title: Neuroserpin polymers cause oxidative stress in a neuronal model of the dementia FENIB.
abstract: The serpinopathies are human pathologies caused by mutations that promote polymerisation and intracellular deposition of proteins of the serpin superfamily, leading to a poorly understood cell toxicity. The dementia FENIB is caused by polymerisation of the neuronal serpin neuroserpin (NS) within the endoplasmic reticulum (ER) of neurons. With the aim of understanding the toxicity due to intracellular accumulation of neuroserpin polymers, we have generated transgenic neural progenitor cell (NPC) cultures from mouse foetal cerebral cortex, stably expressing the control protein GFP (green fluorescent protein), or human wild type, G392E or delta NS. We have characterised these cell lines in the proliferative state and after differentiation to neurons. Our results show that G392E NS formed polymers that were mostly retained within the ER, while wild type NS was correctly secreted as a monomeric protein into the culture medium. Delta NS was absent at steady state due to its rapid degradation, but it was easily detected upon proteasomal block. Looking at their intracellular distribution, wild type NS was found in partial co-localisation with ER and Golgi markers, while G392E NS was localised within the ER only. Furthermore, polymers of NS were detected by ELISA and immunofluorescence in neurons expressing the mutant but not the wild type protein. We used control GFP and G392E NPCs differentiated to neurons to investigate which cellular pathways were modulated by intracellular polymers by performing RNA sequencing. We identified 747 genes with a significant upregulation (623) or downregulation (124) in G392E NS-expressing cells, and we focused our attention on several genes involved in the defence against oxidative stress that were up-regulated in cells expressing G392E NS (Aldh1b1, Apoe, Gpx1, Gstm1, Prdx6, Scara3, Sod2). Inhibition of intracellular anti-oxidants by specific pharmacological reagents uncovered the damaging effects of NS polymers. Our results support a role for oxidative stress in the cellular toxicity underlying the neurodegenerative dementia FENIB.
authors.affiliation: Dpt. of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy.
authors.forename: Noemi A
authors.initials: NA
authors.lastname: Guadagno
authors.affiliation: Dpt. of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy.
authors.forename: Claudia
authors.initials: C
authors.lastname: Moriconi
authors.affiliation: Dpt. of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy; Institute for Systems Analysis and Computer Science 'Antonio Ruberti', National Research Council, Rome, Italy.
authors.forename: Valerio
authors.initials: V
authors.lastname: Licursi
authors.affiliation: Dpt. of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy.
authors.forename: Emanuela
authors.initials: E
authors.lastname: D'Acunto
authors.affiliation: Dpt. of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy.
authors.forename: Paola S
authors.initials: PS
authors.lastname: Nisi
authors.affiliation: Dpt. of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy.
authors.forename: Nicoletta
authors.initials: N
authors.lastname: Carucci
authors.affiliation: Dpt. of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy.
authors.forename: Antonella
authors.initials: A
authors.lastname: De Jaco
authors.affiliation: Dpt. of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy.
authors.forename: Emanuele
authors.initials: E
authors.lastname: Cacci
authors.affiliation: Dpt. of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy; Institute of Biology and Molecular Pathology (IBPM), National Research Council, Rome, Italy.
authors.forename: Rodolfo
authors.initials: R
authors.lastname: Negri
authors.affiliation: Dpt. of Chemistry, Sapienza University of Rome, Italy. Electronic address: giuseppe.lupo@uniroma1.it.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Lupo
authors.affiliation: Dpt. of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy; Pasteur Institute - Cenci Bolognetti Foundation, Sapienza University of Rome, Italy. Electronic address: mariaelena.mirandabanos@uniroma1.it.
authors.forename: Elena
authors.initials: E
authors.lastname: Miranda
mesh_terms: D000818:Animals; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D003704:Dementia / Q000139:chemically induced / Q000378:metabolism* / Q000473:pathology; D004831:Epilepsies, Myoclonic / Q000139:chemically induced / Q000378:metabolism* / Q000473:pathology; D020271:Heredodegenerative Disorders, Nervous System / Q000139:chemically induced / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D009474:Neurons / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D009479:Neuropeptides / Q000633:toxicity*; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D011108:Polymers / Q000633:toxicity*; D015843:Serpins / Q000633:toxicity*; D000097522:Neuroserpin
chemical_list: D009479:Neuropeptides; D011108:Polymers; D015843:Serpins; D000097522:Neuroserpin
keywords: Familial encephalopathy with neuroserpin inclusion bodies; Neural progenitor cells; Neurodegeneration; Neurons; Neuroserpin; Oxidative stress; Protein conformational disease; Serpin polymers