Biology subjects

Galvagnion, C.

Publications and source records attributed to Galvagnion, C..

9 recordsLinked to original sources

bioRxiv2026

Lipidic α-synuclein fibrils catalyze the formation of pathological mature fibrils

The aggregation of -synuclein into amyloid fibrils is a hallmark of a collection of neurodegenerative diseases termed synucleinopathies, including Parkinson's disease (PD). Interactions between -synuclein and lipid membranes modulate -synuclein aggregation and can produce protofibril structures consisting of both protein and lipid, termed lipidic fibrils. These lipidic fibrils can mature into mature fibrils, which resemble ex vivo fibrils extracted from Lewy bodies found in post-mortem brain samples of PD patients. Here, through the combination of experiments and kinetic theory, we study the influence of lipids and lipidic fibrils on the formation of mature fibrils. We find that lipidic fibrils catalyze mature fibril formation via a heterogeneous primary nucleation mechanism, which lowers the activation energy barrier of mature fibril formation relative to lipid-independent primary nucleation. The final stage of the reaction is governed by mature fibril elongation becoming limited by the rate at which protein monomers dissociate from the metastable lipidic fibrils, confirming that fibrils form in order of stability following Ostwald's Rule of Stages. This mechanism is validated using the small molecule inhibitor ambroxol, which selectively suppresses the formation of lipidic fibrils and in this manner shifts the aggregation kinetics to the slow, lipid-independent pathway. Taken together, these findings reveal that lipidic fibrils act as on-pathway intermediates and accelerate the aggregation of mature fibrils, which could provide a quantitative framework to design novel therapeutic strategies to inhibit pathological -synuclein aggregation.

biophysics↗

bioRxiv2025

Metabolic Flexibility of Microglia: Energy Substrate Utilization and Impact on Neuronal Metabolism

Microglia, the main resident immune cells of the brain, play critical roles in maintaining neuronal function and homeostasis. Microglia metabolic flexibility enables rapid adaptation to environmental changes, yet the full extent of their metabolic capabilities and influence on neuronal metabolism remains unclear. While microglia predominantly rely on glucose oxidative metabolism under homeostatic conditions, they shift toward glycolysis upon proinflammatory activation. In this study, we investigated microglial metabolism and its impact on neuronal metabolic homeostasis using isotope tracing with stable carbon 13C-enriched substrates and gas chromatography-mass spectrometry (GC-MS) analysis. Primary microglia were incubated with 13C-labeled glucose, glutamine, or GABA in the presence or absence of lipopolysaccharide (LPS) to assess metabolic adaptations upon an inflammatory challenge. Additionally, neurons co-cultured with quiescent or activated microglia (either with LPS or amyloid-{beta}) were incubated with 13C-enriched glucose to examine microglia-neuron metabolic interactions. Our findings confirm that microglia readily metabolize glucose and glutamine, with LPS stimulation slightly changing the glycolytic activity, as indicated by subtle changes in extracellular lactate. Importantly, we demonstrate for the first time that microglia take up and metabolize the inhibitory neurotransmitter GABA, suggesting a novel metabolic function. Furthermore, microglial presence directly influences neuronal metabolism and neurotransmitter homeostasis, highlighting a previously unrecognized aspect of neuron-microglia metabolic crosstalk. Collectively, these findings provide new insights into microglial metabolism and its role in neuronal function, with implications for neuroinflammatory and neurodegenerative diseases in which microglial metabolism is dysregulated.

neuroscience↗

bioRxiv2025

Ambroxol displaces α-synuclein from the membrane and inhibits the formation of early protein-lipid coaggregates

Parkinsons disease (PD) is a neurological disorder characterized by neuronal loss and the deposition of -synuclein-lipid coaggregates in the brain of patients as well as disruptions in lipid metabolism. Mutations in the gene GBA, which encodes the lysosomal glycoprotein Glucocerebrosidase, are together the most important genetic risk factor for PD and have been associated with lysosomal dysfunction, accumulation of pathological -synuclein as well as major changes in both the levels and properties of lipids. Ambroxol, a small molecule chaperone capable of binding and stabilizing Glucocerebrosidase, was found to revert changes in lipid levels and increase in -synuclein levels due to GBA mutations potentially via restoring lysosomal function. Here, we show that Ambroxol also has a direct effect on -synuclein-lipid co-aggregation by inhibiting the primary nucleation step in the aggregation process. We find that Ambroxol not only displaces -synuclein from negatively charged membranes but also prevents the formation of early -synuclein-lipid coaggregates during primary nucleation. These results suggest that Ambroxol may have beneficial effects on other synucleinopathies, such as multiple system atrophy and dementia with Lewy Bodies, that are also characterised by the aggregation of -synuclein into amyloid fibrils.

biophysics↗

bioRxiv2024

Divergent effects of pathological α-synuclein truncations and mutations on phase separation

