Cellular Oligomerization of α-Synuclein Is Determined by the Interaction of Oxidized Catechols with a C-terminal Sequence

Abstract

The mechanisms that govern the formation of ␣-synuclein (␣-syn) aggregates are not well understood but are considered a central event in the pathogenesis of Parkinson's disease (PD). A critically important modulator of ␣-syn aggregation in vitro is dopamine and other catechols, which can prevent the formation of ␣-syn aggregates in cell-free and cellular model systems. Despite the profound importance of this interaction for the pathogenesis of PD, the processes by which catechols alter ␣-syn aggregation are unclear. Molecular and biochemical approaches were employed to evaluate the mechanism of catechol-␣-syn interactions and the effect on inclusion formation. The data show that the intracellular inhibition of ␣-syn aggregation requires the oxidation of catechols and the specific noncovalent interaction of the oxidized catechols with residues 125 YEMPS 129 in the C-terminal region of the protein. Cell-free studies using novel near infrared fluorescence methodology for the detection of covalent proteinortho-quinone adducts showed that although covalent modification of ␣-syn occurs, this does not affect ␣-syn fibril formation. In addition, oxidized catechols are unable to prevent both thermal and acid-induced protein aggregation as well as fibrils formed from a protein that lacks a YEMPS amino acid sequence, suggesting a specific effect for ␣-syn. These results suggest that inappropriate C-terminal cleavage of ␣-syn, which is known to occur in vivo in PD brain or a decline of intracellular catechol levels might affect disease progression, resulting in accelerated ␣-syn inclusion formation and dopaminergic neurodegeneration. ␣-Synuclein (␣-syn)2 (NACP, synelfin), a small, neuron-specific protein, was first linked to PD by genetic analysis of families with autosomal dominant inheritance of the disease. Genetic analysis discovered a point mutation in the ␣-syn gene (SNCA), resulting in an amino acid conversion of Ala 53 to Thr (1). Subsequently, ␣-syn protein was detected in Lewy bodies within the dopaminergic neurons of the substantia nigra pars compacta (2), the intracellular proteinaceous inclusions characteristic of PD and related disorders. Since this discovery, the process of ␣-syn aggregation has been proposed to underlie dopaminergic degeneration that occurs in PD. Therefore, delineating the mechanisms of ␣-syn aggregation and its pathophysiological role in neurodegeneration has been the focus of many investigations.Although the in vivo factors that regulate ␣-syn aggregation are not well understood, mechanisms involving genetic and environmental factors have been proposed. Genetic analysis has uncovered two additional missense mutations in SNCA (A30P and E46K) (3, 4) as well as triplication of the SNCA genomic region (5). Mutations of ␣-syn or gene triplication may increase the rate of ␣-syn aggregation (6, 7), impair cellular degradation (8), or increase the amount of cytosolic ␣-syn beyond the critical concentration required to initiate polymerization (9). Most PD cases are sporadic and involve aggregation...

Oxidative and nitrative protein modifications in Parkinson's disease

“…Dopamine-modified α-synuclein not only inhibited its own degradation by the CMA but also impairs the degradation of other proteins suggesting why dopaminergic cells may be particularly sensitive to various forms of stress leading to selective SN neurodegeneration in PD patients [58]. More recent data has suggested that dopamine oxidation products on alpha-synuclein may not be derived by adduct formation but rather via noncovalent interaction of oxidized catechols with the protein which may result in its inappropriate C-terminal cleavage and/or conformational changes that impact on its oligomerization [59,60].…”

Protein Aggregation in the Brain: The Molecular Basis for Alzheimer’s and Parkinson’s Diseases

“…Much of this α-synuclein has been posttranslationally modified by sequence truncation at the C-terminus, phosphorylation at Ser129 (120,121), or nitration (122); these modifications have been shown to increase the rate at which oligomers are formed (121,123). Dopamine and related catecholamines have been shown to interact with α-synuclein and stabilize the protofibril stage of aggregation (124,125), providing a possible explanation for the increased susceptibility of dopaminergic neurons. Levels of soluble oligomers of α-synuclein are also affected by fatty acids, being upregulated by polyunsaturated fatty acids (126).…”

Melanocyte-like cells in the heart and pulmonary veins contribute to atrial arrhythmia triggers

“…The melanin synthesis enzyme dopachrome tautomerase (Dct) contributes to, but is not required for, melanin synthesis in dermal melanocytes by catalyzing the conversion of L-dopachrome to DHICA (29). In the absence of Dct, there is decreased DHICA formation with concomitant DHI build up, which promotes the accumulation of reactive species (30,31). Furthermore, Dct is important for intracellular calcium regulation; in fact, eumelanin binds calcium with an affinity similar to calmodulin (32).…”