FOXO3 determines the accumulation of alpha-synuclein and controls the fate of dopaminergic neurons in the substantia nigra

Parkinson's disease (PD) is characterized by the selective degeneration of neuronal populations presumably due to pathogenic interactions between aging and predisposing factors such as increased levels of a-synuclein. Here, we genetically modulate the activity of the transcription factor Forkhead box protein O-3 (FOXO3) in adult nigral dopaminergic neurons using viral vectors and explore how this determinant of longevity impacts on neuronal fate in normal and diseased conditions. We find that dopaminergic neurons are particularly vulnerable to changes in FOXO3 activity in the substantia nigra. While constitutive activation has proapoptotic effects leading to neuronal loss, inhibition of FOXO-mediated transcription by a dominant-negative competitor causes oxidative damage and is detrimental at high vector dose. To address the role of FOXO3 in PD, we modulate its activity in dopaminergic neurons overexpressing human a-synuclein. In this pathogenic condition, we find that FOXO inhibition has protective effects, suggesting that this transcription factor ultimately contributes to neuronal cell death. Nevertheless, mild FOXO3 activity also protects nigral neurons against the accumulation of human a-synuclein, albeit to a lesser extent. FOXO3 reduces the amount of a-synuclein present in the soluble protein fraction and promotes the coalescence of dense proteinase K-resistant aggregates, with an accumulation of autophagic vacuoles containing lipofuscin. Consistent with these in vivo observations, we find that FOXO3 controls autophagic flux in neuronal cells. Altogether, these results point to FOXO3 as an important determinant of neuronal survival in the substantia nigra, which may oppose a-synuclein accumulation and proteotoxicity.

Conditional expression of Parkinson's disease-related R1441C LRRK2 in midbrain dopaminergic neurons of mice causes nuclear abnormalities without neurodegeneration

Shi-Yan Caroline Foo, Liliane Glauser, Meghna Kannan, Graham Knott, Darren Moore, Elpida Tsika

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene cause late-onset, autosomal dominant Parkinson's disease (PD). The clinical and neurochemical features of LRRK2-linked PD are similar to idiopathic disease although neuropathology is somewhat heterogeneous. Dominant mutations in LRRK2 precipitate neurodegeneration through a toxic gain-of-function mechanism which can be modeled in transgenic mice overexpressing human LRRK2 variants. A number of LRRK2 transgenic mouse models have been developed that display abnormalities in dopaminergic neurotransmission and alterations in tau metabolism yet without consistently inducing dopaminergic neurodegeneration. To directly explore the impact of mutant LRRK2 on the nigrostriatal dopaminergic pathway, we developed conditional transgenic mice that selectively express human R1441C LRRK2 in dopaminergic neurons from the endogenous murine ROSA26 promoter. The expression of R1441C LRRK2 does not induce the degeneration of substantia nigra dopaminergic neurons or striatal dopamine deficits in mice up to 2 years of age, and fails to precipitate abnormal protein inclusions containing alpha-synuclein, tau, ubiquitin or autophagy markers (LC3 and p62). Furthermore, mice expressing R1441C LRRK2 exhibit normal motor activity and olfactory function with increasing age. Intriguingly, the expression of R1441C LRRK2 induces age-dependent abnormalities of the nuclear envelope in nigral dopaminergic neurons including reduced nuclear circularity and increased invaginations of the nuclear envelope. In addition, R1441C LRRK2 mice display increased neurite complexity of cultured midbrain dopaminergic neurons. Collectively, these novel R1441C LRRK2 conditional transgenic mice reveal altered dopaminergic neuronal morphology with advancing age, and provide a useful tool for exploring the pathogenic mechanisms underlying the R1441C LRRK2 mutation in PD. (C) 2014 Elsevier Inc All rights reserved.

Elsevier2014

PGC-1 alpha activity in nigral dopamine neurons determines vulnerability to alpha-synuclein

Wojciech Janusz Bobela, Graham Knott, Bernard Schneider, Lu Zheng

Introduction: Mitochondrial dysfunction and oxidative stress are critical factors in the pathogenesis of age-dependent neurodegenerative diseases. PGC-1 alpha, a master regulator of mitochondrial biogenesis and cellular antioxidant defense, has emerged as a possible therapeutic target for Parkinson's disease, with important roles in the function and survival of dopaminergic neurons in the substantia nigra. The objective of this study is to determine if the loss of PGC-1 alpha activity contributes to alpha-synuclein-induced degeneration. Results: We explore the vulnerability of PGC-1 alpha null mice to the accumulation of human alpha-synuclein in nigral neurons, and assess the neuroprotective effect of AAV-mediated PGC-1 alpha expression in this experimental model. Using neuronal cultures derived from these mice, mitochondrial respiration and production of reactive oxygen species are assessed in conditions of human alpha-synuclein overexpression. We find ultrastructural evidence for abnormal mitochondria and fragmented endoplasmic reticulum in the nigral dopaminergic neurons of PGC-1 alpha null mice. Furthermore, PGC-1 alpha null nigral neurons are more prone to degenerate following overexpression of human alpha-synuclein, an effect more apparent in male mice. PGC-1 alpha overexpression restores mitochondrial morphology, oxidative stress detoxification and basal respiration, which is consistent with the observed neuroprotection against alpha-synuclein toxicity in male PGC-1 alpha null mice. Conclusions: Altogether, our results highlight an important role for PGC-1 alpha in controlling the mitochondrial function of nigral neurons accumulating alpha-synuclein, which may be critical for gender-dependent vulnerability to Parkinson's disease.

Biomed Central Ltd2015

Inhibition of alpha-synuclein fibrillization by dopamine is mediated by interactions with five C-terminal residues and with E83 in the NAC region

Paolo Carloni, Alessandra Chesi, Hilal Lashuel, Adrian Schmid

The interplay between dopamine and alpha-synuclein (AS) plays a central role in Parkinson's disease (PD). PD results primarily from a severe and selective devastation of dopaminergic neurons in substantia nigra pars compacta. The neuropathological hallmark of the disease is the presence of intraneuronal proteinaceous inclusions known as Lewy bodies within the surviving neurons, enriched in filamentous AS. In vitro, dopamine inhibits AS fibril formation, but the molecular determinants of this inhibition remain obscure. Here we use molecular dynamic (MD) simulations to investigate the binding of dopamine and several of its derivatives onto conformers representative of an NMR ensemble of AS structures in aqueous solution. Within the limitations inherent to MD simulations of unstructured proteins, our calculations suggest that the ligands bind to the (125)YEMPS(129) region, consistent with experimental findings. The ligands are further stabilized by long-range electrostatic interactions with glutamate 83 (E83) in the NAC region. These results suggest that by forming these interactions with AS, dopamine may affect AS aggregation and fibrillization properties. To test this hypothesis, we investigated in vitro the effects of dopamine on the aggregation of mutants designed to alter or abolish these interactions. We found that point mutations in the (125)YEMPS(129) region do not affect AS aggregation, which is consistent with the fact that dopamine interacts non-specifically with this region. In contrast, and consistent with our modeling studies, the replacement of glutamate by alanine at position 83 (E83A) abolishes the ability of dopamine to inhibit AS fibrillization.

Public Library of Science2008

Conditional expression of Parkinson's disease-related R1441C LRRK2 in midbrain dopaminergic neurons of mice causes nuclear abnormalities without neurodegeneration

Shi-Yan Caroline Foo, Liliane Glauser, Meghna Kannan, Graham Knott, Darren Moore, Elpida Tsika

Elsevier2014