**Alpha-Synuclein, Oxidative Stress and Autophagy Failure: Dangerous Liaisons in Dopaminergic Neurodegeneration**

Giovanni Stefanoni1,2, Gessica Sala1, Lucio Tremolizzo1,2, Laura Brighina1,2 and Carlo Ferrarese1,2

*1Department of Neuroscience and Biomedical Technologies, University of Milano-Bicocca 2Department of Neurology, San Gerardo Hospital, Monza. Italy* 

### **1. Introduction**

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The molecular mechanisms of neurodegeneration in Parkinson's disease and the cause of the selective dopaminergic neuronal loss are mostly unknown. Many pathogenetic factors have been found to play a role but the relationships among these factors, together with the reasons of the high vulnerability of dopaminergic neurons to them, have not been completely defined. Only a small fraction of Parkinson's disease cases have a defined etiology: this fraction include the monogenic hereditary variants of the disease and the sporadic cases determined by prolonged exposition to toxic agents inhibiting mitochondrial complex I, such as 1,1'-dimethyl-4,4'-5 bipyridinium (paraquat), rotenone and 1-methyl-4 phenyl-1,2,3,6-tetrahydropyridine (MPTP). Parkinson's disease-related toxins and pathogenetic mutations have been indispensable to create cell and animal models with the aim to clarify the molecular physiopathology of the disease. Little is known about the primitive causes of idiopathic Parkinson's disease, that probably represents a multi-factorial disease influenced by various genetic and environmental factors, all characterized by high incidence in general population. The different risk factors together would contribute to initiate the complex pathogenetic sequence of events leading to the death of dopaminergic neurons.

Recently, Parkinson's disease has been placed in the large category of neurodegenerative diseases caused by protein misfolding. In particular, alpha-synuclein has been proposed as the central and most specific factor implied in the pathogenesis of this syndrome, which, as a consequence, has been classified among synucleinopathies, together with dementia with Lewy bodies and multiple system atrophy, other neurodegenerative diseases having alphasynuclein pathology as a major feature.

Aim of this chapter is to provide an organic revision of evidences for the involvement of alpha-synuclein in the pathogenesis of Parkinson's disease. We will define the mechanisms responsible for the toxic gain of function of α-synuclein and the processes triggered by aberrant alpha-synuclein and mediating its neurotoxic effect. Particular attention will be paid to establish the links that correlate the deleterious action of alpha-synuclein with oxidative stress and with the efficiency of the processes involved in the clearance of aberrant

Alpha-Synuclein, Oxidative Stress and

bodies in dopaminergic neurons of mouse models.

neurotoxic effect of the protein (Vekrellis et al 2004):

conformation and to promote protein aggregation.

• Binding to microtubules and dysregulation of cytoskeleton functions.

gene have proved to modify the risk of developing Parkinson's disease.

• Damaging of mitochondrial and other cellular membranes. • Interaction with other proteins and modulation of their activity.

transgenic animal models.

mutations.

Autophagy Failure: Dangerous Liaisons in Dopaminergic Neurodegeneration 113

Ala53Thr alpha-synuclein mutant is responsible for the greater in vivo neurotoxicity in

The deleterious effects of these genetic alterations on dopaminergic neurons have been demonstrated in cellular and animal models, confirming the hypothesis that both qualitative and quantitative alterations of alpha-synuclein are able to trigger its toxic effect: mutant alpha-synuclein protein exerts a selective toxicity in dopaminergic neurons when expressed in rats; at the same time, the mutant protein causes aggregation and formation of Lewy

The toxicity of both mutant and wild type alpha-synuclein seems to require the acquisition of a misfolded conformation which prevents alpha-synuclein degradation and favors its fibrillization, firstly into protofibrillar oligomeric species and then to fibrillar aggregates. Recently, it has become clear that insoluble aggregates probably do not have an intrinsic toxic function, as suggested by the finding that Lewy bodies–positive dopaminergic neurons are less vulnerable to degeneration. Furthermore, the increased size of Lewy bodies and the decreased levels of soluble alpha-synuclein correlate with a higher resistance to cytotoxic agents. Therefore, formation of Lewy bodies could represent a protective phenomenon favoring the removal of soluble oligomers. Different mechanisms can explain the negative effects exerted by alpha-synuclein oligomers, which seem to be directly responsible for the

Alpha-synuclein oligomers directly interfere with proteasome subunits. Furthermore, alphasynuclein impairs microtubule stability through binding to alpha-tubulin and to the microtubule associated protein TAU. Alpha synuclein induces the aggregation of TAU, which in turn favors alpha-synuclein fibrillization; interestingly, polymorphisms in the TAU

In monogenic forms of Parkinson's disease the direct neurotoxic effect of mutant alphasynuclein may depend on specific conformational modifications, modulating the propensity of the protein to aggregate, to interact with other proteins and to be processed by clearance mechanisms. Pathogenetic mutations of alpha-synuclein protein are known to confer a tendency to acquire abnormal conformation substantially easier than the wild type protein, which accounts for the development of Parkinson's disease in patients having these

The fact that over-expressed wild type alpha-synuclein is responsible for neurotoxicity suggests that even simple protein accumulation is able to trigger the misfolded

As well as in familial cases, even in sporadic Parkinson's disease the toxic gain of function of alpha-synuclein could derive from the intraneuronal accumulation of the protein or from biochemical modifications enhancing the propensity of the protein to aggregate. Anyway, the ubiquitous expression and the high levels of alpha-synuclein in the brain suggests that its simple presence in dopaminergic neurons is not sufficient to explain the selective degeneration of these cells, which must have specific characteristics able to induce or to accelerate alpha-synuclein misfolding, oligomerization and aggregation. Oxidative stress and production of highly reactive aldehydes, both depending on high levels of dopamine, can modify alpha-synuclein, partially explaining the high tendency of this protein to acquire

proteins; these last include the ubiquitin-proteasome system and the autophagic-lysosomal pathways. Both oxidative stress and impairment of protein degradation machinery exert a neurotoxic effect contributing to the pathogenesis of Parkinson's disease and partially mediated, in dopaminergic neurons, by qualitative and quantitative alterations in alphasynuclein. This protein in turn is able to boost the processes responsible for its toxic gain of function. Therefore, a physiopathologic circuit emerges constituted by a complex interaction among different pathogenetic mechanisms, everyone able to support the others; the death of dopaminergic neurons represents the ultimate and irreversible outcome of all these events. This chapter aims at clarifying this complex dynamics responsible for the neurodegenerative process, with particular regard to mechanisms both determining and mediating the deleterious effect of alpha-synuclein. Furthermore, we will report data on the development of biochemical tests helpful for diagnosis and prognosis and useful as objective criteria to determine the neuroprotective effectiveness of drugs in clinical trials: in this regard, levels and modifications of α-synuclein, as well as parameters of oxidative stress and autophagy, could represent suitable peripheral biomarkers of disease risk and progression. Finally, we will discuss about the opportunity of interfering with all the reported pathogenetic mechanisms as putative neuroprotective pharmacologic strategy.
