Novel suppressors of α-synuclein toxicity identified using yeast

The mechanism by which the Parkinson’s disease-related protein α-synuclein (α-syn) causes neurodegeneration has not been elucidated. To determine the genes that protect cells from α-syn, we used a genetic screen to identify suppressors of the super sensitivity of the yeast Saccharomyces cerevisiae expressing α-syn to killing by hydrogen peroxide. Forty genes in ubiquitin-dependent protein catabolism, protein biosynthesis, vesicle trafficking and the response to stress were identified. Five of the forty genes—ENT3, IDP3, JEM1, ARG2 and HSP82—ranked highest in their ability to block α-syn-induced reactive oxygen species accumulation, and these five genes were characterized in more detail. The deletion of any of these five genes enhanced the toxicity of α-syn as judged by growth defects compared with wild-type cells expressing α-syn, which indicates that these genes protect cells from α-syn. Strikingly, four of the five genes are specific for α-syn in that they fail to protect cells from the toxicity of the two inherited mutants A30P or A53T. This finding suggests that α-syn causes toxicity to cells through a different pathway than these two inherited mutants. Lastly, overexpression of Ent3p, which is a clathrin adapter protein involved in protein transport between the Golgi and the vacuole, causes α-syn to redistribute from the plasma membrane into cytoplasmic vesicular structures. Our interpretation is that Ent3p mediates the transport of α-syn to the vacuole for proteolytic degradation. A similar clathrin adaptor protein, epsinR, exists in humans

Mutation in Archain 1, a Subunit of COPI Coatomer Complex, Causes Diluted Coat Color and Purkinje Cell Degeneration

Intracellular trafficking is critical for delivering molecules and organelles to their proper destinations to carry out normal cellular functions. Disruption of intracellular trafficking has been implicated in the pathogenesis of various neurodegenerative disorders. In addition, a number of genes involved in vesicle/organelle trafficking are also essential for pigmentation, and loss of those genes is often associated with mouse coat-color dilution and human hypopigmentary disorders. Hence, we postulated that screening for mouse mutants with both neurological defects and coat-color dilution will help identify additional factors associated with intracellular trafficking in neuronal cells. In this study, we characterized a mouse mutant with a unique N-ethyl-N-nitrosourea (ENU)–induced mutation, named nur17. nur17 mutant mice exhibit both coat-color dilution and ataxia due to Purkinje cell degeneration in the cerebellum. By positional cloning, we identified that the nur17 mouse carries a T-to-C missense mutation in archain 1 (Arcn1) gene which encodes the δ subunit of the coat protein I (COPI) complex required for intracellular trafficking. Consistent with this function, we found that intracellular trafficking is disrupted in nur17 melanocytes. Moreover, the nur17 mutation leads to common characteristics of neurodegenerative disorders such as abnormal protein accumulation, ER stress, and neurofibrillary tangles. Our study documents for the first time the physiological consequences of the impairment of the ARCN1 function in the whole animal and demonstrates a direct association between ARCN1 and neurodegeneration

Using C. elegans to discover therapeutic compounds for ageing-associated neurodegenerative diseases

Age-associated neurodegenerative disorders such as Alzheimer’s disease are a major public health challenge, due to the demographic increase in the proportion of older individuals in society. However, the relatively few currently approved drugs for these conditions provide only symptomatic relief. A major goal of neurodegeneration research is therefore to identify potential new therapeutic compounds that can slow or even reverse disease progression, either by impacting directly on the neurodegenerative process or by activating endogenous physiological neuroprotective mechanisms that decline with ageing. This requires model systems that can recapitulate key features of human neurodegenerative diseases that are also amenable to compound screening approaches. Mammalian models are very powerful, but are prohibitively expensive for high-throughput drug screens. Given the highly conserved neurological pathways between mammals and invertebrates, Caenorhabditis elegans has emerged as a powerful tool for neuroprotective compound screening. Here we describe how C. elegans has been used to model various human ageing-associated neurodegenerative diseases and provide an extensive list of compounds that have therapeutic activity in these worm models and so may have translational potential

Accelerated turnover of taste bud cells in mice deficient for the cyclin-dependent kinase inhibitor p27Kip1

Background: Mammalian taste buds contain several specialized cell types that coordinately respond to tastants and communicate with sensory nerves. While it has long been appreciated that these cells undergo continual turnover, little is known concerning how adequate numbers of cells are generated and maintained. The cyclin-dependent kinase inhibitor p27Kip1 has been shown to influence cell number in several developing tissues, by coordinating cell cycle exit during cell differentiation. Here, we investigated its involvement in the control of taste cell replacement by examining adult mice with targeted ablation of the p27Kip1 gene.Results: Histological and morphometric analyses of fungiform and circumvallate taste buds reveal no structural differences between wild-type and p27Kip1-null mice. However, when examined in functional assays, mutants show substantial proliferative changes. In BrdU incorporation experiments, more S-phase-labeled precursors appear within circumvallate taste buds at 1 day post-injection, the earliest time point examined. After 1 week, twice as many labeled intragemmal cells are present, but numbers return to wild-type levels by 2 weeks. Mutant taste buds also contain more TUNEL-labeled cells and 50% more apoptotic bodies than wild-type controls. In normal mice, p27 Kip1 is evident in a subset of receptor and presynaptic taste cells beginning about 3 days post-injection, correlating with the onset of taste cell maturation. Loss of gene function, however, does not alter the proportions of distinct immunohistochemically-identified cell types.Conclusions: p27Kip1 participates in taste cell replacement by regulating the number of precursor cells available for entry into taste buds. This is consistent with a role for the protein in timing cell cycle withdrawal in progenitor cells. The equivalence of mutant and wild-type taste buds with regard to cell number, cell types and general structure contrasts with the hyperplasia and tissue disruption seen in certain developing p27Kip1-null sensory organs, and may reflect a compensatory capability inherent in the regenerative taste system

