Fig 6 - uploaded by Rolf O Karlstrom
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Lachesin is linked to the membrane through a glycosyl phosphatidylinositol anchor. (A) 1C10 mAb labelling in one segment of an untreated embryo at 40% of development. Axons of the commissures, longitudinal connectives and median fiber tract as well as nerve cell bodies are labelled. (B) Labelling in an embryo treated with 1.5 U/ml PI-PLC for 2 hours. Lachesin is no longer seen on the axons or cell bodies of the CNS. (C) Western immunoblot of PI-PLC experiment. Left to right: Standard membrane preparation (M). Membrane preparation (M) and supernatant (S) from 10 embryos cultured 2 hours in the absence of PI-PLC. Membrane preparation (M) and supernatant (S) from 10 embryos cultured 2 hours in the presence of PI-PLC. All Lachesin has been removed from the membranes of PI- PLC-treated embryos and is present in the supernatant. The change in protein mobility on a polyacrylamide gel is consistent with the removal of a glycosyl phosphatidyl inositol moiety (see text). Scale bar: 40 µm .
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We describe the developmental expression in grasshopper (Schistocerca americana) and molecular characterization in grasshopper and fruit fly (Drosophila melanogaster) of Lachesin, a novel immunoglobulin superfamily protein. Lachesin is expressed on the surfaces of differentiating neuronal cells from the onset of neurogenesis in both the central and...
Citations
... For double labeling with α-PH3, the primary HRP antibody was polyclonal in goat (Dianova, 123-005-021). Anti-Lachesin (α-Lach, Mab 1C10, mouse, generous gift of M. Bastiani) recognizes the glycosylphosphatidylinositol (GPI)-linked cell surface molecule Lachesin belonging to the Ig superfamily (see Karlstrom et al. 1993). The expression occurs initially on all differentiating neuroepithelial cells, but only neurons and cells involved in neurogenesis, such as precursors, continue to express the molecule later. ...
The antennal flagellum of the locust S. gregaria is an articulated structure bearing a spectrum of sensilla that responds to sensory stimuli. In this study, we focus on the basiconic-type bristles as a model for sensory system development in the antenna. At the end of embryogenesis, these bristles are found at fixed locations and then on only the most distal six articulations of the antenna. They are innervated by a dendrite from a sensory cell cluster in the underlying epithelium, with each cluster directing fused axons topographically to an antennal tract running to the brain. We employ confocal imaging and immunolabeling to (a) identify mitotically active sense organ precursors for sensory cell clusters in the most distal annuli of the early embryonic antenna; (b) observe the subsequent spatial appearance of their neuronal progeny; and (c) map the spatial and temporal organization of axon projections from such clusters into the antennal tracts. We show that early in embryogenesis, proliferative precursors are localized circumferentially within discrete epithelial domains of the flagellum. Progeny first appear distally at the antennal tip and then sequentially in a proximal direction so that sensory neuron populations are distributed in an age-dependent manner along the antenna. Autotracing reveals that axon fasciculation with a tract is also sequential and reflects the location and age of the cell cluster along the most distal annuli. Cell cluster location and bristle location are therefore represented topographically and temporally within the axon profile of the tract and its projection to the brain.
... anti-Lachesin (α-Lach, Mab 1C10, mouse, generous gift of M. Bastiani) recognizes the glycosylphosphatidylinositol (GPI)-linked cell surface molecule Lachesin belonging to the Ig superfamily (see Karlstrom et al. 1993). The expression occurs initially on all differentiating epithelial cells, but only neurons and cells involved in neurogenesis, such as precursors, continue to express the molecule later. ...
