Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling

Autores
Bonnet, Laura V.; Palandri, Anabela; Flores Martin, Jesica B.; Hallak, Marta E.
Año de publicación
2024
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Fil: Bonnet Laura V. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Bonnet Laura V. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Palandri Anabela. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Palandri Anabela. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Flores Martin Jesica B. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Flores Martin Jesica B. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Hallak Marta E. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Hallak Marta E. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Background. Arginyltransferase (Ate1) orchestrates posttranslational protein arginylation, a pivotal regulator of cellular proteolytic processes. In eukaryotic cells, two interconnected systems—the ubiquitin proteasome system (UPS) and macroautophagy—mediate proteolysis and cooperate to maintain quality protein control and cellular homeostasis. Previous studies have shown that N-terminal arginylation facilitates protein degradation through the UPS. Dysregulation of this machinery triggers p62-mediated autophagy to ensure proper substrate processing. Nevertheless, how Ate1 operates through this intricate mechanism remains elusive. Methods. We investigated Ate1 subcellular distribution through confocal microscopy and biochemical assays using cells transiently or stably expressing either endogenous Ate1 or a GFP-tagged Ate1 isoform transfected in CHO-K1 or MEFs, respectively. To assess Ate1 and p62-cargo clustering, we analyzed their colocalization and multimerization status by immunofluorescence and nonreducing immunoblotting, respectively. Additionally, we employed Ate1 KO cells to examine the role of Ate1 in autophagy. Ate1 KO MEFs cells stably expressing GFP-tagged Ate1-1 isoform were used as a model for phenotype rescue. Autophagy dynamics were evaluated by analyzing LC3B turnover and p62/SQSTM1 levels under both steady-state and serum-starvation conditions, through immunoblotting and immunofluorescence. We determined mTORC1/AMPk activation by assessing mTOR and AMPk phosphorylation through immunoblotting, while mTORC1 lysosomal localization was monitored by confocal microscopy. Results. Here, we report a multifaceted role for Ate1 in the autophagic process, wherein it clusters with p62, facilitates autophagic clearance, and modulates its signaling. Mechanistically, we found that cell-specific inactivation of Ate1 elicits overactivation of the mTORC1/AMPk signaling hub that underlies a failure in autophagic flux and subsequent substrate accumulation, which is partially rescued by ectopic expression of Ate1. Statistical significance was assessed using a two-sided unpaired t test with a significance threshold set at P<0.05. Conclusions. Our findings uncover a critical housekeeping role of Ate1 in mTORC1/AMPk-regulated autophagy, as a potential therapeutic target related to this pathway, that is dysregulated in many neurodegenerative and cancer diseases.
info:eu-repo/semantics/publishedVersion
Fil: Bonnet Laura V. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Bonnet Laura V. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Palandri Anabela. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Palandri Anabela. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Flores Martin Jesica B. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Flores Martin Jesica B. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Hallak Marta E. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Hallak Marta E. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Materia
Arginylation
Arginyltransferase 1
Autophagy
mTORC1
Posttranslational modifcation
p62/SQSTM1
Nivel de accesibilidad
acceso abierto
Condiciones de uso
Repositorio
Repositorio Digital Universitario (UNC)
Institución
Universidad Nacional de Córdoba
OAI Identificador
oai:rdu.unc.edu.ar:11086/552355
Acceder
id RDUUNC_f005d98b350a50a29a2c274bfbc51422
oai_identifier_str oai:rdu.unc.edu.ar:11086/552355
network_acronym_str RDUUNC
repository_id_str 2572
network_name_str Repositorio Digital Universitario (UNC)
spelling Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signalingBonnet, Laura V.Palandri, AnabelaFlores Martin, Jesica B.Hallak, Marta E.ArginylationArginyltransferase 1AutophagymTORC1Posttranslational modifcationp62/SQSTM1Fil: Bonnet Laura V. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.Fil: Bonnet Laura V. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.Fil: Palandri Anabela. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.Fil: Palandri Anabela. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.Fil: Flores Martin Jesica B. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.Fil: Flores Martin Jesica B. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.Fil: Hallak Marta E. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.Fil: Hallak Marta E. