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Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons
BACKGROUND: Mutations in leucine-rich repeat kinase 2 (LRRK2) contribute to both familial and idiopathic forms of Parkinson’s disease (PD). Neuroinflammation is a key event in neurodegeneration and aging, and there is mounting evidence of LRRK2 involvement in inflammatory pathways. In a previous stu...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5116223/ https://www.ncbi.nlm.nih.gov/pubmed/27863501 http://dx.doi.org/10.1186/s12974-016-0761-x |
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author | López de Maturana, Rakel Lang, Valérie Zubiarrain, Amaia Sousa, Amaya Vázquez, Nerea Gorostidi, Ana Águila, Julio López de Munain, Adolfo Rodríguez, Manuel Sánchez-Pernaute, Rosario |
author_facet | López de Maturana, Rakel Lang, Valérie Zubiarrain, Amaia Sousa, Amaya Vázquez, Nerea Gorostidi, Ana Águila, Julio López de Munain, Adolfo Rodríguez, Manuel Sánchez-Pernaute, Rosario |
author_sort | López de Maturana, Rakel |
collection | PubMed |
description | BACKGROUND: Mutations in leucine-rich repeat kinase 2 (LRRK2) contribute to both familial and idiopathic forms of Parkinson’s disease (PD). Neuroinflammation is a key event in neurodegeneration and aging, and there is mounting evidence of LRRK2 involvement in inflammatory pathways. In a previous study, we described an alteration of the inflammatory response in dermal fibroblasts from PD patients expressing the G2019S and R1441G mutations in LRRK2. METHODS: Taking advantage of cellular reprogramming, we generated induced pluripotent stem cell (iPSC) lines and neurons thereafter, harboring LRRK2(G2019S) and LRRK2(R1441G) mutations. We used gene silencing and functional reporter assays to characterize the effect of the mutations. We examined the temporal profile of TNFα-induced changes in proteins of the NF-κB pathway and optimized western blot analysis to capture α-synuclein dynamics. The effects of the mutations and interventions were analyzed by two-way ANOVA tests with respect to corresponding controls. RESULTS: LRRK2 silencing decreased α-synuclein protein levels in mutated neurons and modified NF-κB transcriptional targets, such as PTGS2 (COX-2) and TNFAIP3 (A20). We next tested whether NF-κB and α-synuclein pathways converged and found that TNFα modulated α-synuclein levels, although we could not detect an effect of LRRK2 mutations, partly because of the individual variability. Nevertheless, we confirmed NF-κB dysregulation in mutated neurons, as shown by a protracted recovery of IκBα and a clear impairment in p65 nuclear translocation in the LRRK2 mutants. CONCLUSIONS: Altogether, our results show that LRRK2 mutations affect α-synuclein regulation and impair NF-κB canonical signaling in iPSC-derived neurons. TNFα modulated α-synuclein proteostasis but was not modified by the LRRK2 mutations in this paradigm. These results strengthen the link between LRRK2 and the innate immunity system underscoring the involvement of inflammatory pathways in the neurodegenerative process in PD. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12974-016-0761-x) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-5116223 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-51162232016-11-25 Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons López de Maturana, Rakel Lang, Valérie Zubiarrain, Amaia Sousa, Amaya Vázquez, Nerea Gorostidi, Ana Águila, Julio López de Munain, Adolfo Rodríguez, Manuel Sánchez-Pernaute, Rosario J Neuroinflammation Research BACKGROUND: Mutations in leucine-rich repeat kinase 2 (LRRK2) contribute to both familial and idiopathic forms of Parkinson’s disease (PD). Neuroinflammation is a key event in neurodegeneration and aging, and there is mounting evidence of LRRK2 involvement in inflammatory pathways. In a previous study, we described an alteration of the inflammatory response in dermal fibroblasts from PD patients expressing the G2019S and R1441G mutations in LRRK2. METHODS: Taking advantage of cellular reprogramming, we generated induced pluripotent stem cell (iPSC) lines and neurons thereafter, harboring LRRK2(G2019S) and LRRK2(R1441G) mutations. We used gene silencing and functional reporter assays to characterize the effect of the mutations. We examined the temporal profile of TNFα-induced changes in proteins of the NF-κB pathway and optimized western blot analysis to capture α-synuclein dynamics. The effects of the mutations and interventions were analyzed by two-way ANOVA tests with respect to corresponding controls. RESULTS: LRRK2 silencing decreased α-synuclein protein levels in mutated neurons and modified NF-κB transcriptional targets, such as PTGS2 (COX-2) and TNFAIP3 (A20). We next tested whether NF-κB and α-synuclein pathways converged and found that TNFα modulated α-synuclein levels, although we could not detect an effect of LRRK2 mutations, partly because of the individual variability. Nevertheless, we confirmed NF-κB dysregulation in mutated neurons, as shown by a protracted recovery of IκBα and a clear impairment in p65 nuclear translocation in the LRRK2 mutants. CONCLUSIONS: Altogether, our results show that LRRK2 mutations affect α-synuclein regulation and impair NF-κB canonical signaling in iPSC-derived neurons. TNFα modulated α-synuclein proteostasis but was not modified by the LRRK2 mutations in this paradigm. These results strengthen the link between LRRK2 and the innate immunity system underscoring the involvement of inflammatory pathways in the neurodegenerative process in PD. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12974-016-0761-x) contains supplementary material, which is available to authorized users. BioMed Central 2016-11-18 /pmc/articles/PMC5116223/ /pubmed/27863501 http://dx.doi.org/10.1186/s12974-016-0761-x Text en © The Author(s). 2016 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 López de Maturana, Rakel Lang, Valérie Zubiarrain, Amaia Sousa, Amaya Vázquez, Nerea Gorostidi, Ana Águila, Julio López de Munain, Adolfo Rodríguez, Manuel Sánchez-Pernaute, Rosario Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons |
title | Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons |
title_full | Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons |
title_fullStr | Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons |
title_full_unstemmed | Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons |
title_short | Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons |
title_sort | mutations in lrrk2 impair nf-κb pathway in ipsc-derived neurons |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5116223/ https://www.ncbi.nlm.nih.gov/pubmed/27863501 http://dx.doi.org/10.1186/s12974-016-0761-x |
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