The antennal flagellum of the locust S. gregaria is an articulated structure bearing a spectrum of sensory sensilla that encode environmental stimuli mediating adaptive behavior. In this study we focus on the basiconic-type bristles as a model for sensory system development in the antenna. At the end of embryogenesis these bristles are found at conserved locations on each apical segment of the flagellum, are innervated by a dendrite from a sensory cell cluster in the underlying epithelium with each cluster directing an axon topographically to an antennal tract and the brain. We employ confocal imaging and immunolabeling to (a) identify mitotically active sense organ precursors for sensory cell clusters in the most apical annuli of the early embryonic antenna; (b) follow the subsequent differentiation of their neuronal progeny; and (c) map the spatial and temporal organization of axonal projections into the antennal tracts. We show that early in embryogenesis proliferative precursors are localized circumferentially within discrete epithelial domains of the flagellum. Differentiation of their clonal progeny begins apically and proceeds in a basal direction along the antenna so that the oldest neuronal populations are near the antennal tip with progressively younger populations located more basally. Autotracing reveals that axon fasciculation with a tract is sequential and reflects the location and age of the cell cluster along the flagellum. Cell cluster location and bristle location on the cuticle are therefore encoded topographically and temporally within the axon profile of the tract and its projection to the brain.
... Anti-horseradish peroxidase (α-HRP, polyclonal rabbit, and Dianova) recognizes a neuron-specific epitope in insects (see Jan and Jan 1982). Anti-Lachesin (Mab 1C10, mouse, and gift of M. Bastiani) recognizes a GPI-linked cell surface molecule belonging to the Ig superfamily (see Karlstrom et al. 1993). The expression occurs initially on all differentiating epithelial cells, but only cells involved in neurogenesis, such as precursors continue to express the molecule later. ...
... The scape, pedicel, and flagellum of the locust antenna represent true segments, and their development is regulated by molecular mechanisms homologous to those forming other head and body appendages (see Gibson and Gehring 1988;Casares and Mann 1998). Accordingly, antibody labeling shows that common patterns of cell surface epitopes such as Annulin (Bastiani et al. 1992;Boyan et al. 2018) and Lachesin (Karlstrom et al. 1993;Boyan and Ehrhardt 2020) are present in their epithelia. Lachesin expression in the embryonic pedicel takes the form of a single stripe (Fig. 2) within which putative sense-organ precursors (SOPs) generating the cell clusters of the JO are located (Figs. 2d and 3). ...
Johnston’s organ has been shown to act as an antennal auditory organ across a spectrum of insect species. In the hemimetabolous desert locust Schistocerca gregaria, Johnston’s organ must be functional on hatching and so develops in the pedicellar segment of the antenna during embryogenesis. Here, we employ the epithelial cell marker Lachesin to identify the pedicellar domain of the early embryonic antenna and then triple-label against Lachesin, the mitosis marker phosphohistone-3, and neuron-specific horseradish peroxidase to reveal the sense-organ precursors for Johnston’s organ and their lineages. Beginning with a single progenitor at approximately a third of embryogenesis, additional precursors subsequently appear in both the ventral and dorsal pedicellar domains, each generating a lineage or clone. Lineage locations are remarkably conserved across preparations and ages, consistent with the epithelium possessing an underlying topographic coordinate system that determines the cellular organization of Johnston’s organ. By mid-embryogenesis, twelve lineages are arranged circumferentially in the pedicel as in the adult structure. Each sense-organ precursor is associated with a smaller mitotically active cell from which the neuronal complement of each clone may derive. Neuron numbers within a clone increase in discrete steps with age and are invariant between clones and across preparations of a given age. At mid-embryogenesis, each clone comprises five cells consolidated into a tightly bound cartridge. A long scolopale extends apically from each cartridge to an insertion point in the epithelium, and bundled axons project basally toward the brain. Comparative data suggest mechanisms that might also regulate the developmental program of Johnston’s organ in the locust.
... Protocols for immunolabeling with primary and secondary antibodies, the composition of incubation media, incubation conditions, confocal, uorescence and Nomarksi microscopy, and image processing were all as previously described (see Boyan Primary antibodies anti-Horseradish peroxidase (α-HRP, polyclonal rabbit, Dianova) recognizes a neuron-speci c epitope in insects (see Jan and Jan 1982). anti-Lachesin (Mab 1C10, mouse, gift of M. Bastiani) recognizes a GPIlinked cell surface molecule belonging to the Ig superfamily (see Karlstrom et al. 1993). Expression occurs initially on all differentiating epithelial cells but only cells involved in neurogenesis such as precursors continue to express the molecule later. ...