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.Background. Arginyltransferase (Ate1) orchestrates posttranslational protein arginylation, a pivotal regulator of cellular proteolytic processes. In eukaryotic cells, two interconnected systems—the ubiquitin proteasome system (UPS) and macroautophagy—mediate proteolysis and cooperate to maintain quality protein control and cellular homeostasis. Previous studies have shown that N-terminal arginylation facilitates protein degradation through the UPS. Dysregulation of this machinery triggers p62-mediated autophagy to ensure proper substrate processing. Nevertheless, how Ate1 operates through this intricate mechanism remains elusive. Methods. We investigated Ate1 subcellular distribution through confocal microscopy and biochemical assays using cells transiently or stably expressing either endogenous Ate1 or a GFP-tagged Ate1 isoform transfected in CHO-K1 or MEFs, respectively. To assess Ate1 and p62-cargo clustering, we analyzed their colocalization and multimerization status by immunofluorescence and nonreducing immunoblotting, respectively. Additionally, we employed Ate1 KO cells to examine the role of Ate1 in autophagy. Ate1 KO MEFs cells stably expressing GFP-tagged Ate1-1 isoform were used as a model for phenotype rescue. Autophagy dynamics were evaluated by analyzing LC3B turnover and p62/SQSTM1 levels under both steady-state and serum-starvation conditions, through immunoblotting and immunofluorescence. We determined mTORC1/AMPk activation by assessing mTOR and AMPk phosphorylation through immunoblotting, while mTORC1 lysosomal localization was monitored by confocal microscopy. Results. Here, we report a multifaceted role for Ate1 in the autophagic process, wherein it clusters with p62, facilitates autophagic clearance, and modulates its signaling. Mechanistically, we found that cell-specific inactivation of Ate1 elicits overactivation of the mTORC1/AMPk signaling hub that underlies a failure in autophagic flux and subsequent substrate accumulation, which is partially rescued by ectopic expression of Ate1. Statistical significance was assessed using a two-sided unpaired t test with a significance threshold set at P<0.05. Conclusions. Our findings uncover a critical housekeeping role of Ate1 in mTORC1/AMPk-regulated autophagy, as a potential therapeutic target related to this pathway, that is dysregulated in many neurodegenerative and cancer diseases.info:eu-repo/semantics/publishedVersionFil: Bonnet Laura V. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.Fil: Bonnet Laura V. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.Fil: Palandri Anabela. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.Fil: Palandri Anabela. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.Fil: Flores Martin Jesica B. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.Fil: Flores Martin Jesica B. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.Fil: Hallak Marta E. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.Fil: Hallak Marta E. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.BioMed Central Ltd. Part of Springer Naturehttps://orcid.org/0000-0001-5493-0949https://orcid.org/0000-0003-4856-879Xhttps://orcid.org/0000-0002-1619-399X2024-01-31info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfBonnet, L.V., Palandri, A., Flores-Martin, J.B. et al. Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling. Cell Commun Signal 22, 87 (2024). https://doi.org/10.1186/s12964-024-01499-9http://hdl.handle.net/11086/5523551478-811Xhttps://link.springer.com/article/10.1186/s12964-024-01499-9https://doi.org/10.1186/s12964-024-01499-9enginfo:eu-repo/semantics/openAccessreponame:Repositorio Digital Universitario (UNC)instname:Universidad Nacional de Córdobainstacron:UNC2025-09-18T10:08:49Zoai:rdu.unc.edu.ar:11086/552355Institucionalhttps://rdu.unc.edu.ar/Universidad públicaNo correspondehttp://rdu.unc.edu.ar/oai/snrdoca.unc@gmail.comArgentinaNo correspondeNo correspondeNo correspondeopendoar:25722025-09-18 10:08:49.334Repositorio Digital Universitario (UNC) - Universidad Nacional de Córdobafalse
dc.title.none.fl_str_mv Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling
title Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling
spellingShingle Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling
Bonnet, Laura V.
Arginylation
Arginyltransferase 1
Autophagy
mTORC1
Posttranslational modifcation
p62/SQSTM1
title_short Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling
title_full Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling
title_fullStr Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling
title_full_unstemmed Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling
title_sort Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling
dc.creator.none.fl_str_mv Bonnet, Laura V.
Palandri, Anabela
Flores Martin, Jesica B.
Hallak, Marta E.
author Bonnet, Laura V.
author_facet Bonnet, Laura V.
Palandri, Anabela
Flores Martin, Jesica B.
Hallak, Marta E.
author_role author
author2 Palandri, Anabela
Flores Martin, Jesica B.
Hallak, Marta E.
