title: Bcl-xL-mediated antioxidant function abrogates the disruption of mitochondrial dynamics induced by LRRK2 inhibition.
abstract: We have used the human neuroblastoma cell line SH-SY5Y overexpressing Bcl-xL (SH-SY5Y/Bcl-xL) to clarify the effects of this mitochondrial protein on the control of mitochondrial dynamics and the autophagic processes which occur after the inhibition of leucine-rich repeat kinase 2 (LRRK2) with GSK2578215A. In wild type (SH-SY5Y/Neo) cells, GSK2578215A (1nM) caused a disruption of mitochondrial morphology and an imbalance in intracellular reactive oxygen species (ROS) as indicated by an increase in dichlorofluorescein fluorescence and 4-hydroxynonenal. However, SH-SY5Y/Bcl-xL cells under GSK2578215A treatment, unlike the wild type, preserved a high mitochondrial membrane potential and did not exhibit apoptotical chromatins. In contrast to wild type cells, in SH-SY5Y/Bcl-xL cells, GSK2578215A did not induce mitochondrial translocation of neither dynamin related protein-1 nor the proapoptotic protein, Bax. In SH-SY5Y/Neo, but not SH-SY5Y/Bcl-xL cells, mitochondrial fragmentation elicited by GSK2578215A precedes an autophagic response. Furthermore, the overexpression of Bcl-xL protein restores the autophagic flux pathway disrupted by this inhibitor. SH-SY5Y/Neo, but not SH-SY5Y/Bcl-xL cells, responded to LRRK2 inhibition by an increase in the levels of acetylated tubulin, indicating that this was abrogated by Bcl-xL overexpression. This hyperacetylation of tubulin took place earlier than any of the above-mentioned events suggesting that it is involved in the autophagic flux interruption. Pre-treatment with tempol prevented the GSK2578215A-induced mitochondrial fragmentation, autophagy and the rise in acetylated tubulin in SH-SY5Y/Neo cells. Thus, these data support the notion that ROS act as a second messenger connexion between LRRK2 inhibition and these deleterious responses, which are markedly alleviated by the Bcl-xL-mediated ROS generation blockade.
authors.affiliation: Grupo de Neurofarmacología, Dpto. Ciencias Médicas, Facultad de Medicina de Albacete, Universidad de Castilla-La Mancha, IDINE, Albacete, Spain; Facultad de Medicina y Odontología, Universidad Católica de Valencia ¨San Vicente Mártir, Valencia, Spain; Unidad de Neuropsicofarmacología Traslacional, Complejo Hospitalario Universitario de Albacete, Albacete, Spain.
authors.forename: Sara
authors.initials: S
authors.lastname: Saez-Atienzar
authors.affiliation: Facultad de Medicina y Odontología, Universidad Católica de Valencia ¨San Vicente Mártir, Valencia, Spain.
authors.forename: Luis
authors.initials: L
authors.lastname: Bonet-Ponce
authors.affiliation: Grupo de Neurofarmacología, Dpto. Ciencias Médicas, Facultad de Medicina de Albacete, Universidad de Castilla-La Mancha, IDINE, Albacete, Spain; Unidad de Neuropsicofarmacología Traslacional, Complejo Hospitalario Universitario de Albacete, Albacete, Spain.
authors.forename: Carmen
authors.initials: C
authors.lastname: da Casa
authors.affiliation: Grupo de Neurofarmacología, Dpto. Ciencias Médicas, Facultad de Medicina de Albacete, Universidad de Castilla-La Mancha, IDINE, Albacete, Spain; Unidad de Neuropsicofarmacología Traslacional, Complejo Hospitalario Universitario de Albacete, Albacete, Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Perez-Dolz
authors.affiliation: Facultad de Medicina y Odontología, Universidad Católica de Valencia ¨San Vicente Mártir, Valencia, Spain.
authors.forename: Jose R
authors.initials: JR
authors.lastname: Blesa
authors.affiliation: Grupo de Neurofarmacología, Dpto. Ciencias Médicas, Facultad de Medicina de Albacete, Universidad de Castilla-La Mancha, IDINE, Albacete, Spain.
authors.forename: Eduardo
authors.initials: E
authors.lastname: Nava
authors.affiliation: Unidad de Neuropsicofarmacología Traslacional, Complejo Hospitalario Universitario de Albacete, Albacete, Spain. Electronic address: mgalindoa@sescam.jccm.es.
authors.forename: Maria F
authors.initials: MF
authors.lastname: Galindo
authors.affiliation: Grupo de Neurofarmacología, Dpto. Ciencias Médicas, Facultad de Medicina de Albacete, Universidad de Castilla-La Mancha, IDINE, Albacete, Spain. Electronic address: joaquin.jordan@uclm.es.
authors.forename: Joaquín
authors.initials: J
authors.lastname: Jordan
mesh_terms: D000107:Acetylation; D001343:Autophagy; D045744:Cell Line, Tumor; D006801:Humans; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D063154:Mitochondrial Dynamics; D018384:Oxidative Stress; D010300:Parkinson Disease / Q000378:metabolism; D014404:Tubulin / Q000378:metabolism; D051020:bcl-X Protein / Q000378:metabolism*
chemical_list: D014404:Tubulin; D051020:bcl-X Protein; C495280:LRRK2 protein, human; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2
keywords: Acetylated tubulin; DRP-1; LRRK2; Mitocondrial fission; Oxidative stress; Parkinson