... Topographic organization of the pedicellar epithelial domain The scape, pedicel and agellum of the locust antenna represent true segments and their development is regulated by molecular mechanisms homologous to those forming other head and body appendages (see Gibson and Gehring 1988; Casares and Mann 1998). Accordingly their epithelia express conserved expression patterns of cell surface factors such as Annulin (Bastiani et al. 1992; Boyan et al. 2018) andLachesin(Karlstrom et al. 1993;Boyan and Ehrhardt 2020). Lachesin expression in the embryonic pedicel takes the form of a single stripe(Fig. ...
Johnston´s organ has been shown to act as an antennal auditory organ across a spectrum of insect species. In the hemimetabolous desert locust Schistocerca gregaria , Johnston´s organ must be functional on hatching and so develops in the pedicellar segment of the antenna during embryogenesis. Here we employ the epithelial cell marker Lachesin to identify the pedicellar domain of the early embryonic antenna, and then triple-label against Lachesin, the mitosis marker phosphohistone-3, and neuron-specific horseradish peroxidase to reveal the sense-organ precursors for Johnston´s organ and their lineages. Beginning with a single progenitor at approximately a third of embryogenesis, additional precursors subsequently appear in both the ventral and dorsal pedicellar domains, each generating a lineage or clone. Lineage locations are remarkably conserved across preparations and ages, consistent with the epithelium possessing an underlying topographic coordinate system which determines the cellular organization of Johnston´s organ. By mid-embryogenesis twelve lineages are arranged circumferentially in the pedicel as in the adult structure. Each sense-organ precursor is associated with a smaller mitotically active cell from which the neuronal complement of each clone may derive. Neuron numbers within a clone increase in discrete steps with age, are invariant between clones and across preparations of a given age. At mid-embryogenesis each clone comprises five cells consolidated into a tightly bound cartridge. A long scolopale extends apically from each cartridge to an insertion point in the epithelium, and bundled axons project basally towards the brain. We review data from Drosophila to suggest mechanisms which might regulate the developmental program of Johnston´s organ in the locust.
... These actors are known to allow hormonal or neuropeptide modulation of neuronal activity, and they were previously described to be regulated by insecticide exposure [77,78]. Finally, some of our results also suggest that the low lethal dose exposure induced some neurogenesis or axonal growth, since we observed a significant increase in lachesin, an immunoglobulin superfamily protein whose expression correlates with neurogenesis [79] and of the SICKIE protein, which regulates F-actin mediated axonal growth in Drosophila mushroom body neurons [80]. ...
Insect pest management relies mainly on neurotoxic insecticides, including neonicotinoids such as clothianidin. The residual accumulation of low concentrations of these insecticides can have positive effects on target pest insects by enhancing various life traits. Because pest insects often rely on sex pheromones for reproduction and olfactory synaptic transmission is cholinergic, neonicotinoid residues could indeed modify chemical communication. We recently showed that treatments with low doses of clothianidin could induce hormetic effects on behavioral and neuronal sex pheromone responses in the male moth, Agrotis ipsilon. In this study, we used high-throughput RNAseq and proteomic analyses from brains of A. ipsilon males that were intoxicated with a low dose of clothianidin to investigate the molecular mechanisms leading to the observed hormetic effect. Our results showed that clothianidin induced significant changes in transcript levels and protein quantity in the brain of treated moths: 1229 genes and 49 proteins were differentially expressed upon clothianidin exposure. In particular, our analyses highlighted a regulation in numerous enzymes as a possible detoxification response to the insecticide and also numerous changes in neuronal processes, which could act as a form of acclimatization to the insecticide-contaminated environment, both leading to enhanced neuronal and behavioral responses to sex pheromone.
