title: Mitochondrial impairment increases FL-PINK1 levels by calcium-dependent gene expression.
abstract: Mutations of the PTEN-induced kinase 1 (PINK1) gene are a cause of autosomal recessive Parkinson's disease (PD). This gene encodes a mitochondrial serine/threonine kinase, which is partly localized to mitochondria, and has been shown to play a role in protecting neuronal cells from oxidative stress and cell death, perhaps related to its role in mitochondrial dynamics and mitophagy. In this study, we report that increased mitochondrial PINK1 levels observed in human neuroblastoma SH-SY5Y cells after carbonyl cyanide m-chlorophelyhydrazone (CCCP) treatment were due to de novo protein synthesis, and not just increased stabilization of full length PINK1 (FL-PINK1). PINK1 mRNA levels were significantly increased by 4-fold after 24h. FL-PINK1 protein levels at this time point were significantly higher than vehicle-treated, or cells treated with CCCP for 3h, despite mitochondrial content being decreased by 29%. We have also shown that CCCP dissipated the mitochondrial membrane potential (Δψm) and induced entry of extracellular calcium through L/N-type calcium channels. The calcium chelating agent BAPTA-AM impaired the CCCP-induced PINK1 mRNA and protein expression. Furthermore, CCCP treatment activated the transcription factor c-Fos in a calcium-dependent manner. These data indicate that PINK1 expression is significantly increased upon CCCP-induced mitophagy in a calcium-dependent manner. This increase in expression continues after peak Parkin mitochondrial translocation, suggesting a role for PINK1 in mitophagy that is downstream of ubiquitination of mitochondrial substrates. This sensitivity to intracellular calcium levels supports the hypothesis that PINK1 may also play a role in cellular calcium homeostasis and neuroprotection.
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Departamento de Bioquímica y Biología Molecular y Genética, Universidad de Extremadura, F. Enfermería y Terapia Ocupacional, 10003 Cáceres, Spain.
authors.forename: Rubén
authors.initials: R
authors.lastname: Gómez-Sánchez
authors.affiliation: Department of Clinical Neurosciences, Institute of Neurology, University College London, London NW3 2PF, UK.
authors.forename: Matthew E
authors.initials: ME
authors.lastname: Gegg
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Departamento de Bioquímica y Biología Molecular y Genética, Universidad de Extremadura, F. Enfermería y Terapia Ocupacional, 10003 Cáceres, Spain; INSERM, U848, Institut Gustave Roussy, Université Paris Sud, Paris 11, F-94805 Villejuif, France.
authors.forename: José M
authors.initials: JM
authors.lastname: Bravo-San Pedro
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Departamento de Bioquímica y Biología Molecular y Genética, Universidad de Extremadura, F. Enfermería y Terapia Ocupacional, 10003 Cáceres, Spain; INSERM, U848, Institut Gustave Roussy, Université Paris Sud, Paris 11, F-94805 Villejuif, France.
authors.forename: Mireia
authors.initials: M
authors.lastname: Niso-Santano
authors.affiliation: Department of Clinical Neurosciences, Institute of Neurology, University College London, London NW3 2PF, UK.
authors.forename: Lydia
authors.initials: L
authors.lastname: Alvarez-Erviti
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Departamento de Bioquímica y Biología Molecular y Genética, Universidad de Extremadura, F. Enfermería y Terapia Ocupacional, 10003 Cáceres, Spain.
authors.forename: Elisa
authors.initials: E
authors.lastname: Pizarro-Estrella
authors.affiliation: Servicio de Técnicas Aplicadas a las Biociencias, Universidad de Extremadura, 06071 Badajoz, Spain.
authors.forename: Yolanda
authors.initials: Y
authors.lastname: Gutiérrez-Martín
authors.affiliation: Servicio de Técnicas Aplicadas a las Biociencias, Universidad de Extremadura, 06071 Badajoz, Spain.
authors.forename: Alberto
authors.initials: A
authors.lastname: Alvarez-Barrientos
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Departamento de Bioquímica y Biología Molecular y Genética, Universidad de Extremadura, F. Enfermería y Terapia Ocupacional, 10003 Cáceres, Spain. Electronic address: jfuentes@unex.es.
authors.forename: José M
authors.initials: JM
authors.lastname: Fuentes
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Departamento de Bioquímica y Biología Molecular y Genética, Universidad de Extremadura, F. Enfermería y Terapia Ocupacional, 10003 Cáceres, Spain. Electronic address: rosapolo@unex.es.
authors.forename: Rosa Ana
authors.initials: RA
authors.lastname: González-Polo
authors.affiliation: Department of Clinical Neurosciences, Institute of Neurology, University College London, London NW3 2PF, UK.
authors.forename: Anthony H V
authors.initials: AH
authors.lastname: Schapira
mesh_terms: D001343:Autophagy / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000633:toxicity; D045744:Cell Line, Tumor; D015870:Gene Expression; D006801:Humans; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism*; D063306:Mitophagy / Q000187:drug effects / Q000502:physiology; D009447:Neuroblastoma / Q000201:enzymology / Q000378:metabolism; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism; D061209:Proton Ionophores / Q000633:toxicity; D000099202:PTEN-Induced Putative Kinase
chemical_list: D002118:Calcium; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D011494:Protein Kinases; D016760:Proto-Oncogene Proteins c-fos; D061209:Proton Ionophores; D000099202:PTEN-Induced Putative Kinase
keywords: CCCP; Calcium; Mitophagy; PINK1; Parkinson's disease; SH-SY5Y