Benjamin Dehay

Pathogenic Lysosomal Depletion in Parkinson's Disease

Mounting evidence suggests a role for autophagy dysregulation in Parkinsons disease (PD). The bulk degradation of cytoplasmic proteins (including α-synuclein) and organelles (such as mitochondria) is mediated by macroautophagy, which involves the sequestration of cytosolic components into autophagosomes (AP) and its delivery to lysosomes. Accumulation of AP occurs in postmortem brain samples from PD patients, which has been widely attributed to an induction of autophagy. However, the cause and pathogenic significance of these changes remain unknown. Here we found in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model of PD that AP accumulation and dopaminergic cell death are preceded by a marked decrease in the amount of lysosomes within dopaminergic neurons. Lysosomal depletion was secondary to the abnormal permeabilization of lysosomal membranes induced by increased mitochondrial-derived reactive oxygen species. Lysosomal permeabilization resulted in a defective clearance and subsequent accumulation of undegraded AP and contributed directly to neurodegeneration by the ectopic release of lysosomal proteases into the cytosol. Lysosomal breakdown and AP accumulation also occurred in PD brain samples, where Lewy bodies were strongly immunoreactive for AP markers. Induction of lysosomal biogenesis by genetic or pharmacological activation of lysosomal transcription factor EB restored lysosomal levels, increased AP clearance and attenuated 1-methyl-4-phenylpyridinium-induced cell death. Similarly, the autophagy-enhancer compound rapamycin attenuated PD-related dopaminergic neurodegeneration, both in vitro and in vivo, by restoring lysosomal levels. Our results indicate that AP accumulation in PD results from defective lysosomal-mediated AP clearance secondary to lysosomal depletion. Restoration of lysosomal levels and function may thus represent a novel neuroprotective strategy in PD.

Lewy body extracts from Parkinson disease brains trigger α‐synuclein pathology and neurodegeneration in mice and monkeys

Mounting evidence suggests that α‐synuclein, a major protein component of Lewy bodies (LB), may be responsible for initiating and spreading the pathological process in Parkinson disease (PD). Supporting this concept, intracerebral inoculation of synthetic recombinant α‐synuclein fibrils can trigger α‐synuclein pathology in mice. However, it remains uncertain whether the pathogenic effects of recombinant synthetic α‐synuclein may apply to PD‐linked pathological α‐synuclein and occur in species closer to humans.

Loss of P-type ATPase ATP13A2/PARK9 function induces general lysosomal deficiency and leads to Parkinson disease neurodegeneration

Parkinson disease (PD) is a progressive neurodegenerative disorder pathologically characterized by the loss of dopaminergic neurons from the substantia nigra pars compacta and the presence, in affected brain regions, of protein inclusions named Lewy bodies (LBs). The ATP13A2 gene (locus PARK9) encodes the protein ATP13A2, a lysosomal type 5 P-type ATPase that is linked to autosomal recessive familial parkinsonism. The physiological function of ATP13A2, and hence its role in PD, remains to be elucidated. Here, we show that PD-linked mutations in ATP13A2 lead to several lysosomal alterations in ATP13A2 PD patient-derived fibroblasts, including impaired lysosomal acidification, decreased proteolytic processing of lysosomal enzymes, reduced degradation of lysosomal substrates, and diminished lysosomal-mediated clearance of autophagosomes. Similar alterations are observed in stable ATP13A2-knockdown dopaminergic cell lines, which are associated with cell death. Restoration of ATP13A2 levels in ATP13A2-mutant/depleted cells restores lysosomal function and attenuates cell death. Relevant to PD, ATP13A2 levels are decreased in dopaminergic nigral neurons from patients with PD, in which ATP13A2 mostly accumulates within Lewy bodies. Our results unravel an instrumental role of ATP13A2 deficiency on lysosomal function and cell viability and demonstrate the feasibility and therapeutic potential of modulating ATP13A2 levels in the context of PD.