... During embryonic development of the locust, this transmembrane glycoprotein is also transiently expressed in ORNs and all olfactory neuropils . Another useful developmental marker is lachesin, a GPI-linked protein of the immunoglobulin family, that is expressed on the surface of developing locust neurons (Karlstrom et al., 1993;Boyan and Ehrhardt, 2020), including ingrowing larval ORN axons . ...
Regeneration after injury is accompanied by transient and lasting changes in the neuroarchitecture of the nervous system and, thus, a form of structural plasticity. In this review, we introduce the olfactory pathway of a particular insect as a convenient model to visualize neural regeneration at an anatomical level and study functional recovery at an electrophysiological level. The olfactory pathway of the locust (Locusta migratoria) is characterized by a multiglomerular innervation of the antennal lobe by olfactory receptor neurons. These olfactory afferents were axotomized by crushing the base of the antenna. The resulting degeneration and regeneration in the antennal lobe could be quantified by size measurements, dye labeling, and immunofluorescence staining of cell surface proteins implicated in axonal guidance during development. Within 3 days post lesion, the antennal lobe volume was reduced by 30% and from then onward regained size back to normal by 2 weeks post injury. The majority of regenerating olfactory receptor axons reinnervated the glomeruli of the antennal lobe. A few regenerating axons project erroneously into the mushroom body on a pathway that is normally chosen by second-order projection neurons. Based on intracellular responses of antennal lobe output neurons to odor stimulation, regenerated fibers establish functional synapses again. Following complete absence after nerve crush, responses to odor stimuli return to control level within 10–14 days. On average, regeneration of afferents, and re-established synaptic connections appear faster in younger fifth instar nymphs than in adults. The initial degeneration of olfactory receptor axons has a trans-synaptic effect on a second order brain center, leading to a transient size reduction of the mushroom body calyx. Odor-evoked oscillating field potentials, absent after nerve crush, were restored in the calyx, indicative of regenerative processes in the network architecture. We conclude that axonal regeneration in the locust olfactory system appears to be possible, precise, and fast, opening an avenue for future mechanistic studies. As a perspective of biomedical importance, the current evidence for nitric oxide/cGMP signaling as positive regulator of axon regeneration in connectives of the ventral nerve cord is considered in light of particular regeneration studies in vertebrate central nervous systems.
... In this study, we visualize the development of epithelial domains in the apical meristal annuli of the early embryonic antenna using the expression pattern of the grasshopper immunoglobulin Lachesin (Karlstrom et al. 1993). Lachesin is a glycosyl-phosphatidylinositol (GPI)-linked cell surface molecule with 70% sequence homology to Drosophila Lachesin and is known to function in neurite outgrowth, cell surface recognition, and cell adhesion (Llimargas et al. 2003;Strigini et al. 2006). ...
... Protocols for immunolabeling with primary and secondary antibodies, as well as for nuclear staining, followed Boyan and Williams (2004) and Ehrhardt et al. (2015aEhrhardt et al. ( , b, 2016. The following primary antibodies were employed: anti-horseradish peroxidase (α-HRP, Dianova, 323-005-021) which recognizes a neuron-specific cell surface epitope in insects (see Jan and Jan 1982;Haase et al. 2001); anti-Lachesin (Mab 1C10, gift of Dr. M. Bastiani) which recognizes the glycosyl-phosphatidylinositol (GPI)-linked cell surface molecule Lachesin expressed by differentiating epithelial cells (see Karlstrom et al. 1993); anti-phospho-Histone H3 (Millipore 06-570) which reveals mitotically active cells by recognizing and binding the phosphorylated form of the amine terminal of the histone H3, one of the four core histones wrapped inside genomic DNA and forming the core nucleosome complex (see Takizawa and Meshorer 2008); and anti-Lazarillo (Mab 10E6, a gift of Dr. M. Bastiani) which recognizes the GPI-linked cell surface lipocalin Lazarillo expressed by insect pioneer and sensory cells (see Ganfornina et al. 1995;Sánchez et al. 1995). The nuclear stain 4,6-diamidino-2-phenylindole (DAPI, Sigma) binds to the minor groove of double-stranded DNA and thus labels cell nuclei (see Naimski et al. 1980). ...
