Aggregated alpha-synuclein proteins form brain lesions that are hallmarks of neurodegenerative synucleinopathies, and oxidative stress has been implicated in the pathogenesis of some of these disorders. Using antibodies to specific nitrated tyrosine residues in alpha-synuclein, we demonstrate extensive and widespread accumulations of nitrated alpha-synuclein in the signature inclusions of Parkinson's disease, dementia with Lewy bodies, the Lewy body variant of Alzheimer's disease, and multiple system atrophy brains. We also show that nitrated alpha-synuclein is present in the major filamentous building blocks of these inclusions, as well as in the insoluble fractions of affected brain regions of synucleinopathies. The selective and specific nitration of alpha-synuclein in these disorders provides evidence to directly link oxidative and nitrative damage to the onset and progression of neurodegenerative synucleinopathies.

Neuronal and oligodendrocytic aggregates of fibrillar ␣-synuclein define several diseases of the nervous system. It is likely that these inclusions impair vital metabolic processes and compromise vialibity of affected cells. Here, we report that a 12-amino acid stretch ( 71 VT-GVTAVAQKTV 82 ) in the middle of the hydrophobic domain of human ␣-synuclein is necessary and sufficient for its fibrillization based on the following observations: 1) human ␤-synuclein is highly homologous to ␣-synuclein but lacks these 12 residues, and it does not assemble into filaments in vitro; 2) the rate of ␣-synuclein polymerization in vitro decreases after the introduction of a single charged amino acid within these 12 residues, and a deletion within this region abrogates assembly; 3) this stretch of 12 amino acids appears to form the core of ␣-synuclein filaments, because it is resistant to proteolytic digestion in ␣-synuclein filaments; and 4) synthetic peptides corresponding to this 12-amino acid stretch self-polymerize to form filaments, and these peptides promote fibrillization of full-length human ␣-synuclein in vitro. Thus, we have identified key sequence elements necessary for the assembly of human ␣-synuclein into filaments, and these elements may be exploited as targets for the design of drugs that inhibit ␣-synuclein fibrillization and might arrest disease progression.

No abstract

Alpha-synuclein (alpha-syn) is the major component of intracellular inclusions in several neurodegenerative diseases, and the conversion of soluble alpha-syn into filamentous aggregates may contribute to disease pathogenesis. Since mechanisms leading to the formation of alpha-syn inclusions are unclear, in vitro models of alpha-syn aggregation may yield insights into this process. To that end, we examined the consequences on the progressive deletion of the carboxy-terminus of alpha-syn in regulating fibril formation, and we show here that carboxy-terminal truncated alpha-syn proteins aggregate faster than the full-length molecule. Protease digestion and immunoelectron microscopy indicate that the alpha-syn amino- and carboxy-termini are more solvent exposed than the central core and that filaments formed from carboxy-terminal truncated alpha-syn are narrower in diameter than the full-length molecule. Moreover, seeding experiments under conditions where full-length alpha-syn did not readily aggregate revealed that carboxy-truncated alpha-syn extending from amino acids 1-102 and 1-110 but not 1-120 were efficient in seeding full-length alpha-syn aggregation over a range of concentrations. Using site-directed mutagenesis, the negatively charged residues 104, 105 and 114, 115 in the carboxy-terminus were implicated in this reduced aggregation and the lack of seeding of full-length alpha-syn fibrillogenesis by 1-120. Our data support the view that the middle region of alpha-syn forms the core of alpha-syn filaments and that negative charges in the carboxy-terminus counteract alpha-syn aggregation. Thus, the carboxy-terminus of alpha-syn may regulate aggregation of full-length alpha-syn and determine the diameter of alpha-syn filaments.

SUMMARYFatty acid derivatives are of central importance for plant immunity against insect herbivores; however, major regulatory genes and the signals that modulate these defense metabolites are vastly understudied, especially in important agro-economic monocot species. Here we show that products and signals derived from a single Zea mays (maize) lipoxygenase (LOX), ZmLOX10, are critical for both direct and indirect defenses to herbivory. We provide genetic evidence that two 13-LOXs, ZmLOX10 and ZmLOX8, specialize in providing substrate for the green leaf volatile (GLV) and jasmonate (JA) biosynthesis pathways, respectively. Supporting the specialization of these LOX isoforms, LOX8 and LOX10 are localized to two distinct cellular compartments, indicating that the JA and GLV biosynthesis pathways are physically separated in maize. Reduced expression of JA biosynthesis genes and diminished levels of JA in lox10 mutants indicate that LOX10-derived signaling is required for LOX8-mediated JA. The possible role of GLVs in JA signaling is supported by their ability to partially restore wound-induced JA levels in lox10 mutants. The impaired ability of lox10 mutants to produce GLVs and JA led to dramatic reductions in herbivore-induced plant volatiles (HIPVs) and attractiveness to parasitoid wasps. Because LOX10 is under circadian rhythm regulation, this study provides a mechanistic link to the diurnal regulation of GLVs and HIPVs. GLV-, JA-and HIPV-deficient lox10 mutants display compromised resistance to insect feeding, both under laboratory and field conditions, which is strong evidence that LOX10-dependent metabolites confer immunity against insect attack. Hence, this comprehensive gene to agro-ecosystem study reveals the broad implications of a single LOX isoform in herbivore defense.

Mutations in the beta-synuclein gene may predispose to DLB.

Senile plaques in the cerebral parenchyma are a pathognomonic feature of Alzheimer's disease (AD) and are mainly composed of aggregated fibrillar amyloid β (Aβ) proteins. The plaques are associated with neuronal degeneration, lipid membrane abnormalities, and chemical evidence of oxidative stress. The view that Aβ proteins cause these pathological changes has been challenged by suggestions that they have a protective function or that they are merely byproducts of the pathological process. This investigation was conducted to determine whether Aβ proteins promote or inhibit oxidative damage to lipid membranes. Using a mass spectrometric assay of oxidative lipid damage, the 42-residue form of Aβ (Aβ42) was found to accelerate the oxidative lipid damage caused by physiological concentrations of ascorbate and submicromolar concentrations of copper(II) ion. Under these conditions, Aβ42 was aggregated, but nonfibrillar. Ascorbate and copper produced H 2 O 2 , but Aβ42 reduced H 2 O 2 concentrations, and its ability to accelerate oxidative damage was not affected by catalase. Lipids could be oxidized by H 2 O 2 and copper-(II) in the absence of ascorbate, but only at significantly higher concentrations, and Aβ42 inhibited this reaction. These results indicate that the ability of Aβ42 to promote oxidative damage is more potent and more likely to be manifest in vivo than its ability to inhibit oxidative damage. In conjunction with prior results demonstrating that oxidatively damaged membranes cause Aβ42 to misfold and form fibrils, these results suggest a specific chemical mechanism linking Aβ42-promoted oxidative lipid damage to amyloid fibril formation.

Amyloid β proteins and oxidative stress are believed to have central roles in the development of Alzheimer's disease. Lipid membranes are among the most vulnerable cellular components to oxidative stress, and membranes in susceptible regions of the brain are compositionally distinct from those in other tissues. This review considers the evidence that membranes are either a source of neurotoxic lipid oxidation products or the target of pathogenic processes involving amyloid β proteins that cause permeability changes or ion channel formation. Progress toward a comprehensive theory of Alzheimer's disease pathogenesis is discussed in which lipid membranes assume both roles and promote the conversion of monomeric amyloid β proteins into fibrils, the pathognomonic histopathological lesion of the disease.