Wild-type PINK1 Prevents Basal and Induced Neuronal Apoptosis, a Protective Effect Abrogated by Parkinson Disease-related Mutations

Abstract

Mutations in the PTEN-induced kinase 1 (PINK1) gene have recently been implicated in autosomal recessive early onset Parkinson Disease (1, 2). To investigate the role of PINK1 in neurodegeneration, we designed human and murine neuronal cell lines expressing either wild-type PINK1 or PINK1 bearing a mutation associated with Parkinson Disease. We show that under basal and staurosporine-induced conditions, the number of terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL)-positive cells was lower in wild-type PINK1 expressing SH-SY5Y cells than in mock-transfected cells. This phenotype was due to a PINK1-mediated reduction in cytochrome c release from mitochondria, which prevents subsequent caspase-3 activation. We show that overexpression of wild-type PINK1 strongly reduced both basal and staurosporine-induced caspase 3 activity. Overexpression of wild-type PINK1 also reduced the levels of cleaved caspase-9, caspase-3, caspase-7, and activated poly(ADP-ribose) polymerase under both basal and staurosporine-induced conditions. In contrast, Parkinson disease-related mutations and a kinase-inactive mutation in PINK1 abrogated the protective effect of PINK1. Together, these results suggest that PINK1 reduces the basal neuronal pro-apoptotic activity and protects neurons from staurosporine-induced apoptosis. Loss of this protective function may therefore underlie the degeneration of nigral dopaminergic neurons in patients with PINK1 mutations. Parkinson disease (PD)2 is the most common neurodegenerative movement disorder, affecting ϳ1% of the population by age 65 years (3, 4). It is characterized by the predominant degeneration of midbrain dopaminergic neurons. Although most patients with PD are sporadic, familial cases represent ϳ10% of all diagnoses. To date, six genes responsible for inherited forms of PD have been identified. Mutations in the ␣-synuclein (5), LRRK2 (leucine-rich repeat kinase 2) and UCH-L1 (ubiquitin C-terminal esterase L1) genes cause dominant forms of familial PD. In contrast, mutations in parkin (6), DJ-1 (7,8), and the newly identified PTEN (phosphatase and tensin homologue on chromosome 10)-induced kinase 1 (PINK1) (1, 2) are responsible for recessive forms of familial PD.PINK1 encodes a highly conserved, 581-amino acid, putative serinethreonine protein kinase and is a member of a small family of novel kinases including CLIK1 (CLP-36 interacting kinase)/PDLIM1 kinases. Bioinformatic analysis suggests that residues Gly-193 to Leu-507 comprise the catalytic domain, residues Gly-193 to Lys-219 form the ATPbinding cassette (with Tyr-166 as an autophosphorylated regulatory residue), and residues Asp-384 to Glu-417 form an activation loop (Fig. 1). PINK1 is transcriptionally transactivated by the PTEN gene (9) and is expressed at variable levels in different cancer cell types. Valente et al. showed that overexpressed, epitope-tagged PINK1 localized to mitochondria and may have a protective function against cell death (1).To further investigate the role of PINK1 in neuronal ap...

Revisiting oxidative stress and mitochondrial dysfunction in the pathogenesis of Parkinson disease—resemblance to the effect of amphetamine drugs of abuse

“…These mutants also showed reduced mitochondrial mass with disorganized morphology. Moreover, PINK1 seems to exert neuroprotective properties, as shown by Petit et al(2005) [248]where WT, but not mutant PINK1 attenuated staurosporineͲinduced apoptosis and reduced mitochondrial cytochrome c release when overexpressed in SHͲSY5Y cells. Also, silencing of PINK1 increased susceptibility to MPP+ or rotenone [248].…”

“…Moreover, PINK1 seems to exert neuroprotective properties, as shown by Petit et al(2005) [248]where WT, but not mutant PINK1 attenuated staurosporineͲinduced apoptosis and reduced mitochondrial cytochrome c release when overexpressed in SHͲSY5Y cells. Also, silencing of PINK1 increased susceptibility to MPP+ or rotenone [248]. In this context, decreased expression of PINK1 in human dopaminergic neurons led to reduced longͲterm survival, along with morphological / structural mitochondrial abnormalities and higher levels of oxidative stress [249].…”

E2F1-dependent miR-421 regulates mitochondrial fragmentation and myocardial infarction by targeting Pink1

“…Inhibition of Pink1 induces fragmented mitochondria, altered cristae morphology and reduced membrane potential 34 . Overexpression of Pink1 rescues mitochondrial fragmentation 35 , increases mitochondrial interconnectivity 12 and reduces mitochondrial dysfunction and apoptosis as well 36 . Our present study for the first time reveals that Pink1 can maintain the cardiac structure and function through its protective effect on the mitochondria.…”

Reprint of: Revisiting oxidative stress and mitochondrial dysfunction in the pathogenesis of Parkinson disease—resemblance to the effect of amphetamine drugs of abuse

“…These mutants also showed reduced mitochondrial mass with disorganized morphology. Moreover, PINK1 seems to exert neuroprotective properties, as shown by Petit et al [248], in which WT, but not mutant PINK1, attenuated staurosporine-induced apoptosis and reduced mitochondrial cytochrome c release when overexpressed in SH-SY5Y cells. Also, silencing of PINK1 increased susceptibility to MPP þ or rotenone [248].…”

“…Moreover, PINK1 seems to exert neuroprotective properties, as shown by Petit et al [248], in which WT, but not mutant PINK1, attenuated staurosporine-induced apoptosis and reduced mitochondrial cytochrome c release when overexpressed in SH-SY5Y cells. Also, silencing of PINK1 increased susceptibility to MPP þ or rotenone [248]. In this context, decreased expression of PINK1 in human dopaminergic neurons led to reduced long-term survival, along with morphological/structural mitochondrial abnormalities and higher levels of oxidative stress [249].…”