Lysosomal Impairment in Parkinson's Disease

Impairment of autophagy‐lysosomal pathways (ALPs) is increasingly regarded as a major pathogenic event in neurodegenerative diseases, including Parkinsons disease (PD). ALP alterations are observed in sporadic PD brains and in toxic and genetic rodent models of PD‐related neurodegeneration. In addition, PD‐linked mutations and post‐translational modifications of α‐synuclein impair its own lysosomal‐mediated degradation, thereby contributing to its accumulation and aggregation. Furthermore, other PD‐related genes, such as leucine‐rich repeat kinase‐2 (LRRK2), parkin, and phosphatase and tensin homolog (PTEN)‐induced putative kinase 1 (PINK1), have been mechanistically linked to alterations in ALPs. Conversely, mutations in lysosomal‐related genes, such as glucocerebrosidase (GBA) and lysosomal type 5 P‐type ATPase (ATP13A2), have been linked to PD. New data offer mechanistic molecular evidence for such a connection, unraveling a causal link between lysosomal impairment, α‐synuclein accumulation, and neurotoxicity. First, PD‐related GBA deficiency/mutations initiate a positive feedback loop in which reduced lysosomal function leads to α‐synuclein accumulation, which, in turn, further decreases lysosomal GBA activity by impairing the trafficking of GBA from the endoplasmic reticulum‐Golgi to lysosomes, leading to neurodegeneration. Second, PD‐related mutations/deficiency in the ATP13A2 gene lead to a general lysosomal impairment characterized by lysosomal membrane instability, impaired lysosomal acidification, decreased processing of lysosomal enzymes, reduced degradation of lysosomal substrates, and diminished clearance of autophagosomes, collectively contributing to α‐synuclein accumulation and cell death. According to these new findings, primary lysosomal defects could potentially account for Lewy body formation and neurodegeneration in PD, laying the groundwork for the prospective development of new neuroprotective/disease‐modifying therapeutic strategies aimed at restoring lysosomal levels and function.

Two molecular pathways initiate mitochondria-dependent dopaminergic neurodegeneration in experimental Parkinson's disease.

Dysfunction of mitochondrial complex I is associated with a wide spectrum of neurodegenerative disorders, including Parkinsons disease (PD). In rodents, inhibition of complex I leads to degeneration of dopaminergic neurons of the substantia nigra pars compacta (SNpc), as seen in PD, through activation of mitochondria-dependent apoptotic molecular pathways. In this scenario, complex I blockade increases the soluble pool of cytochrome c in the mitochondrial intermembrane space through oxidative mechanisms, whereas activation of pro-cell death protein Bax is actually necessary to trigger neuronal death by permeabilizing the outer mitochondrial membrane and releasing cytochrome c into the cytosol. Activation of Bax after complex I inhibition relies on its transcriptional induction and translocation to the mitochondria. How complex I deficiency leads to Bax activation is currently unknown. Using gene-targeted mice, we show that the tumor suppressor p53 mediates Bax transcriptional induction after PD-related complex I blockade in vivo, but it does not participate in Bax mitochondrial translocation in this model, either by a transcription-independent mechanism or through the induction of BH3-only proteins Puma or Noxa. Instead, Bax mitochondrial translocation in this model relies mainly on the JNK-dependent activation of the BH3-only protein Bim. Targeting either Bax transcriptional induction or Bax mitochondrial translocation results in a marked attenuation of SNpc dopaminergic cell death caused by complex I inhibition. These results provide further insight into the pathogenesis of PD neurodegeneration and identify molecular targets of potential therapeutic significance for this disabling neurological illness.

Alpha-synuclein spreading in Parkinson’s disease

Formation and accumulation of misfolded protein aggregates are a central hallmark of several neurodegenerative diseases. In Parkinson’s disease (PD), the aggregation-prone protein alpha-synuclein (α-syn) is the culprit. In the past few years, another piece of the puzzle has been added with data suggesting that α-syn may self-propagate, thereby contributing to the progression and extension of PD. Of particular importance, it was the seminal observation of Lewy bodies (LB), a histopathological signature of PD, in grafted fetal dopaminergic neurons in the striatum of PD patients. Consequently, these findings were a conceptual breakthrough, generating the “host to graft transmission” hypothesis, also called the “prion-like hypothesis.” Several in vitro and in vivo studies suggest that α-syn can undergo a toxic templated conformational change, spread from cell to cell and from region to region, and initiate the formation of “LB–like aggregates,” contributing to the PD pathogenesis. Here, we will review and discuss the current knowledge for such a putative mechanism on the prion-like nature of α-syn, and discuss about the proper use of the term prion-like.

PSD-95 expression controls l-DOPA dyskinesia through dopamine D1 receptor trafficking

L-DOPA-induced dyskinesia (LID), a detrimental consequence of dopamine replacement therapy for Parkinsons disease, is associated with an alteration in dopamine D1 receptor (D1R) and glutamate receptor interactions. We hypothesized that the synaptic scaffolding protein PSD-95 plays a pivotal role in this process, as it interacts with D1R, regulates its trafficking and function, and is overexpressed in LID. Here, we demonstrate in rat and macaque models that disrupting the interaction between D1R and PSD-95 in the striatum reduces LID development and severity. Single quantum dot imaging revealed that this benefit was achieved primarily by destabilizing D1R localization, via increased lateral diffusion followed by increased internalization and diminished surface expression. These findings indicate that altering D1R trafficking via synapse-associated scaffolding proteins may be useful in the treatment of dyskinesia in Parkinsons patients.

