These data strongly suggest a role for RH5 and RH6, as well as NORPA, signaling in both larval and adult extra-retinal circadian photoreception. The Hofbauer-Buchner eyelet therefore does not appear to account for the previously described norpA-independent light input to the adult clock. This supports the existence of yet uncharacterized photoreceptive structures in Drosophila.

Published
2002

9. The last 59 amino acids of Smoothened cytoplasmic tail directly bind the protein kinase Fused and negatively regulate the Hedgehog pathway

Author

Sébastien Malpel, Matthieu Sanial, Laurent Daviet, Séverine Martin-Lannerée, Tristan Piolot, Anne Plessis, Sandra Claret, Amira Brigui, Institut Jacques Monod (IJM (UMR_7592)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Biologie systémique (BS (IFR_117)), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Diderot - Paris 7 (UPD7), Hybrigenics [Paris], and Hybrigenics

Subjects
Biology, Protein Serine-Threonine Kinases, Drosophila development, Models, Biological, Cell Line, Receptors, G-Protein-Coupled, 03 medical and health sciences, 0302 clinical medicine, GPCR, Two-Hybrid System Techniques, Animals, Drosophila Proteins, Wings, Animal, Fluorescent imaging, Hedgehog Proteins, Protein kinase A, Hedgehog, Molecular Biology, [SDV.BDD]Life Sciences [q-bio]/Development Biology, 030304 developmental biology, G protein-coupled receptor, DNA Primers, Smoothened, 0303 health sciences, Activator (genetics), Two hybrid, Fused, Cell Biology, Smoothened Receptor, Imaginal disc, Transmembrane protein, Hedgehog signaling pathway, Signaling, Clone 8 cells, Cell biology, Biochemistry, Gene Expression Regulation, Microscopy, Fluorescence, Drosophila, Signal transduction, 030217 neurology & neurosurgery, Protein Binding, Signal Transduction, Developmental Biology
Abstract

International audience; The Hedgehog (HH) signaling pathway is crucial for the development of many organisms and its inappropriate activation is involved in numerous cancers. HH signal controls the traffic and activity of the seven-pass transmembrane protein Smoothened (SMO), leading to the transcriptional regulation of HH-responsive genes. In Drosophila, the intracellular transduction events following SMO activation depend on cytoplasmic multimeric complexes that include the Fused (FU) protein kinase. Here we show that the regulatory domain of FU physically interacts with the last 52 amino acids of SMO and that the two proteins colocalize in vivo to vesicles. The deletion of this region of SMO leads to a constitutive activation of SMO, promoting the ectopic transcription of HH target genes. This activation is partially dependent of FU activity. Thus, we identify a novel link between SMO and the cytoplasmic complex(es) and reveal a negative role of the SMO C-terminal region that interacts with FU. We propose that FU could act as a switch, activator in presence of HH signal or inhibitor in absence of HH.

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