title: Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons.
abstract: 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.
  
  
  
  Taking advantage of cellular reprogramming, we generated induced pluripotent stem cell (iPSC) lines and neurons thereafter, harboring LRRK2G2019Sand LRRK2R1441Gmutations. 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.
  
  
  
  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.
  
  
  
  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.
authors.affiliation: Laboratory of Stem Cells and Neural Repair, Inbiomed, Paseo Mikeletegi, 81, E-20009, San Sebastian, Spain.
authors.forename: Rakel
authors.initials: R
authors.lastname: López de Maturana
authors.affiliation: Laboratory of Ubiquitylation and Cancer Molecular Biology, Inbiomed, San Sebastian, Spain.
authors.forename: Valérie
authors.initials: V
authors.lastname: Lang
authors.affiliation: Laboratory of Stem Cells and Neural Repair, Inbiomed, Paseo Mikeletegi, 81, E-20009, San Sebastian, Spain.
authors.forename: Amaia
authors.initials: A
authors.lastname: Zubiarrain
authors.affiliation: Laboratory of Stem Cells and Neural Repair, Inbiomed, Paseo Mikeletegi, 81, E-20009, San Sebastian, Spain.
authors.forename: Amaya
authors.initials: A
authors.lastname: Sousa
authors.affiliation: Laboratory of Stem Cells and Neural Repair, Inbiomed, Paseo Mikeletegi, 81, E-20009, San Sebastian, Spain.
authors.forename: Nerea
authors.initials: N
authors.lastname: Vázquez
authors.affiliation: Genomics Platform and Neuroscience Area, Biodonostia Research Institute, San Sebastian, Spain.
authors.forename: Ana
authors.initials: A
authors.lastname: Gorostidi
authors.affiliation: Laboratory of Stem Cells and Neural Repair, Inbiomed, Paseo Mikeletegi, 81, E-20009, San Sebastian, Spain.
authors.forename: Julio
authors.initials: J
authors.lastname: Águila
authors.affiliation: Neurology Department, Donostia Universitary Hospital, Neuroscience Area, Instituto Biodonostia, San Sebastián, Spain.|Center for Biomedical Research Network in Neurodegenerative Diseases (CIBERNED), Institute Carlos III, Ministry of Economy and Competitiveness, Madrid, Spain.|Department of Neurosciences, University of the Basque Country, UPV/EHU, San Sebastian, Spain.
authors.forename: Adolfo
authors.initials: A
authors.lastname: López de Munain
authors.affiliation: Laboratory of Ubiquitylation and Cancer Molecular Biology, Inbiomed, San Sebastian, Spain.
authors.forename: Manuel
authors.initials: M
authors.lastname: Rodríguez
authors.affiliation: Laboratory of Stem Cells and Neural Repair, Inbiomed, Paseo Mikeletegi, 81, E-20009, San Sebastian, Spain. rpernaute@inbiomed.org.
authors.forename: Rosario
authors.initials: R
authors.lastname: Sánchez-Pernaute
mesh_terms: D002454:Cell Differentiation / Q000235:genetics; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism; D004252:DNA Mutational Analysis; D004298:Dopamine / Q000378:metabolism; D005347:Fibroblasts; D005786:Gene Expression Regulation / Q000235:genetics; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D006801:Humans; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2 / Q000235:genetics* / Q000378:metabolism; D009154:Mutation / Q000235:genetics*; D016328:NF-kappa B / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism*; D010300:Parkinson Disease / Q000235:genetics / Q000473:pathology; D039904:Pluripotent Stem Cells / Q000502:physiology*; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D055746:Stage-Specific Embryonic Antigens / Q000378:metabolism; D014162:Transfection; D014404:Tubulin / Q000378:metabolism; D051844:alpha-Synuclein / Q000378:metabolism
chemical_list: D016207:Cytokines; D005904:Glial Fibrillary Acidic Protein; D016328:NF-kappa B; D055746:Stage-Specific Embryonic Antigens; C498585:TUBB3 protein, human; D014404:Tubulin; D051844:alpha-Synuclein; C066584:stage-specific embryonic antigen-4; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; D004298:Dopamine
keywords: Inflammation; LRRK2; NF-κB; Parkinson’s disease; iPSCs; α-Synuclein