alpha-Synuclein implicated in Parkinson's disease catalyses the formation of hydrogen peroxide in vitro

Some rare inherited forms of Parkinson's disease (PD) are due to mutations in the gene encoding a 140-amino acid presynaptic protein called alpha-synuclein. In PD, and some other related disorders such as dementia with Lewy bodies, alpha-synuclein accumulates in the brain in the form of fibrillar aggregates, which are found inside the neuronal cytoplasmic inclusions known as Lewy bodies. By means of an electron spin resonance (ESR) spin trapping method, we show here that solutions of full-length alpha-synuclein, and a synthetic peptide fragment of alpha-synuclein corresponding to residues 61-95 (the so-called non-Abeta component or NAC), both liberate hydroxyl radicals upon incubation in vitro followed by the addition of Fe(II). We did not observe this property for the related beta- and gamma-synucleins, which are not found in Lewy bodies, and are not linked genetically to any neurodegenerative disorder. There is abundant evidence for the involvement of free radicals and oxidative stress in the pathogenesis of nigral damage in PD. Our new data suggest that the fundamental molecular mechanism underlying this pathological process could be the production of hydrogen peroxide by alpha-synuclein.     

In utero fate mapping reveals distinct migratory pathways and fates of neurons born in the mammalian basal forebrain

Recent studies suggest that neurons born in the developing basal forebrain migrate long distances perpendicularly to radial glia and that many of these cells reach the developing neocortex. This form of tangential migration, however, has not been demonstrated in vivo, and the sites of origin, pathways of migration and final destinations of these neurons in the postnatal brain are not fully understood. Using ultrasound-guided transplantation in utero, we have mapped the migratory pathways and fates of cells born in the lateral and medial ganglionic eminences (LGE and MGE) in 13.5-day-old mouse embryos. We demonstrate that LGE and MGE cells migrate along different routes to populate distinct regions in the developing brain. We show that LGE cells migrate ventrally and anteriorly, and give rise to the projecting medium spiny neurons in the striatum, nucleus accumbens and olfactory tubercle, and to granule and periglomerular cells in the olfactory bulb. By contrast, we show that the MGE is a major source of neurons migrating dorsally and invading the developing neocortex. MGE cells migrate into the neocortex via the neocortical subventricular zone and differentiate into the transient subpial granule neurons in the marginal zone and into a stable population of GABA-, parvalbumin- or somatostatin-expressing interneurons throughout the cortical plate.     

An analysis of cancer prevention by selenium

The nutritional functions of selenium (Se) are recognized as being due to a number of Se-containing proteins. It is not clear, however, whether any of these function in the anti-tumorigenic effects of Se most of which have been demonstrated for Se exposures greater than those required for selenoprotein expression. Indeed, other anti-tumorigenic mechanisms have been demonstrated for certain Se-metabolites. The Nutritional Prevention of Cancer Trial found supplemental Se (200 microg/day, as Se-enriched yeast) to be associated with significant reductions in cancer risks in subjects with pre-treatment plasma Se concentrations below ca. 120 ng/ml (1.5 nmoles/ml), which level would appear to require food-Se intakes of ca. 1.5 microg/kg body weight/day. However, the putative anti-carcinogenic Se-metabolite(s) should be more relevant than total plasma Se as a supplementation target for cancer prevention. These may be components of the non-protein-bound fraction of Se in plasma, which constitutes 2-4% of total plasma Se.     

Calbindin28kDa is specifically associated with extranuclear constituents of the dense particulate fraction