... We investigated the appearance of epithelial domains in the apical region of the early embryonic antenna via immunolabeling against the GPI-linked epithelial cell surface antigen Lachesin (Karlstrom et al. 1993) and then related these domains to the subsequent articulation of this region. Lachesin is expressed in discrete domains in all appendages such as the leg (Fig. 2a) and antenna (Fig. 2b) of the early embryonic grasshopper. ...
The antenna is a key sensory organ in insects. Factors which pattern its epithelium and the spacing of sensillae will play an important role in shaping its contribution to adaptive behavior. The antenna of the grasshopper S. gregaria has three major articulations: scape, pedicel, and flagellum. During postembryonic development, the flagellum lengthens as segments (so-called meristal annuli) are added at each molt. However, the five most apical annuli do not subdivide; thus, their epithelial domains must already be defined during embryogenesis. We investigated epithelial compartmentalization and its relationship to the developing primordial nervous system of the antenna by simultaneous immunolabeling against the epithelial cell surface molecule Lachesin, against neuron-specific horseradish peroxidase, and against the mitosis marker phospho-histone 3. We found that Lachesin is initially expressed in a highly ordered pattern of “rings” and a “sock” in the apical antennal epithelium of the early embryo. These expression domains appear in a stereotypic order and prefigure later articulations. Proliferative cells segregate into these developing domains and pioneer- and sensory-cell precursors were molecularly identified. Our study allows pioneer neurons, guidepost cells, and the earliest sensory cell clusters of the primordial nervous system to be allocated to their respective epithelial domain. As the apical-most five domains remain stable through subsequent development, lengthening of the flagellum must originate from more basal regions and is likely to be under the control of factors homologous to those which regulate boundary and joint formation in the antenna of Drosophila.
... Results revealed the neuron specific expression of lachesin in mid-gut, salivary gland and ovary which were again confirmed by co-localisation of ELAV protein with lachesin. The expression of lachesin in neuron and axons was first reported in grasshopper embryo as a membrane protein (Karlstrom, Wilder, & Bastiani, 1993). A study conducted on lachesin mutant embryo of Drosophila suggested that lachesin expresses on the cell surfaces of the specific tissues such as trachea, hindgut, foregut and nervous system which is required for the proper morphogenesis of the tracheal system and cell adhesion (Llimargas, Strigini, Katidou, Karagogeos, & Casanova, 2004). ...
Dengue virus (DENV) comprises of 4 serotypes (DENV‐1 to 4) and is medically one of the most important arboviruses (arthropod‐borne virus). DENV infection is a major human health burden and is transmitted between humans by the insect vector, Aedes aegypti. Ae. aegypti ingests DENV while feeding on infected humans, which traverses through its gut, haemolymph and salivary glands of the mosquito before being injected into a healthy human. During this process of transmission, DENV must interact with many proteins of the insect vector, which are important for its successful transmission. Our study focused on the identification and characterization of interacting protein partners in Ae. aegypti to DENV. Since domain III (DIII) of envelope protein (E) is exposed on the virion surface and is involved in virus entry into various cells, we performed phage display library screening against domain III of the envelope protein (EDIII) of DENV‐2. A peptide sequence showing similarity to lachesin protein was found interacting with EDIII. The lachesin protein was cloned, heterologously expressed, purified and used for in vitro interaction studies. Lachesin protein interacted with EDIII and also with DENV. Further, lachesin protein was localized in neuronal cells of different organs of Ae. aegypti by confocal microscopy. Blocking of lachesin protein in Ae. aegypti with anti‐lachesin antibody resulted in a significant reduction in DENV replication. This article is protected by copyright. All rights reserved.
