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In vivo assembly and trafficking of olfactory Ionotropic Receptors
BACKGROUND: Ionotropic receptors (IRs) are a large, divergent subfamily of ionotropic glutamate receptors (iGluRs) that are expressed in diverse peripheral sensory neurons and function in olfaction, taste, hygrosensation and thermosensation. Analogous to the cell biological properties of their synap...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2019
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6472016/ https://www.ncbi.nlm.nih.gov/pubmed/30995910 http://dx.doi.org/10.1186/s12915-019-0651-7 |
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author | Abuin, Liliane Prieto-Godino, Lucia L. Pan, Haiyun Gutierrez, Craig Huang, Lan Jin, Rongsheng Benton, Richard |
author_facet | Abuin, Liliane Prieto-Godino, Lucia L. Pan, Haiyun Gutierrez, Craig Huang, Lan Jin, Rongsheng Benton, Richard |
author_sort | Abuin, Liliane |
collection | PubMed |
description | BACKGROUND: Ionotropic receptors (IRs) are a large, divergent subfamily of ionotropic glutamate receptors (iGluRs) that are expressed in diverse peripheral sensory neurons and function in olfaction, taste, hygrosensation and thermosensation. Analogous to the cell biological properties of their synaptic iGluR ancestors, IRs are thought to form heteromeric complexes that localise to the ciliated dendrites of sensory neurons. IR complexes are composed of selectively expressed ‘tuning’ receptors and one of two broadly expressed co-receptors (IR8a or IR25a). While the extracellular ligand-binding domain (LBD) of tuning IRs is likely to define the stimulus specificity of the complex, the role of this domain in co-receptors is unclear. RESULTS: We identify a sequence in the co-receptor LBD, the ‘co-receptor extra loop’ (CREL), which is conserved across IR8a and IR25a orthologues but not present in either tuning IRs or iGluRs. The CREL contains a single predicted N-glycosylation site, which we show bears a sugar modification in recombinantly expressed IR8a. Using the Drosophila olfactory system as an in vivo model, we find that a transgenically encoded IR8a mutant in which the CREL cannot be N-glycosylated is impaired in localisation to cilia in some, though not all, populations of sensory neurons expressing different tuning IRs. This defect can be complemented by the presence of endogenous wild-type IR8a, indicating that IR complexes contain at least two IR8a subunits and that this post-translational modification is dispensable for protein folding or complex assembly. Analysis of the subcellular distribution of the mutant protein suggests that its absence from sensory cilia is due to a failure in exit from the endoplasmic reticulum. Protein modelling and in vivo analysis of tuning IR and co-receptor subunit interactions by a fluorescent protein fragment complementation assay reveal that the CREL N-glycosylation site is likely to be located on the external face of a heterotetrameric IR complex. CONCLUSIONS: Our data reveal an important role for the IR co-receptor LBD in control of intracellular transport, provide novel insights into the stoichiometry and assembly of IR complexes and uncover an unexpected heterogeneity in the trafficking regulation of this sensory receptor family. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12915-019-0651-7) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-6472016 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-64720162019-04-24 In vivo assembly and trafficking of olfactory Ionotropic Receptors Abuin, Liliane Prieto-Godino, Lucia L. Pan, Haiyun Gutierrez, Craig Huang, Lan Jin, Rongsheng Benton, Richard BMC Biol Research Article BACKGROUND: Ionotropic receptors (IRs) are a large, divergent subfamily of ionotropic glutamate receptors (iGluRs) that are expressed in diverse peripheral sensory neurons and function in olfaction, taste, hygrosensation and thermosensation. Analogous to the cell biological properties of their synaptic iGluR ancestors, IRs are thought to form heteromeric complexes that localise to the ciliated dendrites of sensory neurons. IR complexes are composed of selectively expressed ‘tuning’ receptors and one of two broadly expressed co-receptors (IR8a or IR25a). While the extracellular ligand-binding domain (LBD) of tuning IRs is likely to define the stimulus specificity of the complex, the role of this domain in co-receptors is unclear. RESULTS: We identify a sequence in the co-receptor LBD, the ‘co-receptor extra loop’ (CREL), which is conserved across IR8a and IR25a orthologues but not present in either tuning IRs or iGluRs. The CREL contains a single predicted N-glycosylation site, which we show bears a sugar modification in recombinantly expressed IR8a. Using the Drosophila olfactory system as an in vivo model, we find that a transgenically encoded IR8a mutant in which the CREL cannot be N-glycosylated is impaired in localisation to cilia in some, though not all, populations of sensory neurons expressing different tuning IRs. This defect can be complemented by the presence of endogenous wild-type IR8a, indicating that IR complexes contain at least two IR8a subunits and that this post-translational modification is dispensable for protein folding or complex assembly. Analysis of the subcellular distribution of the mutant protein suggests that its absence from sensory cilia is due to a failure in exit from the endoplasmic reticulum. Protein modelling and in vivo analysis of tuning IR and co-receptor subunit interactions by a fluorescent protein fragment complementation assay reveal that the CREL N-glycosylation site is likely to be located on the external face of a heterotetrameric IR complex. CONCLUSIONS: Our data reveal an important role for the IR co-receptor LBD in control of intracellular transport, provide novel insights into the stoichiometry and assembly of IR complexes and uncover an unexpected heterogeneity in the trafficking regulation of this sensory receptor family. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12915-019-0651-7) contains supplementary material, which is available to authorized users. BioMed Central 2019-04-17 /pmc/articles/PMC6472016/ /pubmed/30995910 http://dx.doi.org/10.1186/s12915-019-0651-7 Text en © The Author(s). 2019 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Research Article Abuin, Liliane Prieto-Godino, Lucia L. Pan, Haiyun Gutierrez, Craig Huang, Lan Jin, Rongsheng Benton, Richard In vivo assembly and trafficking of olfactory Ionotropic Receptors |
title | In vivo assembly and trafficking of olfactory Ionotropic Receptors |
title_full | In vivo assembly and trafficking of olfactory Ionotropic Receptors |
title_fullStr | In vivo assembly and trafficking of olfactory Ionotropic Receptors |
title_full_unstemmed | In vivo assembly and trafficking of olfactory Ionotropic Receptors |
title_short | In vivo assembly and trafficking of olfactory Ionotropic Receptors |
title_sort | in vivo assembly and trafficking of olfactory ionotropic receptors |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6472016/ https://www.ncbi.nlm.nih.gov/pubmed/30995910 http://dx.doi.org/10.1186/s12915-019-0651-7 |
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