{"id":{"repo_id":"sherbrooke","oai_identifier":"oai:usherbrooke.scholaris.ca:11143/22548"},"canonical_url":"https://search.dev.ndltd.org/etd/sherbrooke/oai:usherbrooke.scholaris.ca:11143/22548","repository":{"repo_id":"sherbrooke","name":"Université de Sherbrooke","base_url":"https://usherbrooke.scholaris.ca/server/oai/request"},"display":{"title":"Régulation du récepteur aux opioïdes de type delta par les protéines Homer1","abstract":"Compared to other opioid receptors, the delta opioid receptor (DOP), a G proteincoupled receptor (GPCR), is considered a promising therapeutic target for chronic pain management. However, DOP mainly localizes to intracellular compartments whereas its analgesic effects occur when translocated to the plasma membrane. The mechanisms involved in its trafficking remain also poorly known. Recently, liquid chromatographytandem mass spectrometry (LC-MS/MS) analyses established the DOP interactome, which included Homer Protein Homolog 1 (Homer1). Interestingly, it was reported that its short and long isoforms, namely Homer1a and Homer1b, regulate the cell-surface expression of various GPCR. Furthermore, neuropathic and inflammatory models demonstrated pain modulation by Homer1. We thus hypothesized that Homer1 isoforms regulate DOP. Resuming preliminary work on this project (Degrandmaison, 2023) with DOP and Homer1 human orthologs (hDOP, Homer1a and Homer1b), we first determined that Homer1b interacts with the hDOP C-terminal (CT) and third intracellular loop (ICL3) in GSTPulldowns binding assays. We then generated truncated, alanine and phosphomimetic GSThDOP-CT mutants to investigate a potential Homer1 consensus binding motif. Whereas amino acids 339 to 354 of hDOP-CT were important to Homer1 binding, individual residues within this sequence were not identified as essential for this interaction. Indirect ELISA assays in HEK 293 cells resulted in a decrease in hDOP cell-surface expression with transient overexpression of Homer1b; preliminary results showed also a decrease of hDOP cellsurface expression with CT 339 and 354 deletions. Furthermore, endogenous Homer1b colocalization with stably expressed hDOP in HEK 293 cells was observed at the plasma membrane and in intracellular compartments by confocal fluorescence microscopy. Lastly, stimulation of hDOP with SNC80 and [D-Ala2] Deltorphine II did not show any significant Homer1 regulatory effect on ERK1/2 signaling pathway. Altogether, these results will further our understanding of the role and pharmacological potential of Homer1 interaction with DOP.","abstract_html":"Compared to other opioid receptors, the delta opioid receptor (DOP), a G proteincoupled receptor (GPCR), is considered a promising therapeutic target for chronic pain management. However, DOP mainly localizes to intracellular compartments whereas its analgesic effects occur when translocated to the plasma membrane. The mechanisms involved in its trafficking remain also poorly known. Recently, liquid chromatographytandem mass spectrometry (LC-MS/MS) analyses established the DOP interactome, which included Homer Protein Homolog 1 (Homer1). Interestingly, it was reported that its short and long isoforms, namely Homer1a and Homer1b, regulate the cell-surface expression of various GPCR. Furthermore, neuropathic and inflammatory models demonstrated pain modulation by Homer1. We thus hypothesized that Homer1 isoforms regulate DOP. Resuming preliminary work on this project (Degrandmaison, 2023) with DOP and Homer1 human orthologs (hDOP, Homer1a and Homer1b), we first determined that Homer1b interacts with the hDOP C-terminal (CT) and third intracellular loop (ICL3) in GSTPulldowns binding assays. We then generated truncated, alanine and phosphomimetic GSThDOP-CT mutants to investigate a potential Homer1 consensus binding motif. Whereas amino acids 339 to 354 of hDOP-CT were important to Homer1 binding, individual residues within this sequence were not identified as essential for this interaction. Indirect ELISA assays in HEK 293 cells resulted in a decrease in hDOP cell-surface expression with transient overexpression of