Using C. elegans to decipher the cellular and molecular mechanisms underlying neurodevelopmental disorders

Prova tipográfica (uncorrected proof)Neurodevelopmental disorders such as epilepsy, intellectual disability (ID), and autism spectrum disorders (ASDs) occur in over 2 % of the population, as the result of genetic mutations, environmental factors, or combination of both. In the last years, use of large-scale genomic techniques allowed important advances in the identification of genes/loci associated with these disorders. Nevertheless, following association of novel genes with a given disease, interpretation of findings is often difficult due to lack of information on gene function and effect of a given mutation in the corresponding protein. This brings the need to validate genetic associations from a functional perspective in model systems in a relatively fast but effective manner. In this context, the small nematode, Caenorhabditis elegans, presents a good compromise between the simplicity of cell models and the complexity of rodent nervous systems. In this article, we review the features that make C. elegans a good model for the study of neurodevelopmental diseases. We discuss its nervous system architecture and function as well as the molecular basis of behaviors that seem important in the context of different neurodevelopmental disorders. We review methodologies used to assess memory, learning, and social behavior as well as susceptibility to seizures in this organism. We will also discuss technological progresses applied in C. elegans neurobiology research, such as use of microfluidics and optogenetic tools. Finally, we will present some interesting examples of the functional analysis of genes associated with human neurodevelopmental disorders and how we can move from genes to therapies using this simple model organism.The authors would like to acknowledge Fundação para a Ciência e Tecnologia (FCT) (PTDC/SAU-GMG/112577/2009). AJR and CB are recipients of FCT fellowships: SFRH/BPD/33611/2009 and SFRH/BPD/74452/2010, respectively

Cyclized NDGA modifies dynamic α-synuclein monomers preventing aggregation and toxicity.

Growing evidence implicates α-synuclein aggregation as a key driver of neurodegeneration in Parkinson's disease (PD) and other neurodegenerative disorders. Herein, the molecular and structural mechanisms of inhibiting α-synuclein aggregation by novel analogs of nordihydroguaiaretic acid (NDGA), a phenolic dibenzenediol lignan, were explored using an array of biochemical and biophysical methodologies. NDGA analogs induced modest, progressive compaction of monomeric α-synuclein, preventing aggregation into amyloid-like fibrils. This conformational remodeling preserved the dynamic adoption of α-helical conformations, which are essential for physiological membrane interactions. Oxidation-dependent NDGA cyclization was required for the interaction with monomeric α-synuclein. NDGA analog-pretreated α-synuclein did not aggregate even without NDGA-analogs in the aggregation mixture. Strikingly, NDGA-pretreated α-synuclein suppressed aggregation of naïve untreated aggregation-competent monomeric α-synuclein. Further, cyclized NDGA reduced α-synuclein-driven neurodegeneration in Caenorhabditis elegans. The cyclized NDGA analogs may serve as a platform for the development of small molecules that stabilize aggregation-resistant α-synuclein monomers without interfering with functional conformations yielding potential therapies for PD and related disorders

The University of Tokyo Atacama Observatory 6.5m telescope: Site development

One of remarkable features of the University of Tokyo Atacama Observatory (TAO) is an altitude of the site (5,640 m.a.s.l.) While this provides us an excellent condition for astronomical observations, the site development is attended with difficulty due to the hard conditions such as low pressure, low temperature, and limited access. Site preparation for the TAO 6.5 m telescope started in Apr. 2018. Firstly, we have constructed an access road from Pampa la bola plateau (∼ 5,000 m.a.s.l) to the summit. It has a width of < 6.5 meter for transportation of telescope parts including the 6.5meter mirror. In order to prevent collapse, angle of side slope is carefully determined based on ground condition and frozen soils. All workers always use oxygen during their work as a measure against hypobaropathy. Since the site temperature in night is lower than 0 degree even in the summer season, it is difficult to ensure quality of foundation concrete if we cast it in-situ. We use pre-cast concrete for the foundation of the telescope, the enclosure, and the support building. The biggest part is the telescope foundation. It has a weight of 600 ton. Considering the transportation to the summit, it is divided into 43 parts and unified at the summit. This is a new trial to make a massive foundation for a large telescope with pre-cast concrete. © COPYRIGHT SPIE. Downloading of the abstract is permitted for personal use only.Immediate accessThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

The University of Tokyo Atacama Observatory 6.5m telescope: Safety management at the extremely high altitude at Chajnantor mountain

Since the University of Tokyo Atacama Observatory (TAO) is located in extremely high altitude (5,640 m.a.s.l.), safety management is one of the most important issues for the project. We have developed safety management program for work at the site in order to prevent medical illness for humans. In this program all staff have to take medical examinations such as Electrocardiogram and Hipobaria before their work starts. The results are reviewed by medical doctor. Only authorized staff can be permitted to work at the site. During stay in site, all staff need to always use oxygen supply because there is only half of the oxygen at the site. It is also important to understand physical workload at the site. Our safety staff reviews it and determines necessary resting time for each worker, e.g. great load works need to work 50 minutes and rest 10 minutes. In addition to low air pressure, very low temperature, extremely high UV radiation, and extremely dry atmosphere should be concerned. Our program requests all staff to use winter clothes, sunblock and UV protection sunglasses and a lot of potable water consumption. Keep communication is also very important to secure the safety. Normal telephone communication is not available as well as internet connectivity. We have established satellite communication as well as UHF internal communication for general works and safety coordination with other observatories in Atacama area. © COPYRIGHT SPIE. Downloading of the abstract is permitted for personal use only.Immediate accessThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]