Recent attempts to understand the function of calbindin28kDa, a widely expressed calcium-binding protein, are confounded by uncertainties over its subcellular location. Using immunoblot analysis of rat brain subregions, we found that the proportion of particulate calbindin28kDa (24-43% of total) was independent of expression level and location. The association of calbindin28kDa with particulate structures appeared to be specific, since it persisted when soluble calbindin28kDa was sequestered by antibodies added before tissue disruption. Moreover, when exogenous calbindin28kDa was added during homogenisation of brain from calbindin28kDa-nullmutant mice, only 10% partitioned to the particulate fraction compared with 33% of endogenous calbindin28kDa in wildtype controls. Confocal microscopy showed that calbindin28kDa was predominantly extranuclear in all tissues analysed (i.e. various brain regions, isolated neurons, and dental enamel epithelium). Dual-label microscopy of neural dense particulate fractions confirmed the extranuclear location of calbindin28kDa and also showed that it partly colocalised with synaptosome and microtubule markers. Using sucrose step gradients, calbindin28kDa was separated from nuclei in parallel with synaptosome and endoplasmic reticulum markers. However, no association with the marker proteins (synaptophysin, ERp29, alpha/beta-tubulin) was detected by calbindin28kDa-immunoprecipitation analysis. Together these findings provide the first consistent picture that calbindin28kDa is located predominantly outside of the nucleus, irrespective of tissue type (neuronal vs. non-neuronal) and experimental approach (biochemical vs morphological). The evidence of a substantial, strong and specific association with insoluble cellular structures challenges the widely held view of calbindin28kDa as a mobile calcium buffer, and supports the existence of important alternative roles that involve target proteins.     

Topoisomerase 3α Is Required for Decatenation and Segregation of Human mtDNA

1. Download high-res image (186KB)
2. Download full-size image

     

Deficiency of a Glycogen Synthase-associated Protein,Epm2aip1, Causes Decreased Glycogen Synthesis and Hepatic Insulin Resistance

Glycogen synthesis is a major component of the insulin response, and defective glycogen synthesis is a major portion of insulin resistance. Insulin regulates glycogen synthase (GS) through incompletely defined pathways that activate the enzyme through dephosphorylation and, more potently, allosteric activation. We identify Epm2aip1 as a GS-associated protein. We show that the absence of Epm2aip1 in mice impairs allosteric activation of GS by glucose 6-phosphate, decreases hepatic glycogen synthesis, increases liver fat, causes hepatic insulin resistance, and protects against age-related obesity. Our work identifies a novel GS-associated GS activity-modulating component of insulin resistance.     

Effects of Experimental Exclusion of Scavengers from Carcasses of Anthrax-Infected Herbivores on Bacillus anthracis Sporulation,Survival, and Distribution

Scavenging of anthrax carcasses has long been hypothesized to play a critical role in the production of the infectious spore stage of Bacillus anthracis after host death, though empirical studies assessing this are lacking. We compared B. anthracis spore production, distribution, and survival at naturally occurring anthrax herbivore carcasses that were either experimentally caged to exclude vertebrate scavengers or left unmanipulated. We found no significant effect of scavengers on soil spore density (P > 0.05). Soil stained with terminally hemorrhaged blood and with nonhemorrhagic fluids exhibited high levels of B. anthracis spore contamination (ranging from 10³ to 10⁸ spores/g), even in the absence of vertebrate scavengers. At most of the carcass sites, we also found that spore density in samples taken from hemorrhagic-fluid-stained soil continued to increase for >4 days after host death. We conclude that scavenging by vertebrates is not a critical factor in the life cycle of B. anthracis and that anthrax control measures relying on deterrence or exclusion of vertebrate scavengers to prevent sporulation are unlikely to be effective.     

Identification of a serine protease inhibitor which causes inclusion vacuole reduction and is lethal to Chlamydia trachomatis

The mechanistic details of the pathogenesis of Chlamydia, an obligate intracellular pathogen of global importance, have eluded scientists due to the scarcity of traditional molecular genetic tools to investigate this organism. Here we report a chemical biology strategy that has uncovered the first essential protease for this organism. Identification and application of a unique CtHtrA inhibitor (JO146) to cultures of Chlamydia resulted in a complete loss of viable elementary body formation. JO146 treatment during the replicative phase of development resulted in a loss of Chlamydia cell morphology, diminishing inclusion size, and ultimate loss of inclusions from the host cells. This completely prevented the formation of viable Chlamydia elementary bodies. In addition to its effect on the human Chlamydia trachomatis strain, JO146 inhibited the viability of the mouse strain, Chlamydia muridarum, both in vitro and in vivo. Thus, we report a chemical biology approach to establish an essential role for Chlamydia CtHtrA. The function of CtHtrA for Chlamydia appears to be essential for maintenance of cell morphology during replicative the phase and these findings provide proof of concept that proteases can be targeted for antimicrobial therapy for intracellular pathogens.