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Title: Study of the role of PINK1 in mitochondrial function and morphology
Other Titles: Poster
Authors: Van Den Haute, Chris
Paesen, Kirsten
Coun, Frea
Heeman, Bavo
Coussée, E
Koopman, W J H
De Smet, P
Willems, P H G M
Callewaert, Geert
Debyser, Zeger
Baekelandt, Veerle #
Issue Date: 2007
Conference: Annual Meeting of the Society for Neuroscience edition:37 location:San Diego, USA date:3-7 November 2007
Abstract: Parkinson disease (PD) is the second most common neurodegenerative disorder. Mutations in the recently identified PTEN-induced putative kinase (PINK1) have been linked to familial forms of PD. PINK1 is a serine-threonine kinase with a mitochondrial localization. It is thought that the mutations induce a loss-of-function. However, the exact role of PINK1 in the physiology of the cell is still unknown.
To mimick the mutations and the loss-of-function we induced RNA interference against PINK1 mRNA. By means of a lentiviral vector system expressing short-hairpins against PINK1 we created stable PINK1 knock-down Neuro2a, SHSY-5Y and PC-12 cell lines. Efficient suppression of PINK1 expression was confirmed using quantitative-RT-PCR. We also overexpressed wild-type and two clinical mutants (G309D and L347P) of human PINK1 with our lentiviral vector sytem. We confirmed the overexpression using Western blot.To investigate the effects of PINK1 knock-down and overexpression we focused on mitochondrial Ca2+-metabolism and mitochondrial morphology. Mitochondrial apo-aequorin was transfected into the different cell lines. Apo-aequorin in combination with coelenterazine and Ca2+ results in light emission, which allows to to measure the Ca2+ flux in the mitochondria. Incubation of the cells with Rhodamine 123 was used to visualize the mitochondrial network and to quantify the mitochondrial morphology. Small but significant differences in Ca2+-metabolism and mitochondrial morphology were observed in the knock-down cell lines. This suggests that knock-down of PINK1 affects mitochondrial function in neuronal cells.
Publication status: published
KU Leuven publication type: IMa
Appears in Collections:Research Group for Neurobiology and Gene Therapy
Physiology Section (-)
Molecular Virology and Gene Therapy
# (joint) last author

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