author2_role author
author
author
dc.contributor.none.fl_str_mv https://orcid.org/0000-0001-5493-0949
https://orcid.org/0000-0003-4856-879X
https://orcid.org/0000-0002-1619-399X
dc.subject.none.fl_str_mv Arginylation
Arginyltransferase 1
Autophagy
mTORC1
Posttranslational modifcation
p62/SQSTM1
topic Arginylation
Arginyltransferase 1
Autophagy
mTORC1
Posttranslational modifcation
p62/SQSTM1
dc.description.none.fl_txt_mv Fil: Bonnet Laura V. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Bonnet Laura V. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Palandri Anabela. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Palandri Anabela. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Flores Martin Jesica B. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Flores Martin Jesica B. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Hallak Marta E. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Hallak Marta E. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Background. Arginyltransferase (Ate1) orchestrates posttranslational protein arginylation, a pivotal regulator of cellular proteolytic processes. In eukaryotic cells, two interconnected systems—the ubiquitin proteasome system (UPS) and macroautophagy—mediate proteolysis and cooperate to maintain quality protein control and cellular homeostasis. Previous studies have shown that N-terminal arginylation facilitates protein degradation through the UPS. Dysregulation of this machinery triggers p62-mediated autophagy to ensure proper substrate processing. Nevertheless, how Ate1 operates through this intricate mechanism remains elusive. Methods. We investigated Ate1 subcellular distribution through confocal microscopy and biochemical assays using cells transiently or stably expressing either endogenous Ate1 or a GFP-tagged Ate1 isoform transfected in CHO-K1 or MEFs, respectively. To assess Ate1 and p62-cargo clustering, we analyzed their colocalization and multimerization status by immunofluorescence and nonreducing immunoblotting, respectively. Additionally, we employed Ate1 KO cells to examine the role of Ate1 in autophagy. Ate1 KO MEFs cells stably expressing GFP-tagged Ate1-1 isoform were used as a model for phenotype rescue. Autophagy dynamics were evaluated by analyzing LC3B turnover and p62/SQSTM1 levels under both steady-state and serum-starvation conditions, through immunoblotting and immunofluorescence. We determined mTORC1/AMPk activation by assessing mTOR and AMPk phosphorylation through immunoblotting, while mTORC1 lysosomal localization was monitored by confocal microscopy. Results. Here, we report a multifaceted role for Ate1 in the autophagic process, wherein it clusters with p62, facilitates autophagic clearance, and modulates its signaling. Mechanistically, we found that cell-specific inactivation of Ate1 elicits overactivation of the mTORC1/AMPk signaling hub that underlies a failure in autophagic flux and subsequent substrate accumulation, which is partially rescued by ectopic expression of Ate1. Statistical significance was assessed using a two-sided unpaired t test with a significance threshold set at P<0.05. Conclusions. Our findings uncover a critical housekeeping role of Ate1 in mTORC1/AMPk-regulated autophagy, as a potential therapeutic target related to this pathway, that is dysregulated in many neurodegenerative and cancer diseases.
info:eu-repo/semantics/publishedVersion
Fil: Bonnet Laura V. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Bonnet Laura V. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Palandri Anabela. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Palandri Anabela. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Flores Martin Jesica B. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Flores Martin Jesica B. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
Fil: Hallak Marta E. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
Fil: Hallak Marta E. Consejo Nacional de Investigaciones Científicas y Técnicas. Argentina. Centro de Investigaciones en Química Biológica de Córdoba, Argentina.
description Fil: Bonnet Laura V. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica Ranwel Caputto, Argentina.
publishDate 2024
dc.date.none.fl_str_mv 2024-01-31
dc.type.none.fl_str_mv info:eu-repo/semantics/publishedVersion
info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
status_str publishedVersion
format article
dc.identifier.none.fl_str_mv Bonnet, L.V., Palandri, A., Flores-Martin, J.B. et al. Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling. Cell Commun Signal 22, 87 (2024). https://doi.org/10.1186/s12964-024-01499-9
http://hdl.handle.net/11086/552355
1478-811X
https://link.springer.com/article/10.1186/s12964-024-01499-9
https://doi.org/10.1186/s12964-024-01499-9
identifier_str_mv Bonnet, L.V., Palandri, A., Flores-Martin, J.B. et al. Arginyltransferase 1 modulates p62-driven autophagy via mTORC1/AMPk signaling. Cell Commun Signal 22, 87 (2024). https://doi.org/10.1186/s12964-024-01499-9
1478-811X
url http://hdl.handle.net/11086/552355
https://link.springer.com/article/10.1186/s12964-024-01499-9
https://doi.org/10.1186/s12964-024-01499-9
dc.language.none.fl_str_mv eng
language eng
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv BioMed Central Ltd. Part of Springer Nature
publisher.none.fl_str_mv BioMed Central Ltd. Part of Springer Nature
dc.source.none.fl_str_mv reponame:Repositorio Digital Universitario (UNC)
instname:Universidad Nacional de Córdoba
instacron:UNC
reponame_str Repositorio Digital Universitario (UNC)
collection Repositorio Digital Universitario (UNC)
instname_str Universidad Nacional de Córdoba
instacron_str UNC
institution UNC
repository.name.fl_str_mv Repositorio Digital Universitario (UNC) - Universidad Nacional de Córdoba
repository.mail.fl_str_mv oca.unc@gmail.com
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score 13.001348

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