... Neuronal acetylcholine receptor subunit alpha-7 has been found to be associated with a number of behaviors including social recognition, spatial learning and memory, and attention in rats and mice (reviewed by Pandya & Yakel 2013). (Karlstrom et al. 1993), and is particularly important in the development of the blood-brain barrier (Strigini et al. 2006). Lachesin mutants demonstrate an abnormal behavioral phenotype as embryos as they undergo a period of hyperactivity followed by 89 paralysis and an inability to hatch (Strigini et al. 2006). ...
Collective behavior is widespread in nature and examples include schools of fish and nest building in social insects. Although collective behavior and other group-level phenotypes are assumed to be shaped by selection, we do not know to what degree they are heritable and how selection acts on them. Furthermore, we have identified relatively few genes underlying variation in group-level phenotypes, hindering our understanding of the molecular mechanisms by which genes influence these traits and how they evolve. Elucidating the genetic architecture underlying group-level phenotypes is especially diffuclt because it depends on the genotypes of multiple interacting individuals. In this thesis, we use a new pharaoh ant (Monomorium pharaonis) laboratory mapping population to investigate the genetic architecture underlying a number of group-level phenotypes. These group-level phenotypes include collective behaviors (foraging, aggression, and exploration) and cuticular hydrocarbons, which play a vital role in chemical communication within social insect colonies. We demomonstrate that these phenotypes are heritable and have fitness consequences – the two prerequites for evolution via natural selection. Next, we perform genome-wide association studies to identify many interesting candidate genes associated with variation in group-level phenotypes, including genes associated with variation in collective behavior that have been implicated in neurological disorders or in the development of the visual system. Next, we explore how the genetic makeup of groups affects collective behavior and find that the specific combinations of genotypes within a group influence group-level outputs. Finally, we focus on the important social interactions between nurse workers and larvae. We first explore the evolutionary origin of sibling care and find that it likely shares a genetic basis with maternal care. Next, we demonstare that some nurse workers are behaviorally specicialized to care for larvae of different development stages and identify genes differentially expressed between nurses caring for different larval types. These specialized nurse workers likely play a large role in regulating divion of labor within social insect colonies. Overall, this work begins to identify the genetic architecture underlying group-level phenotypes, highlights the importance of within-group genetic composition on group-level output, and demonstrates the important role of nurse workers in modulating group-level phenotypes.
... In addition, the single nucleotide polymorphism (SNP) of OPCML, rs3016384, was found to be significantly associated with schizophrenia in both European and Thai populations (Athanasiu et al., 2010;Panichareon et al., 2012). Functional studies have revealed that OPCML is required for neurite outgrowth and cell surface recognition events in brain development (Faivre-Sarrailh and Rougon, 1997;Karlstrom et al., 1993;Rougon and Hobert, 2003). Furthermore, OPCML was reported to regulate synaptogenesis and synaptic plasticity, the disruption of which might contribute to neurodevelopmental disorders (Hashimoto et al., 2009;Yamada et al., 2007). ...
Previous genetic and biological evidence converge on the involvement of synaptic dysfunction in schizophrenia, and OPCML, encoding a synaptic membrane protein, is reported to be genetically associated with schizophrenia. However, its role in the pathophysiology of schizophrenia remains largely unknown. Here, we found that Opcml is strongly expressed in the mouse hippocampus; ablation of Opcml leads to reduced phosphorylated cofilin and dysregulated F-actin dynamics, which disturbs the spine maturation. Furthermore, Opcml interacts with EphB2 to control the stability of spines by regulating the ephrin-EphB2-cofilin signaling pathway. Opcml-deficient mice display impaired cognitive behaviors and abnormal sensorimotor gating, which are similar to features in neuropsychiatric disorders such as schizophrenia. Notably, the administration of aripiprazole partially restores the abnormal behaviors in Opcml-/- mice by increasing the phosphorylated cofilin level and facilitating spine maturation. We demonstrated a critical role of the schizophrenia-susceptible gene OPCML in spine maturation and cognitive behaviors via regulating the ephrin-EphB2-cofilin signaling pathway, providing further insights into the characteristics of schizophrenia.