Homer1b; preliminary results showed also a decrease of hDOP cellsurface expression with CT 339 and 354 deletions. Furthermore, endogenous Homer1b colocalization with stably expressed hDOP in HEK 293 cells was observed at the plasma membrane and in intracellular compartments by confocal fluorescence microscopy. Lastly, stimulation of hDOP with SNC80 and [D-Ala2] Deltorphine II did not show any significant Homer1 regulatory effect on ERK1/2 signaling pathway. Altogether, these results will further our understanding of the role and pharmacological potential of Homer1 interaction with DOP.","abstract_has_math":false,"creators":["Grisé, Olivier"],"institution":"Université de Sherbrooke","degree_name":"M. Sc.","degree_level":"Maîtrise","degree_discipline":"Pharmacologie","degree_department":null,"school":null,"contributors":[],"advisors":["Parent, Jean-Luc","Gendron, Louis"],"committee_chairs":[],"committee_members":[],"year":2025,"date_issued":"2025","date_published":"2025","updated_at":"2026-07-27T21:07:49Z","subjects":["Récepteur couplé aux protéines G (RCPG)","Récepteur aux opioïdes de type delta (DOP)","Homer1","Douleur","G protein-coupled receptors (GPCR)","Delta-opioid receptor (DOP)","Pain"],"languages":["fr"],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"http://hdl.handle.net/11143/22548","outbound_label":"Handle","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.advisor","label":"Advisor","values":["Parent, Jean-Luc","Gendron, Louis"]},{"key":"dc:creator","label":"Author","values":["Grisé, Olivier"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.accessioned","label":"Dc Date Accessioned","values":["2025-03-11T14:08:04Z"]},{"key":"dc:date.available","label":"Dc Date Available","values":["2025-03-11T14:08:04Z"]},{"key":"dc:date.issued","label":"Date","values":["2025"]},{"key":"dc:publisher","label":"Institution","values":["Université de Sherbrooke"]},{"key":"dc:type","label":"Dc Type","values":["Mémoire de maîtrise"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Pharmacologie"]},{"key":"thesis:degree_level","label":"Degree Level","values":["Maîtrise"]},{"key":"thesis:degree_name","label":"Degree Name","values":["M. 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However, DOP mainly localizes to intracellular compartments whereas its analgesic effects occur when translocated to the plasma membrane. The mechanisms involved in its trafficking remain also poorly known. Recently, liquid chromatographytandem mass spectrometry (LC-MS/MS) analyses established the DOP interactome, which included Homer Protein Homolog 1 (Homer1). Interestingly, it was reported that its short and long isoforms, namely Homer1a and Homer1b, regulate the cell-surface expression of various GPCR. Furthermore, neuropathic and inflammatory models demonstrated pain modulation by Homer1. We thus hypothesized that Homer1 isoforms regulate DOP. Resuming preliminary work on this project (Degrandmaison, 2023) with DOP and Homer1 human orthologs (hDOP, Homer1a and Homer1b), we first determined that Homer1b interacts with the hDOP C-terminal (CT) and third intracellular loop (ICL3) in GSTPulldowns binding assays. We then generated truncated, alanine and phosphomimetic GSThDOP-CT mutants to investigate a potential Homer1 consensus binding motif. Whereas amino acids 339 to 354 of hDOP-CT were important to Homer1 binding, individual residues within this sequence were not identified as essential for this interaction. Indirect ELISA assays in HEK 293 cells resulted in a decrease in hDOP cell-surface expression with transient overexpression of Homer1b; preliminary results showed also a decrease of hDOP cellsurface expression with CT 339 and 354 deletions. Furthermore, endogenous Homer1b colocalization with stably expressed hDOP in HEK 293 cells was observed at the plasma membrane and in intracellular compartments by confocal fluorescence microscopy. Lastly, stimulation of hDOP with SNC80 and [D-Ala2] Deltorphine II did not show any significant Homer1 regulatory effect on ERK1/2 signaling pathway. Altogether, these results will further our understanding of the role and pharmacological