Phase separated condensates can accelerate -synuclein (-Syn) amyloid fibril formation implicated in Parkinsons disease pathogenesis. The effects of pathological modifications, i.e., truncations and familial mutations on the thermodynamics, material properties, and the extent of amyloid aggregation within -Syn condensates remain elusive. Here, we quantitatively demonstrate that terminal truncations significantly alter -Syn phase separation, while familial mutations impart minimal effects. Spontaneous sol-gel phase transitions of the truncated -Syn variants could give rise to amyloid fibrils almost instantly within condensates, suggesting similarities between molecular interactions driving both processes. Extending our study to model coacervate and condensate systems where -Syn acts as a client, we find -Syn can dissolve coacervates and form Pickering clusters on condensate surfaces--regulating their size. Additionally, the C-terminal region of -Syn modulates nucleic acid sequestration within condensates. Together, our findings reveal diverse effects of -Syn modifications on phase separation, both in pathological and physiological contexts. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=151 SRC="FIGDIR/small/624073v1_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@1760f9dorg.highwire.dtl.DTLVardef@1fb155forg.highwire.dtl.DTLVardef@138402dorg.highwire.dtl.DTLVardef@1fd5229_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2024

Global kinetic model of lipid-induced alpha-synuclein aggregation and its inhibition by small molecules

The aggregation of -synuclein into amyloid fibrils is a hallmark of Parkinsons disease. This process has been shown to directly involve interactions between proteins and lipid surfaces when the latter are present. Despite this importance, the molecular mechanisms of lipid-induced amyloid aggregation have remained largely elusive. Here, we present a global kinetic model to describe lipid-induced amyloid aggregation of -synuclein. Using this framework we find that -synuclein fibrils form via a two-step primary nucleation mechanism and that lipid molecules are directly involved in both the nucleation and fibril elongation steps, giving rise to lipid-protein coaggregates. To illustrate the applicability of this kinetic approach to drug discovery, we identify the mechanism of action of squalamine, a known inhibitor of -synuclein aggregation, finding that this small molecule reduces the rate of lipid-dependent primary nucleation. Our work will likely guide the rational design of -synuclein aggregation inhibitors. Significance StatementAmyloid aggregation is a hallmark of a diverse range of diseases including Parkinsons Disease, where the protein -synuclein is a major con-stituent of proteinaceous deposits found in patients. It is well established that interactions between -synuclein and lipids modulate aggregation. However, the molecular mechanisms driving this lipid-induced aggregation have remained largely elusive, which has frustrated so far the discovery of drugs that prevent lipid-induced aggregation. In this work, we present the first global kinetic model describing lipid-induced aggregation and by integrating this theoretical framework with in vitro experimental data of lipid-induced -synuclein aggregation, we reveal the role of lipid membranes in the aggregation process and uncover the mechanism by which small molecule inhibitors interfere with this process.

biophysics↗

bioRxiv2024

Lipidomic Profiling Reveals Shared and Distinct Pathological Signatures in Sporadic Parkinson's Disease and GBA Mutation Carriers: Implications for Disease Mechanisms

Parkinsons Disease (PD) is a neurodegenerative disorder characterised by the deposition of protein-lipid inclusions, containing alpha-synuclein, neuronal cell loss and disruptions in lipid metabolism such as those associated with GBA mutations. GBA mutations are together an important genetic risk factor for PD and are associated with a decrease in glucocerebrosidase, a lysosomal glycoprotein encoded by GBA, increase in alpha-synuclein and changes in sphingolipids levels and composition. However, the extent of lipid metabolism disruptions associated to PD and their contributions to disease progression remain unclear. In this study, we used a combination of biochemical and lipidomic analyses of amygdala from healthy controls (HC) and people with sporadic (sPD) or GBA-associated PD (PD-GBA) to investigate the correlation between alpha-synuclein, glucocerebrosidase and lipids. We found extensive metabolic remodelling of brain lipids, including increased free cholesterol, diacylglycerides, sphingolipids and specific glycerophospholipids in amygdala from people with sPD and disease duration above 30 years (sPD>30y) and from people with PD carriers of a GBA risk mutation (PD-GBArisk) relative to HC. The levels of free cholesterol, diacylglycerides, sphingolipids and specific glycerophospholipids all correlated positively with pathological S and negatively with GCase activity. In contrast, the levels of phosphatidylethanolamine and cardiolipin only correlated positively with GCase activity. Moreover, we observed changes in the distribution of species for sphingolipids and glycerophospholipids in opposite directions for two categories of PD cases. We found a shift from short to long sphingomyelin and ceramide and from long to short phosphatidylserine and phosphatidylethanolamine in sPD>30y and PD-GBArisk cases and the opposite in sPD<10y and PD-GBAsevere cases. The relative proportion of lipid species affected in these samples all correlated with glucocerebrosidase activity and pathological alpha-synuclein levels. Together, these findings highlight the correlation between glucocerebrosidase, pathological alpha-synuclein and lipid levels in PD. Moreover, the identified opposite changes in lipid distribution for two categories of people with sPD and PD-GBA underscore the importance of patient stratification in clinical trials aiming at reverting PD-related lipid changes.

neuroscience↗

bioRxiv2023

Lipid levels correlate with neuronal and dopaminergic markers during the differentiation of SH-SY5Y cells

