<i>C-elegans</i> model identifies genetic modifiers of alpha-synuclein inclusion formation during aging

Inclusions in the brain containing alpha-synuclein are the pathological hallmark of Parkinson's disease, but how these inclusions are formed and how this links to disease is poorly understood. We have developed a &lt;i&gt;C-elegans&lt;/i&gt; model that makes it possible to monitor, in living animals, the formation of alpha-synuclein inclusions. In worms of old age, inclusions contain aggregated alpha-synuclein, resembling a critical pathological feature. We used genome-wide RNA interference to identify processes involved in inclusion formation, and identified 80 genes that, when knocked down, resulted in a premature increase in the number of inclusions. Quality control and vesicle-trafficking genes expressed in the ER/Golgi complex and vesicular compartments were overrepresented, indicating a specific role for these processes in alpha-synuclein inclusion formation. Suppressors include aging-associated genes, such as sir-2.1/SIRT1 and lagr-1/LASS2. Altogether, our data suggest a link between alpha-synuclein inclusion formation and cellular aging, likely through an endomembrane-related mechanism. The processes and genes identified here present a framework for further study of the disease mechanism and provide candidate susceptibility genes and drug targets for Parkinson's disease and other alpha-synuclein related disorders

Improved Immunodetection of Endogenous α-Synuclein

α-Synuclein is a key molecule in understanding the pathogenesis of neurodegenerative α-synucleinopathies such as Parkinson's disease. Despite extensive research, however, its precise function remains unclear partly because of a difficulty in immunoblotting detection of endogenous α-synuclein. This difficulty has largely restricted the progress for α-synucleinopathy research. Here, we report that α-synuclein monomers tend to easily detach from blotted membranes, resulting in no or very poor detection. To prevent this detachment, a mild fixation of blotted membranes with paraformaldehyde was applied to the immunoblotting method. Amazingly, this fixation led to clear and strong detection of endogenous α-synuclein, which has been undetectable by a conventional immunoblotting method. Specifically, we were able to detect endogenous α-synuclein in various human cell lines, including SH-SY5Y, HEK293, HL60, HeLa, K562, A375, and Daoy, and a mouse cell line B16 as well as in several mouse tissues such as the spleen and kidney. Moreover, it should be noted that we could clearly detect endogenous α-synuclein phosphorylated at Ser-129 in several human cell lines. Thus, in some tissues and cultured cells, endogenous α-synuclein becomes easily detectable by simply fixing the blotted membranes. This improved immunoblotting method will allow us to detect previously undetectable endogenous α-synuclein, thereby facilitating α-synuclein research

Site-specific perturbations of alpha-synuclein fibril structure by the Parkinson's disease associated mutations A53T and E46K.

PMCID: PMC3591419This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Parkinson's disease (PD) is pathologically characterized by the presence of Lewy bodies (LBs) in dopaminergic neurons of the substantia nigra. These intracellular inclusions are largely composed of misfolded α-synuclein (AS), a neuronal protein that is abundant in the vertebrate brain. Point mutations in AS are associated with rare, early-onset forms of PD, although aggregation of the wild-type (WT) protein is observed in the more common sporadic forms of the disease. Here, we employed multidimensional solid-state NMR experiments to assess A53T and E46K mutant fibrils, in comparison to our recent description of WT AS fibrils. We made de novo chemical shift assignments for the mutants, and used these chemical shifts to empirically determine secondary structures. We observe significant perturbations in secondary structure throughout the fibril core for the E46K fibril, while the A53T fibril exhibits more localized perturbations near the mutation site. Overall, these results demonstrate that the secondary structure of A53T has some small differences from the WT and the secondary structure of E46K has significant differences, which may alter the overall structural arrangement of the fibrils

TOM40 Mediates Mitochondrial Dysfunction Induced by α-Synuclein Accumulation in Parkinson's Disease.

Alpha-synuclein (α-Syn) accumulation/aggregation and mitochondrial dysfunction play prominent roles in the pathology of Parkinson's disease. We have previously shown that postmortem human dopaminergic neurons from PD brains accumulate high levels of mitochondrial DNA (mtDNA) deletions. We now addressed the question, whether alterations in a component of the mitochondrial import machinery -TOM40- might contribute to the mitochondrial dysfunction and damage in PD. For this purpose, we studied levels of TOM40, mtDNA deletions, oxidative damage, energy production, and complexes of the respiratory chain in brain homogenates as well as in single neurons, using laser-capture-microdissection in transgenic mice overexpressing human wildtype α-Syn. Additionally, we used lentivirus-mediated stereotactic delivery of a component of this import machinery into mouse brain as a novel therapeutic strategy. We report here that TOM40 is significantly reduced in the brain of PD patients and in α-Syn transgenic mice. TOM40 deficits were associated with increased mtDNA deletions and oxidative DNA damage, and with decreased energy production and altered levels of complex I proteins in α-Syn transgenic mice. Lentiviral-mediated overexpression of Tom40 in α-Syn-transgenic mice brains ameliorated energy deficits as well as oxidative burden. Our results suggest that alterations in the mitochondrial protein transport machinery might contribute to mitochondrial impairment in α-Synucleinopathies