potential of Homer1 interaction with DOP.","Le récepteur aux opioïdes de type delta (DOP), un récepteur couplé aux protéines G (RCPG), serait une cible thérapeutique prometteuse comparé aux autres récepteurs opioïdes pour traiter la douleur chronique. Cependant, DOP est largement retenu au niveau de compartiments intracellulaires et ses effets analgésiques corrèlent avec sa translocation vers la membrane plasmique. Les mécanismes impliqués sont d’ailleurs méconnus. Récemment, des analyses par chromatographie liquide couplée à la spectrométrie de masse (LC-MS/MS) ont permis d’identifier l’interactome de DOP, dont la protéine échafaud Homer Protein Homolog 1 (Homer1) faisait partie. Des études ont rapporté que les isoformes courtes et longues, Homer1a et Homer1b respectivement, régulent l’expression à la membrane plasmique de certains RCPG. De plus, des modèles neuropathiques et inflammatoires supportent une modulation de la douleur par Homer1. Nous avons donc émis l’hypothèse que les isoformes d’Homer1 régulent le trafic du récepteur DOP. Poursuivant des travaux préliminaires sur ce projet (Degrandmaison, 2023) avec les orthologues humains de DOP (hDOP) et d’Homer1, Homer1a et Homer1b, des essais GST-Pulldowns ont d’abord déterminé que les domaines C-terminal (CT) et la boucle intracellulaire 3 (ICL3) du récepteur hDOP interagissent avec Homer1b. Nous avons ensuite généré des mutants tronqués, alanines et phosphomimétiques de GST-hDOP-CT afin d’investiguer un potentiel motif d’interaction avec Homer1 dans ce domaine. Bien que l’importance des acides aminés 339 à 354 de hDOP-CT ait été confirmée, les résidus individuels du motif n’ont pas été identifiés comme essentiels à l’interaction. Une diminution de l’expression en surface de cellules HEK 293 de hDOP a ensuite été observée par ELISA indirect en surexpression transitoire d’Homer1b; un résultat préliminaire montrait aussi une diminution de l’expression en surface cellulaire de hDOP lorsque tronqué aux positions 339 et 354. De plus, une colocalisation de l’isoforme endogène Homer1b avec hDOP exprimé de manière constitutive dans les cellules HEK 293 a été observée à la membrane plasmique et au niveau intracellulaire par microscopie confocale à fluorescence. Enfin, des stimulations de hDOP avec le SNC80 ou avec la [D-Ala2] Deltorphine II n’ont montré aucun effet modulateur des isoformes d’Homer1 sur la voie de signalisation des ERK1/2. Ces résultats contribueront à la compréhension du rôle et du potentiel pharmacologique de l’interaction entre Homer1 et DOP."]},{"key":"dc:title","label":"Title","values":["Régulation du récepteur aux opioïdes de type delta par les protéines Homer1"]}]}],"canonical_facts":{"dc:contributor.advisor":["Parent, Jean-Luc","Gendron, Louis"],"dc:creator":["Grisé, Olivier"],"dc:date.accessioned":["2025-03-11T14:08:04Z"],"dc:date.available":["2025-03-11T14:08:04Z"],"dc:date.issued":["2025"],"dc:description.abstract":["Compared to other opioid receptors, the delta opioid receptor (DOP), a G proteincoupled receptor (GPCR), is considered a promising therapeutic target for chronic pain management. However, DOP mainly localizes to intracellular compartments whereas its analgesic effects occur when translocated to the plasma membrane. The mechanisms involved in its trafficking remain also poorly known. Recently, liquid chromatographytandem mass spectrometry (LC-MS/MS) analyses established the DOP interactome, which included Homer Protein Homolog 1 (Homer1). Interestingly, it was reported that its short and long isoforms, namely Homer1a and Homer1b, regulate the cell-surface expression of various GPCR. Furthermore, neuropathic and inflammatory models demonstrated pain modulation by Homer1. We thus hypothesized that Homer1 isoforms regulate DOP. Resuming preliminary work on this project (Degrandmaison, 2023) with DOP and Homer1 human orthologs (hDOP, Homer1a and Homer1b), we first determined that Homer1b interacts with the hDOP C-terminal (CT) and third intracellular loop (ICL3) in GSTPulldowns binding assays. We then generated truncated, alanine and phosphomimetic GSThDOP-CT mutants to