Phosphorylation Modulates Clearance of Alpha-Synuclein Inclusions in a Yeast Model of Parkinson's Disease

Alpha-synuclein (aSyn) is the main component of proteinaceous inclusions known as Lewy bodies (LBs), the typical pathological hallmark of Parkinson''s disease (PD) and other synucleinopathies. Although aSyn is phosphorylated at low levels under physiological conditions, it is estimated that ∼90% of aSyn in LBs is phosphorylated at S129 (pS129). Nevertheless, the significance of pS129 in the biology of aSyn and in PD pathogenesis is still controversial. Here, we harnessed the power of budding yeast in order to assess the implications of phosphorylation on aSyn cytotoxicity, aggregation and sub-cellular distribution. We found that aSyn is phosphorylated on S129 by endogenous kinases. Interestingly, phosphorylation reduced aSyn toxicity and the percentage of cells with cytosolic inclusions, in comparison to cells expressing mutant forms of aSyn (S129A or S129G) that mimic the unphosphorylated form of aSyn. Using high-resolution 4D imaging and fluorescence recovery after photobleaching (FRAP) in live cells, we compared the dynamics of WT and S129A mutant aSyn. While WT aSyn inclusions were very homogeneous, inclusions formed by S129A aSyn were larger and showed FRAP heterogeneity. Upon blockade of aSyn expression, cells were able to clear the inclusions formed by WT aSyn. However, this process was much slower for the inclusions formed by S129A aSyn. Interestingly, whereas the accumulation of WT aSyn led to a marked induction of autophagy, cells expressing the S129A mutant failed to activate this protein quality control pathway. The finding that the phosphorylation state of aSyn on S129 can alter the ability of cells to clear aSyn inclusions provides important insight into the role that this posttranslational modification may have in the pathogenesis of PD and other synucleinopathies, opening novel avenues for investigating the molecular basis of these disorders and for the development of therapeutic strategies.     

How to build a paraspeckle

Noncoding RNAs have recently been identified as essential components of the nuclear suborganelles called paraspeckles. This finding will facilitate our understanding of the molecular dynamics and physiological role of these enigmatic macromolecular structures.     

Increased serum HSP70 levels are associated with the duration of diabetes

The evolutionary conserved family of heat shock proteins (HSP) is responsible for protecting cells against different types of stress, including oxidative stress. Although the levels of HSPs can be readily measured in blood serum, the levels of HSP70 in patients with different durations of diabetes have not been studied before. We quantified serum HSP70 levels in a healthy control group (n = 36) and two groups of type 2 diabetic patients, defined as newly diagnosed diabetes (n = 36) and patients with diabetes duration of more than 5 years (n = 37). The clinical characteristics and biochemical parameters were evaluated in the studied population. We found that serum HSP70 levels were significantly higher in patients with diabetes when compared with controls (p < 0.001) and it was higher in patients with disease for more than 5 years than in newly diagnosed patients (p < 0.001). Serum HSP70 was inversely correlated with fasting blood sugar in patients with diabetes for more than 5 years (r = −0.500, p = 0.002), positively correlated with the history of hypertension in newly diagnosed patients (p < 0.001), and positively correlated with age in patients with diabetes (r = 0.531, p = 0.001). Serum level of HSP70 is significantly higher in patients with diabetes and correlates with the duration of disease. Higher HSP70 in prolonged diabetes versus newly diagnosed diabetes may be an indicator of metabolic derangement in the course of diabetes.     

Harnessing the power of yeast to unravel the molecular basis of neurodegeneration

Several neurodegenerative diseases, such as Parkinson's disease (PD), Alzheimer's disease (AD), Huntington's disease (HD), amyotrophic lateral sclerosis (ALS), or prion diseases, are known for their intimate association with protein misfolding and aggregation. These disorders are characterized by the loss of specific neuronal populations in the brain and are highly associated with aging, suggesting a decline in proteostasis capacity may contribute to pathogenesis. Nevertheless, the precise molecular mechanisms that lead to the selective demise of neurons remain poorly understood. As a consequence, appropriate therapeutic approaches and effective treatments are largely lacking. The development of cellular and animal models that faithfully reproduce central aspects of neurodegeneration has been crucial for advancing our understanding of these diseases. Approaches involving the sequential use of different model systems, starting with simpler cellular models and ending with validation in more complex animal models, resulted in the discovery of promising therapeutic targets and small molecules with therapeutic potential. Within this framework, the simple and well‐characterized eukaryote Saccharomyces cerevisiae, also known as budding yeast, is being increasingly used to study the molecular basis of several neurodegenerative disorders. Yeast provides an unprecedented toolbox for the dissection of complex biological processes and pathways. Here, we summarize how yeast models are adding to our current understanding of several neurodegenerative disorders.     

