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Comparative Proteomics Unveils LRRFIP1 as a New Player in the DAPK1 Interactome of Neurons Exposed to Oxygen and Glucose Deprivation
Death-associated protein kinase 1 (DAPK1) is a pleiotropic hub of a number of networked distributed intracellular processes. Among them, DAPK1 is known to interact with the excitotoxicity driver NMDA receptor (NMDAR), and in sudden pathophysiological conditions of the brain, e.g., stroke, several li...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7761126/ https://www.ncbi.nlm.nih.gov/pubmed/33265962 http://dx.doi.org/10.3390/antiox9121202 |
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author | DeGregorio-Rocasolano, Núria Guirao, Verónica Ponce, Jovita Melià-Sorolla, Marc Aliena-Valero, Alicia García-Serran, Alexia Salom, Juan B. Dávalos, Antoni Martí-Sistac, Octavi Gasull, Teresa |
author_facet | DeGregorio-Rocasolano, Núria Guirao, Verónica Ponce, Jovita Melià-Sorolla, Marc Aliena-Valero, Alicia García-Serran, Alexia Salom, Juan B. Dávalos, Antoni Martí-Sistac, Octavi Gasull, Teresa |
author_sort | DeGregorio-Rocasolano, Núria |
collection | PubMed |
description | Death-associated protein kinase 1 (DAPK1) is a pleiotropic hub of a number of networked distributed intracellular processes. Among them, DAPK1 is known to interact with the excitotoxicity driver NMDA receptor (NMDAR), and in sudden pathophysiological conditions of the brain, e.g., stroke, several lines of evidence link DAPK1 with the transduction of glutamate-induced events that determine neuronal fate. In turn, DAPK1 expression and activity are known to be affected by the redox status of the cell. To delineate specific and differential neuronal DAPK1 interactors in stroke-like conditions in vitro, we exposed primary cultures of rat cortical neurons to oxygen/glucose deprivation (OGD), a condition that increases reactive oxygen species (ROS) and lipid peroxides. OGD or control samples were co-immunoprecipitated separately, trypsin-digested, and proteins in the interactome identified by high-resolution LC-MS/MS. Data were processed and curated using bioinformatics tools. OGD increased total DAPK1 protein levels, cleavage into shorter isoforms, and dephosphorylation to render the active DAPK1 form. The DAPK1 interactome comprises some 600 proteins, mostly involving binding, catalytic and structural molecular functions. OGD up-regulated 190 and down-regulated 192 candidate DAPK1-interacting proteins. Some differentially up-regulated interactors related to NMDAR were validated by WB. In addition, a novel differential DAPK1 partner, LRRFIP1, was further confirmed by reverse Co-IP. Furthermore, LRRFIP1 levels were increased by pro-oxidant conditions such as ODG or the ferroptosis inducer erastin. The present study identifies novel partners of DAPK1, such as LRRFIP1, which are suitable as targets for neuroprotection. |
format | Online Article Text |
id | pubmed-7761126 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-77611262020-12-26 Comparative Proteomics Unveils LRRFIP1 as a New Player in the DAPK1 Interactome of Neurons Exposed to Oxygen and Glucose Deprivation DeGregorio-Rocasolano, Núria Guirao, Verónica Ponce, Jovita Melià-Sorolla, Marc Aliena-Valero, Alicia García-Serran, Alexia Salom, Juan B. Dávalos, Antoni Martí-Sistac, Octavi Gasull, Teresa Antioxidants (Basel) Article Death-associated protein kinase 1 (DAPK1) is a pleiotropic hub of a number of networked distributed intracellular processes. Among them, DAPK1 is known to interact with the excitotoxicity driver NMDA receptor (NMDAR), and in sudden pathophysiological conditions of the brain, e.g., stroke, several lines of evidence link DAPK1 with the transduction of glutamate-induced events that determine neuronal fate. In turn, DAPK1 expression and activity are known to be affected by the redox status of the cell. To delineate specific and differential neuronal DAPK1 interactors in stroke-like conditions in vitro, we exposed primary cultures of rat cortical neurons to oxygen/glucose deprivation (OGD), a condition that increases reactive oxygen species (ROS) and lipid peroxides. OGD or control samples were co-immunoprecipitated separately, trypsin-digested, and proteins in the interactome identified by high-resolution LC-MS/MS. Data were processed and curated using bioinformatics tools. OGD increased total DAPK1 protein levels, cleavage into shorter isoforms, and dephosphorylation to render the active DAPK1 form. The DAPK1 interactome comprises some 600 proteins, mostly involving binding, catalytic and structural molecular functions. OGD up-regulated 190 and down-regulated 192 candidate DAPK1-interacting proteins. Some differentially up-regulated interactors related to NMDAR were validated by WB. In addition, a novel differential DAPK1 partner, LRRFIP1, was further confirmed by reverse Co-IP. Furthermore, LRRFIP1 levels were increased by pro-oxidant conditions such as ODG or the ferroptosis inducer erastin. The present study identifies novel partners of DAPK1, such as LRRFIP1, which are suitable as targets for neuroprotection. MDPI 2020-11-30 /pmc/articles/PMC7761126/ /pubmed/33265962 http://dx.doi.org/10.3390/antiox9121202 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article DeGregorio-Rocasolano, Núria Guirao, Verónica Ponce, Jovita Melià-Sorolla, Marc Aliena-Valero, Alicia García-Serran, Alexia Salom, Juan B. Dávalos, Antoni Martí-Sistac, Octavi Gasull, Teresa Comparative Proteomics Unveils LRRFIP1 as a New Player in the DAPK1 Interactome of Neurons Exposed to Oxygen and Glucose Deprivation |
title | Comparative Proteomics Unveils LRRFIP1 as a New Player in the DAPK1 Interactome of Neurons Exposed to Oxygen and Glucose Deprivation |
title_full | Comparative Proteomics Unveils LRRFIP1 as a New Player in the DAPK1 Interactome of Neurons Exposed to Oxygen and Glucose Deprivation |
title_fullStr | Comparative Proteomics Unveils LRRFIP1 as a New Player in the DAPK1 Interactome of Neurons Exposed to Oxygen and Glucose Deprivation |
title_full_unstemmed | Comparative Proteomics Unveils LRRFIP1 as a New Player in the DAPK1 Interactome of Neurons Exposed to Oxygen and Glucose Deprivation |
title_short | Comparative Proteomics Unveils LRRFIP1 as a New Player in the DAPK1 Interactome of Neurons Exposed to Oxygen and Glucose Deprivation |
title_sort | comparative proteomics unveils lrrfip1 as a new player in the dapk1 interactome of neurons exposed to oxygen and glucose deprivation |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7761126/ https://www.ncbi.nlm.nih.gov/pubmed/33265962 http://dx.doi.org/10.3390/antiox9121202 |
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