investigate a potential Homer1 consensus binding motif. Whereas amino acids 339 to 354 of hDOP-CT were important to Homer1 binding, individual residues within this sequence were not identified as essential for this interaction. Indirect ELISA assays in HEK 293 cells resulted in a decrease in hDOP cell-surface expression with transient overexpression of Homer1b; preliminary results showed also a decrease of hDOP cellsurface expression with CT 339 and 354 deletions. Furthermore, endogenous Homer1b colocalization with stably expressed hDOP in HEK 293 cells was observed at the plasma membrane and in intracellular compartments by confocal fluorescence microscopy. Lastly, stimulation of hDOP with SNC80 and [D-Ala2] Deltorphine II did not show any significant Homer1 regulatory effect on ERK1/2 signaling pathway. Altogether, these results will further our understanding of the role and pharmacological potential of Homer1 interaction with DOP.","Le récepteur aux opioïdes de type delta (DOP), un récepteur couplé aux protéines G (RCPG), serait une cible thérapeutique prometteuse comparé aux autres récepteurs opioïdes pour traiter la douleur chronique. Cependant, DOP est largement retenu au niveau de compartiments intracellulaires et ses effets analgésiques corrèlent avec sa translocation vers la membrane plasmique. Les mécanismes impliqués sont d’ailleurs méconnus. Récemment, des analyses par chromatographie liquide couplée à la spectrométrie de masse (LC-MS/MS) ont permis d’identifier l’interactome de DOP, dont la protéine échafaud Homer Protein Homolog 1 (Homer1) faisait partie. Des études ont rapporté que les isoformes courtes et longues, Homer1a et Homer1b respectivement, régulent l’expression à la membrane plasmique de certains RCPG. De plus, des modèles neuropathiques et inflammatoires supportent une modulation de la douleur par Homer1. Nous avons donc émis l’hypothèse que les isoformes d’Homer1 régulent le trafic du récepteur DOP. Poursuivant des travaux préliminaires sur ce projet (Degrandmaison, 2023) avec les orthologues humains de DOP (hDOP) et d’Homer1, Homer1a et Homer1b, des essais GST-Pulldowns ont d’abord déterminé que les domaines C-terminal (CT) et la boucle intracellulaire 3 (ICL3) du récepteur hDOP interagissent avec Homer1b. Nous avons ensuite généré des mutants tronqués, alanines et phosphomimétiques de GST-hDOP-CT afin d’investiguer un potentiel motif d’interaction avec Homer1 dans ce domaine. Bien que l’importance des acides aminés 339 à 354 de hDOP-CT ait été confirmée, les résidus individuels du motif n’ont pas été identifiés comme essentiels à l’interaction. Une diminution de l’expression en surface de cellules HEK 293 de hDOP a ensuite été observée par ELISA indirect en surexpression transitoire d’Homer1b; un résultat préliminaire montrait aussi une diminution de l’expression en surface cellulaire de hDOP lorsque tronqué aux positions 339 et 354. De plus, une colocalisation de l’isoforme endogène Homer1b avec hDOP exprimé de manière constitutive dans les cellules HEK 293 a été observée à la membrane plasmique et au niveau intracellulaire par microscopie confocale à fluorescence. Enfin, des stimulations de hDOP avec le SNC80 ou avec la [D-Ala2] Deltorphine II n’ont montré aucun effet modulateur des isoformes d’Homer1 sur la voie de signalisation des ERK1/2. Ces résultats contribueront à la compréhension du rôle et du potentiel pharmacologique de l’interaction entre Homer1 et DOP."],"dc:identifier.uri":["http://hdl.handle.net/11143/22548"],"dc:language.iso":["fr"],"dc:publisher":["Université de Sherbrooke"],"dc:subject":["Récepteur couplé aux protéines G (RCPG)","Récepteur aux opioïdes de type delta (DOP)","Homer1","Douleur","G protein-coupled receptors (GPCR)","Delta-opioid receptor (DOP)","Pain"],"dc:title":["Régulation du récepteur aux opioïdes de type delta par les protéines Homer1"],"dc:type":["Mémoire de maîtrise"],"thesis:degree_discipline":["Pharmacologie"],"thesis:degree_level":["Maîtrise"],"thesis:degree_name":["M. Sc."],"thesis:institution_name":["Faculté de médecine et des sciences de la santé"]},"updated_at":"2026-07-27T21:07:49Z"}