Optic atrophy 1 mediates mitochondria remodeling and dopaminergic neurodegeneration linked to complex I deficiency

Mitochondrial complex I dysfunction has long been associated with Parkinson’s disease (PD). Recent evidence suggests that mitochondrial involvement in PD may extend beyond a sole respiratory deficit and also include perturbations in mitochondrial fusion/fission or ultrastructure. Whether and how alterations in mitochondrial dynamics may relate to the known complex I defects in PD is unclear. Optic atrophy 1 (OPA1), a dynamin-related GTPase of the inner mitochondrial membrane, participates in mitochondrial fusion and apoptotic mitochondrial cristae remodeling. Here we show that complex I inhibition by parkinsonian neurotoxins leads to an oxidative-dependent disruption of OPA1 oligomeric complexes that normally keep mitochondrial cristae junctions tight. As a consequence, affected mitochondria exhibit major structural abnormalities, including cristae disintegration, loss of matrix density and swelling. These changes are not accompanied by mitochondrial fission but a mobilization of cytochrome c from cristae to intermembrane space, thereby lowering the threshold for activation of mitochondria-dependent apoptosis by cell death agonists in compromised neurons. All these pathogenic changes, including mitochondrial structural remodeling and dopaminergic neurodegeneration, are abrogated by OPA1 overexpression, both in vitro and in vivo. Our results identify OPA1 as molecular link between complex I deficiency and alterations in mitochondrial dynamics machinery and point to OPA1 as a novel therapeutic target for complex I cytopathies, such as PD.

Lysosomal membrane permeabilization in Parkinson disease.

*Correspondence to: Miquel Vila and Patricia Boya; Email: mvila@ir.vhebron.net and pboya@cib.csic.es Mounting evidence supports a role for autophagy dysregulation in Parkinson disease (PD). For instance, pathogenic variants of α-synuclein have been shown to block chaperone-mediated autophagy (CMA) in vitro, resulting in a reduced degradation of this protein and other CMA substrates. Furthermore, accumulation of autophagosomes (AP) has been observed in post-mortem PD brains, which has been largely attributed to an activation of macroautophagy, perhaps secondary to CMA blockage. However, the actual cause and pathogenic significance of AP accumulation in PD remains unknown. In this context, we have recently observed in an experimental mouse model of PD that AP accumulation and dopaminergic cell death are preceded by an early disruption of lysosomal integrity caused by the abnormal permeabilization of lysosomal membranes through mitochondriallydriven oxidative attack. Besides overloading the system with undegraded AP, lysosomal breakdown directly contributes to dopaminergic neuron cell death by the ectopic release of lysosomal proteases into the cytosol. Lysosomal depletion and AP accumulation also occurred in PD postmortem brain samples, where characteristic intraneuronal cytoplasmic inclusions, called Lewy bodies, were strongly immunoreactive for AP markers. Our results indicate that accumulation of AP in PD mostly results from defective AP clearance because of early lysosomal disruption. Supporting this concept, genetic or pharmacological restoration of lysosomal levels results in increased AP clearance, reduced AP accumulation and attenuated dopaminergic cell death in experimental PD. Lysosomal membrane permeabilization in Parkinson disease

Selective Inactivation of Striatal FosB/ΔFosB-Expressing Neurons Alleviates L-DOPA–Induced Dyskinesia

BACKGROUNDnΔFosB is a surrogate marker of L-DOPA-induced dyskinesia (LID), the unavoidable disabling consequence of Parkinsons disease L-DOPA long-term treatment. However, the relationship between the electrical activity of FosB/ΔFosB-expressing neurons and LID manifestation is unknown.nnnMETHODSnWe used the Daun02 prodrug-inactivation method associated with lentiviral expression of β-galactosidase under the control of the FosB promoter to investigate a causal link between the activity of FosB/ΔFosB-expressing neurons and dyskinesia severity in both rat and monkey models of Parkinsons disease and LID. Whole-cell recordings of medium spiny neurons (MSNs) were performed to assess the effects of Daun02 and daunorubicin on neuronal excitability.nnnRESULTSnWe first show that daunorubicin, the active product of Daun02 metabolism by β-galactosidase, decreases the activity of MSNs in rat brain slices and that Daun02 strongly decreases the excitability of rat MSN primary cultures expressing β-galactosidase upon D1 dopamine receptor stimulation. We then demonstrate that the selective, and reversible, inhibition of FosB/ΔFosB-expressing striatal neurons with Daun02 decreases the severity of LID while improving the beneficial effect of L-DOPA.nnnCONCLUSIONSnThese results establish that FosB/ΔFosB accumulation ultimately results in altered neuronal electrical properties sustaining maladaptive circuits leading not only to LID but also to a blunted response to L-DOPA. These findings further reveal that targeting dyskinesia can be achieved without reducing the antiparkinsonian properties of L-DOPA when specifically inhibiting FosB/ΔFosB-accumulating neurons.