PLK2 Modulates α-Synuclein Aggregation in Yeast and Mammalian Cells

Phosphorylation of α-synuclein (aSyn) on serine 129 is one of the major post-translation modifications found in Lewy bodies, the typical pathological hallmark of Parkinson’s disease. Here, we found that both PLK2 and PLK3 phosphorylate aSyn on serine 129 in yeast. However, only PLK2 increased aSyn cytotoxicity and the percentage of cells presenting cytoplasmic foci. Consistently, in mammalian cells, PLK2 induced aSyn phosphorylation on serine 129 and induced an increase in the size of the inclusions. Our study supports a role for PLK2 in the generation of aSyn inclusions by a mechanism that does not depend directly on serine 129 phosphorylation.     

Assessment of the Efficacy of Solutes from Extremophiles on Protein Aggregation in Cell Models of Huntington’s and Parkinson’s Diseases

Protein misfolding and deposition in the brain are implicated in the etiology of numerous neurodegenerative disorders. Here, organic solutes characteristic of microorganisms adapted to hot environments, were tested on experimental cell models of Huntington’s and Parkinson’s diseases. Diglycerol phosphate, di-myo-inositol phosphate, mannosylglycerate, and mannosylglyceramide were not toxic to the cells, at 10 mM concentration, but caused a decrease in cell density, which suggested an effect on proliferation. In contrast, mannosyl-lactate, an artificial analogue of mannosylglycerate, had a negative impact on cell viability. Concerning protein aggregation, inclusions of mutant huntingtin were reduced in the presence of diglycerol phosphate and di-myo-inositol phosphate, increased with mannosylglycerate, while mannosyl-lactate and mannosylglyceramide had no significant effect. α-Synuclein aggregation was not affected by the solutes tested, except for di-myo-inositol phosphate that led to a slight increased percentage of cells displaying visible aggregates. These solutes might be useful in the development of therapies for protein misfolding diseases.     

Dopamine-Induced Conformational Changes in Alpha-Synuclein

Background

Oligomerization and aggregation of α-synuclein molecules play a major role in neuronal dysfunction and loss in Parkinson''s disease [1]. However, α-synuclein oligomerization and aggregation have mostly been detected indirectly in cells using detergent extraction methods [2], [3], [4]. A number of in vitro studies showed that dopamine can modulate the aggregation of α-synuclein by inhibiting the formation of or by disaggregating amyloid fibrils [5], [6], [7].

Methodology/Principal Findings

Here, we show that α-synuclein adopts a variety of conformations in primary neuronal cultures using fluorescence lifetime imaging microscopy (FLIM). Importantly, we found that dopamine, but not dopamine agonists, induced conformational changes in α-synuclein which could be prevented by blocking dopamine transport into the cell. Dopamine also induced conformational changes in α-synuclein expressed in neuronal cell lines, and these changes were also associated with alterations in oligomeric/aggregated species.

Conclusion/Significance

Our results show, for the first time, a direct effect of dopamine on the conformation of α-synuclein in neurons, which may help explain the increased vulnerability of dopaminergic neurons in Parkinson''s disease.     

Conservation status of freshwater mussels in Europe: state of the art and future challenges

Freshwater mussels of the Order Unionida provide important ecosystem functions and services, yet many of their populations are in decline. We comprehensively review the status of the 16 currently recognized species in Europe, collating for the first time their life‐history traits, distribution, conservation status, habitat preferences, and main threats in order to suggest future management actions. In northern, central, and eastern Europe, a relatively homogeneous species composition is found in most basins. In southern Europe, despite the lower species richness, spatially restricted species make these basins a high conservation priority. Information on freshwater mussels in Europe is unevenly distributed with considerable differences in data quality and quantity among countries and species. To make conservation more effective in the future, we suggest greater international cooperation using standardized protocols and methods to monitor and manage European freshwater mussel diversity. Such an approach will not only help conserve this vulnerable group but also, through the protection of these important organisms, will offer wider benefits to freshwater ecosystems.