Parkinsons Disease (PD) is characterized by the loss of dopaminergic neurons and the deposition in the remaining cells of protein inclusions called Lewy Bodies (LBs). LBs are heterogeneous structures composed of protein and lipid molecules and their main constituent is the presynaptic protein -synuclein. SH-SY5Y cells are neuroblastoma cells commonly used to model PD because they express dopaminergic markers and -synuclein and they can be differentiated into neuronal cells using established protocols. Despite increasing evidence pointing towards a role of lipids in the initiation of PD, limited knowledge is available on the lipidome of undifferentiated and differentiated SH-SY5Y cells. In this study, we show that the levels and chemical properties of negatively charged phospholipids, diacyl glycerol and sphingolipids are specifically altered along the differentiation process of SH-SY5Y cells and that the levels of these lipids species correlate with those of dopaminergic and neuronal markers. These results are supported by proteomic data showing that the main biological processes affected by the differentiation of SH-SY5Y cells are lipid metabolism and processes associated with neuron maturation. Finally, our results show that electrophysiological activity can be detected in differentiated SH-SY5Y cells at a stage where most of the lipid changes have reached their maximal value. These results provide the first complete and quantitative characterisation of the changes in lipidome associated with the differentiation of SH-SY5Y cells into more neuronal and dopaminergic-like phenotype and serve as a basis for further characterisation of lipid disruptions in association with PD and its risk factors in this dopaminergic-like neuronal cell model.

cell biology↗

bioRxiv2020

Sphingolipid changes in Parkinson L444P GBA mutation fibroblasts promote α-synuclein aggregation

Intraneuronal accumulation of aggregated -synuclein is a pathological hallmark of Parkinsons disease. Therefore, mechanisms capable of promoting -synuclein deposition bear important pathogenetic implications. Mutations of the glucocerebrosidase 1 (GBA) gene represent a prevalent Parkinsons disease risk factor. They are associated with loss of activity of a key enzyme involved in lipid metabolism, glucocerebrosidase, supporting a mechanistic relationship between abnormal -synuclein-lipid interactions and the development of Parkinson pathology. In this study, the lipid membrane composition of fibroblasts isolated from control subjects, patients with idiopathic Parkinsons disease (iPD) and Parkinson patients carrying the L444P GBA mutation (PD-GBA) was assayed using shotgun lipidomics. The lipid profile of PD-GBA fibroblasts differed significantly from that of control and iPD cells. It was characterized by an overall increase in sphingolipid levels. It also featured a significant change in the proportion of ceramide, sphingomyelin and hexosylceramide molecules with shorter and longer hydrocarbon chain length; levels of shorter-chain molecules were increased while the percent of longer-chain sphingolipids was decreased in PD-GBA lipid extracts. The extent of this shift was correlated to the degree of reduction of fibroblast glucocerebrosidase activity. In a second set of experiments, lipid extracts from control and PD-GBA fibroblasts were added to incubations of recombinant -synuclein. The kinetics of -synuclein aggregation, as assessed by the binding of thioflavin T to amyloid structures, was significantly accelerated after addition of PD-GBA extracts as compared to control samples. Amyloid fibrils collected at the end of these incubations contained lipids, indicating -synuclein-lipid co-assembly. Lipids extracted from -synuclein fibrils were also analysed by shotgun lipidomics. Data revealed that the lipid content of these fibrils was significantly enriched of shorter-chain sphingolipids. Taken together, findings of this study indicate that the L444P GBA mutation and consequent enzymatic loss are associated with a distinctly altered membrane lipid profile that provides a biological fingerprint of this mutation in Parkinson fibroblasts. This altered lipid profile, which includes an increased content of shorter-chain sphingolipids, could also be an indicator of increased risk for -synuclein aggregate pathology. Shorter-chain molecules may act as preferred reactants during lipid-induced -synuclein fibrillation.

biochemistry↗

bioRxiv2019

An engineered monomer binding-protein for α-synucleinefficiently inhibits the proliferation of amyloid fibrils

Removing or preventing the formation of -synuclein aggregates is a plausible strategy against Parkinsons disease. To this end we have engineered the {beta}-wrapin AS69 to bind monomeric -synuclein with high affinity. In cultured cells, AS69 reduced the occurrence of -synuclein oligomers and of visible -synuclein aggregates. In flies, AS69 reduced -synuclein aggregates and the locomotor deficit resulting from -synuclein expression in neuronal cells. In a mouse model based on the intracerebral injection of pre-formed -synuclein seed fibrills (PFFs), AS69 co-injection reduced the density of dystrophic neurites observed three months later. In biophysical experiments in vitro, AS69 highly sub-stoichiometrically inhibited auto-catalytic secondary nucleation processes, even in the presence of a large excess of monomer. We present evidence that the AS69--synuclein complex, rather than the free AS69, is the inhibitory species responsible for sub-stoichiometric inhibition. These results represent a new paradigm that high affinity monomer binders can be strongly sub-stoichiometric inhibitors of nucleation processes.

bioengineering↗