Biology subjects

Chiki, A.

Publications and source records attributed to Chiki, A..

15 recordsLinked to original sources

bioRxiv2024

Dissecting the differential role of C-terminal truncations in the regulation of aSyn pathology formation and the biogenesis of Lewy bodies

Alpha-synuclein (aSyn) post-translational modifications (PTMs), particularly phosphorylation at serine 129 and C-terminal truncations, are highly enriched in Lewy bodies (LBs), Lewy neurites, and other types of aSyn pathological aggregates in the brain of patients with Parkinsons disease (PD) and other synucleinopathies. However, our knowledge about the precise role of PTMs in regulating the different stages of pathology formation, neurodegeneration, and aSyn pathology spreading remains incomplete. In this work, we applied a systematic approach to address this knowledge gap with an emphasis on mapping and elucidating the role of post-fibrillization C-terminal aSyn truncations in regulating the uptake, processing, seeding activity, and formation of LB-like inclusions and maturations in a well-established neuronal seeding model that recapitulates all the stages leading to LB formation and neurodegeneration. Our work shows that C-terminal cleavage of aSyn fibrils at multiple sites is a conserved process that occurs rapidly after and during the formation of intracellular LB-like aSyn inclusions in all neuronal seeding models. Interestingly, blocking the cleavage of internalized fibrils does not influence their seeding activity, whereas inhibiting the enzymes that regulate the cleavage of newly formed fibrils (e.g., calpains 1 and 2) significantly alters the formation of LB-like inclusions. We also show that C-terminal truncations, in combination with other PTMs, play a crucial role in regulating the interactome and remodeling of newly formed aSyn fibrils, including their shortening, lateral association, and packing during LB formation and maturation. Altogether, our results demonstrate that post-fibrillization C-terminal truncations have a differential role at different stages of aSyn aggregation and pathology formation. These insights, combined with the abundance of truncated aSyn species in LBs, have significant implications in understanding aSyn pathological diversity and developing therapeutic strategies targeting the C-terminus of aSyn or its proteolytic processing.

neuroscience↗

bioRxiv2023

Deep learning-assisted single-molecule detection of protein post-translational modifications with a biological nanopore

Protein post-translational modifications (PTMs) play a crucial role in countless biological processes, profoundly modulating protein properties on both the spatial and temporal scales. Protein PTMs have also emerged as reliable biomarkers for several diseases. However, only a handful of techniques are available to accurately measure their levels, capture their complexity at a single molecule level and characterize their multifaceted roles in health and disease. Nanopore sensing provides high sensitivity for the detection of low-abundance proteins, holding the potential to impact single-molecule proteomics and PTM detection in particular. Here, we demonstrate the ability of a biological nanopore, the pore-forming toxin aerolysin, to detect and distinguish -synuclein-derived peptides bearing single or multiple PTMs, namely phosphorylation, nitration and oxidation occurring at different positions and in various combinations. The characteristic current signatures of the -synuclein peptide and its PTM variants could be confidently identified using a deep learning model for signal processing. We further demonstrate that this framework can quantify -synuclein peptides at picomolar concentration and detect the C-terminal peptides generated by digestion of full-length -synuclein. Collectively, our work highlights the unique advantage of using nanopore as a tool for simultaneous detection of multiple PTMs and paves the way for their use in biomarker discovery and diagnostics.

biophysics↗

bioRxiv2023

Post-fibrillization nitration of alpha-synuclein abolishes its seeding activity and pathology formation in primary neurons and in vivo

Increasing evidence points to post-translational modifications (PTMs) as key regulators of alpha-synuclein (-Syn) function in health and disease. However, whether these PTMs occur before or after -Syn pathology formation and their role in regulating -Syn toxicity remain unclear. In this study, we demonstrate that post-fibrillization nitration of -Syn fibrils induced their fragmentation, modified their surface and dynamic properties but not their structure, and nearly abolished their seeding activity in primary neurons and in vivo. Furthermore, we show that the dynamic and surface properties of the fibrils, rather than simply their length, are important determinants of -Syn fibril seeding activity. Altogether, our work demonstrates that post-aggregation modifications of -Syn may provide novel approaches to target a central process that contributes to pathology formation and disease progression. Finally, our results suggest that the pattern of PTMs on pathological aggregates, rather than simply their presence, could be a key determinant of their toxicity and neurodegeneration. This calls for reconsidering current approaches relying solely on quantifying and correlating the level of pathology to assess the efficacy of novel therapies, as not all -Syn aggregates in the brain are pathogenic.