Copy number variation in Parkinson's disease

A central theme of human genetic studies is to understand genomic variation and how this underlies the inherited basis of disease. Genomic variation can provide increased biological understanding of disease processes, which is necessary to develop future treatments. Recent technological advances have highlighted the role of copy number variants in normal and pathological phenotypic expression. These applications have been used in studies of Parkinson's disease, a common, late-onset, progressive neurodegenerative disorder. At present the main therapeutic approach is administration of symptom-alleviating drugs, which neither reverses the disease process nor halts its progression. However, the generation of in vivo model systems and development of novel disease intervention strategies for Parkinson's disease have come from research on monogenic forms of the disorder, including those caused by copy number variants. Here, we review the role of copy number variants and the mechanistic insights they have provided on the pathogenesis of Parkinson's disease

Growth charts for children with Ellis–van Creveld syndrome

Ellis–van Creveld (EvC) syndrome is a congenital malformation syndrome with marked growth retardation. In this study, specific growth charts for EvC patients were derived to allow better follow-up of growth and earlier detection of growth patterns unusual for EvC. With the use of 235 observations of 101 EvC patients (49 males, 52 females), growth charts for males and females from 0 to 20 years of age were derived. Longitudinal and cross-sectional data were collected from an earlier review of growth data in EvC, a database of EvC patients, and from recent literature. To model the growth charts, the GAMLSS package for the R statistical program was used. Height of EvC patients was compared to healthy children using Dutch growth charts. Data are presented both on a scale for age and on a scale for the square root of age. Compared to healthy Dutch children, mean height standard deviation score values for male and female EvC patients were −3.1 and −3.0, respectively. The present growth charts should be useful in the follow-up of EvC patients. Most importantly, early detection of growth hormone deficiency, known to occur in EvC, will be facilitated

LRRK2 in Parkinson's disease – drawing the curtain of penetrance: a commentary

Parkinson's disease is the most common neurodegenerative movement disorder and affects about 2% of the population over the age of 60 years. In 2004, mutations in the LRRK2 gene were first described and turned out to be the most frequent genetic cause of familial and sporadic Parkinson's disease and may account for up to 40% of patients in distinct populations. Based on these findings, Latourelle and colleagues show that the penetrance of the most common LRRK2 mutation is higher in patients with familial compared with sporadic Parkinson's disease and identified a substantial number of affected relatives of mutation carriers not presenting with a LRRK2 mutation themselves. This commentary discusses the role of genetic and/or environmental susceptibility factors modulating the expressivity of the disease trait, how these factors may contribute to the phenomenon of phenocopies in genetically defined Parkinson's disease pedigrees, and how the findings of Latourelle and colleagues, published this month in BMC Medicine, relate to current concepts of genetic counselling

Spinocerebellar ataxia type 17: Report of a family with reduced penetrance of an unstable Gln(49 )TBP allele, haplotype analysis supporting a founder effect for unstable alleles and comparative analysis of SCA17 genotypes

BACKGROUND: Spinocerebellar ataxia type 17 (SCA17), a neurodegenerative disorder in man, is caused by an expanded polymorphic polyglutamine-encoding trinucleotide repeat in the gene for TATA-box binding protein (TBP), a main transcription factor. Observed pathogenic expansions ranged from 43 – 63 glutamine (Gln) codons (Gln(43–63)). Reduced penetrance is known for Gln(43–48 )alleles. In the vast majority of families with SCA17 an expanded CAG repeat interrupted by a CAA CAG CAA element is inherited stably. RESULTS: Here, we report the first pedigree with a Gln(49 )allele that is a) not interrupted, b) unstable upon transmission, and c) associated with reduced penetrance or very late age of onset. The 76-year-old father of two SCA17 patients carries the Gln(49 )TBP allele but presents without obvious neurological symptoms. His children with Gln(53 )and Gln(52 )developed ataxia at the age of 41 and 50. Haplotype analysis of this and a second family both with uninterrupted expanded and unstable pathological SCA17 alleles revealed a common core genotype not present in the interrupted expansion of an unrelated SCA17 patient. Review of the literature did not present instability in SCA17 families with expanded alleles interrupted by the CAA CAG CAA element. CONCLUSION: The presence of a Gln(49 )SCA17 allele in an asymptomatic 76-year-old male reams the discussion of reduced penetrance and genotypes producing very late disease onset. In SCA17, uninterrupted expanded alleles of TBP are associated with repeat instability and a common founder haplotype. This suggests for uninterrupted expanded alleles a mutation mechanism and some clinical genetic features distinct from those alleles interrupted by a CAA CAG CAA element

Does hyperthermia constrain flight duration in a short-distance migrant?

While some migratory birds perform non-stop flights of over 11 000 km, many species only spend around 15% of the day in flight during migration, posing a question as to why flight times for many species are so short. Here, we test the idea that hyperthermia might constrain flight duration (FD) in a short-distance migrant using remote biologging technology to measure heart rate, hydrostatic pressure and body temperature in 19 migrating eider ducks (Somateria mollissima), a short-distance migrant. Our results reveal a stop-and-go migration strategy where migratory flights were frequent (14 flights day(−1)) and short (15.7 min), together with the fact that body temperature increases by 1°C, on average, during such flights, which equates to a rate of heat storage index (HSI) of 4°C h(−1). Furthermore, we could not find any evidence that short flights were limited by heart rate, together with the fact that the numerous stops could not be explained by the need to feed, as the frequency of dives and the time spent feeding were comparatively small during the migratory period. We thus conclude that hyperthermia appears to be the predominant determinant of the observed migration strategy, and suggest that such a physiological limitation to FD may also occur in other species. This article is part of the themed issue ‘Moving in a moving medium: new perspectives on flight’