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      De novo assembly of a transcriptome for the cricket Gryllus bimaculatus prothoracic ganglion: An invertebrate model for investigating adult central nervous system compensatory plasticity

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          Abstract

          The auditory system of the cricket, Gryllus bimaculatus, demonstrates an unusual amount of anatomical plasticity in response to injury, even in adults. Unilateral removal of the ear causes deafferented auditory neurons in the prothoracic ganglion to sprout dendrites across the midline, a boundary they typically respect, and become synaptically connected to the auditory afferents of the contralateral ear. The molecular basis of this sprouting and novel synaptogenesis in the adult is not understood. We hypothesize that well-conserved developmental guidance cues may recapitulate their guidance functions in the adult in order to facilitate this compensatory growth. As a first step in testing this hypothesis, we have generated a de novo assembly of a prothoracic ganglion transcriptome derived from control and deafferented adult individuals. We have mined this transcriptome for orthologues of guidance molecules from four well-conserved signaling families: Slit, Netrin, Ephrin, and Semaphorin. Here we report that transcripts encoding putative orthologues of most of the candidate developmental ligands and receptors from these signaling families were present in the assembly, indicating expression in the adult G. bimaculatus prothoracic ganglion.

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          Slit proteins bind Robo receptors and have an evolutionarily conserved role in repulsive axon guidance.

          Extending axons in the developing nervous system are guided in part by repulsive cues. Genetic analysis in Drosophila, reported in a companion to this paper, identifies the Slit protein as a candidate ligand for the repulsive guidance receptor Roundabout (Robo). Here we describe the characterization of three mammalian Slit homologs and show that the Drosophila Slit protein and at least one of the mammalian Slit proteins, Slit2, are proteolytically processed and show specific, high-affinity binding to Robo proteins. Furthermore, recombinant Slit2 can repel embryonic spinal motor axons in cell culture. These results support the hypothesis that Slit proteins have an evolutionarily conserved role in axon guidance as repulsive ligands for Robo receptors.
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            Roundabout controls axon crossing of the CNS midline and defines a novel subfamily of evolutionarily conserved guidance receptors.

            The robo gene in Drosophila was identified in a large-scale mutant screen for genes that control the decision by axons to cross the CNS midline. In robo mutants, too many axons cross and recross the midline. Here we show that robo encodes an axon guidance receptor that defines a novel subfamily of immunoglobulin superfamily proteins that is highly conserved from fruit flies to mammals. For those axons that never cross the midline, Robo is expressed on their growth cones from the outset; for the majority of axons that do cross the midline, Robo is expressed at high levels on their growth cones only after they cross the midline. Transgenic rescue experiments reveal that Robo can function in a cell-autonomous fashion. Robo appears to function as the gatekeeper controlling midline crossing.
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              Slit is the midline repellent for the robo receptor in Drosophila.

              Previous studies suggested that Roundabout (Robo) is a repulsive guidance receptor on growth cones that binds to an unknown midline ligand. Here we present genetic evidence that Slit is the midline Robo ligand; a companion paper presents biochemical evidence that Slit binds Robo. Slit is a large extracellular matrix protein expressed by midline glia. In slit mutants, growth cones enter the midline but never leave it; they abnormally continue to express high levels of Robo while at the midline. slit and robo display dosage-sensitive genetic interactions, indicating that they function in the same pathway. slit is also required for migration of muscle precursors away from the midline. Slit appears to function as a short-range repellent controlling axon crossing of the midline and as a long-range chemorepellent controlling mesoderm migration away from the midline.
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                Author and article information

                Contributors
                Role: ConceptualizationRole: MethodologyRole: Project administrationRole: ValidationRole: Writing – original draft
                Role: Data curationRole: MethodologyRole: Project administration
                Role: Formal analysisRole: ValidationRole: Writing – review & editing
                Role: ValidationRole: Writing – review & editing
                Role: ValidationRole: Writing – review & editing
                Role: ValidationRole: Writing – review & editing
                Role: ConceptualizationRole: Data curationRole: Formal analysisRole: MethodologyRole: Project administrationRole: ResourcesRole: SupervisionRole: Writing – original draftRole: Writing – review & editing
                Role: ConceptualizationRole: Formal analysisRole: MethodologyRole: Project administrationRole: ResourcesRole: Writing – original draft
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                11 July 2018
                2018
                : 13
                : 7
                : e0199070
                Affiliations
                [1 ] Department of Biology, Bowdoin College, Brunswick, Maine, United States of America
                [2 ] Békésy Laboratory of Neurobiology, Pacific Biosciences Research Center, School of Ocean and Earth Science and Technology, University of Hawaii at Manoa, Honolulu, Hawaii, United States of America
                Federal University of Rio de Janeiro, BRAZIL
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Author information
                http://orcid.org/0000-0001-6867-6865
                Article
                PONE-D-17-33294
                10.1371/journal.pone.0199070
                6040699
                29995882
                773886da-1a16-4a09-8c2e-083fe0b48e49
                © 2018 Fisher et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 12 September 2017
                : 25 May 2018
                Page count
                Figures: 6, Tables: 4, Pages: 28
                Funding
                Funded by: funder-id http://dx.doi.org/10.13039/100000057, National Institute of General Medical Sciences;
                Award ID: P20GM0103423
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/100000154, Division of Integrative Organismal Systems;
                Award ID: IOS-1353023
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/100008889, Cades Foundation;
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/100008783, University of Hawai'i at Mānoa;
                Award ID: Undergraduate Research Opportunities Program
                Award Recipient :
                Research reported in this publication was supported by an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under grant number P20GM0103423.
                Categories
                Research Article
                Biology and Life Sciences
                Biochemistry
                Proteins
                Protein Domains
                Research and Analysis Methods
                Experimental Organism Systems
                Model Organisms
                Drosophila Melanogaster
                Research and Analysis Methods
                Model Organisms
                Drosophila Melanogaster
                Research and Analysis Methods
                Experimental Organism Systems
                Animal Models
                Drosophila Melanogaster
                Biology and Life Sciences
                Organisms
                Eukaryota
                Animals
                Invertebrates
                Arthropoda
                Insects
                Drosophila
                Drosophila Melanogaster
                Biology and Life Sciences
                Computational Biology
                Genome Analysis
                Transcriptome Analysis
                Biology and Life Sciences
                Genetics
                Genomics
                Genome Analysis
                Transcriptome Analysis
                Biology and Life Sciences
                Anatomy
                Biological Tissue
                Ganglia
                Medicine and Health Sciences
                Anatomy
                Biological Tissue
                Ganglia
                Biology and Life Sciences
                Organisms
                Eukaryota
                Animals
                Invertebrates
                Arthropoda
                Insects
                Crickets
                Research and Analysis Methods
                Database and Informatics Methods
                Database Searching
                Sequence Similarity Searching
                Biology and Life Sciences
                Cell Biology
                Cellular Types
                Animal Cells
                Neurons
                Neuronal Dendrites
                Biology and Life Sciences
                Neuroscience
                Cellular Neuroscience
                Neurons
                Neuronal Dendrites
                Biology and Life Sciences
                Neuroscience
                Cellular Neuroscience
                Neuronal Plasticity
                Custom metadata
                Much of the data discussed and mined from the transcriptome is included in the first supplemental figure. The complete, quality filtered dataset (509,779 of the original 511,724 contigs), is available at the NCBI under BioProject No. PRJNA376023.

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