neuroscience↗

bioRxiv2022

Neighbouring modifications interfere with the detection of phosphorylated alpha-synuclein at Serine 129: Revisiting the specificity of pS129 antibodies

Alpha-synuclein (aSyn) within Lewy bodies, Lewy neurites, and other pathological hallmarks of Parkinsons disease and synucleinopathies have consistently been shown to accumulate in aggregated and phosphorylated forms of the protein, predominantly at Serine 129 (S129). Antibodies against phosphorylated S129 (pS129) have emerged as the primary tools to investigate, monitor, and quantify aSyn pathology in the brain and peripheral tissues. However, most of the antibodies and immunoassays aimed at detecting pS129-aSyn were developed based on the assumption that neighbouring post-translational modifications (PTMs) either do not co-occur with pS129 or do not influence its detection. Herein, we demonstrate that the co-occurrence of multiple pathology-associated C-terminal PTMs (e.g., phosphorylation at Tyrosine 125 or truncation at residue 133 or 135) differentially influences the detection of pS129-aSyn species by pS129-aSyn antibodies. These observations prompted us to systematically reassess the specificity of the most commonly used pS129 antibodies against monomeric and aggregated forms of pS129-aSyn in mouse brain slices, primary neurons, mammalian cells and seeding models of aSyn pathology formation. We identified two antibodies that are insensitive to pS129 neighbouring PTMs. However, consistent with previous reports, most pS129 antibodies showed cross-reactivity towards other proteins and often detected low and high molecular weight bands in aSyn knock-out samples that could be easily mistaken for monomeric or High Molecular Weight aggregates of aSyn. Our observations suggest that the pS129 antibodies do not capture the biochemical and morphological diversity of aSyn pathology. They also underscore the need for more specific pS129 antibodies, more thorough characterization and validation of existing antibodies, and the use of the appropriate protein standards and controls in future studies.

neuroscience↗

bioRxiv2021

The structural basis of huntingtin (Htt) fibril polymorphism, revealed by cryo-EM of exon 1 Htt fibrils

The lack of detailed insight into the structure of aggregates formed by the huntingtin protein has hampered efforts to develop therapeutics and diagnostics targeting pathology formation in the brain of patients with Huntingtons disease. To address this knowledge gap, we investigated the structural properties of in vitro generated fibrils from exon1 of the huntingtin protein by electron cryo-microscopy and single-particle analysis. We show that wildtype and mutant exon1 of the huntingtin protein form non-helical fibrils with a polygultamine amyloid core composed of {beta}-hairpins with unique characteristics that have not been previously observed with other amyloid filaments. The stacks of {beta}-hairpins form long planar {beta}- sheets (protofilaments) with variable stacking angle and occasional out-of-register state of individual {beta}-hairpins. These features and the propensity of protofilament to undergo lateral association results in a high degree of fibril polymorphism, including fibrils composed of varying numbers of protofilaments. Our results also represent the first direct observation of how the flanking domains are organized around the polyglutamine core of the fibril and provide insight into how they might affect huntingtin fibril structure, polymorphism, and stacking of {beta}-hairpins within its core structure. Removal of the first 17 amino acids at the N-terminus resulted in surprising intra-fibril structural heterogeneity and reduced fibrils propensity to lateral associations. Overall, this work provides valuable insights that could guide future mechanistic studies to elucidate the sequence and structural determinants of huntingtin aggregation, as well as cryo-EM and structural studies of fibrils derived from huntingtin proteins and other disease-associated polyglutamine-containing proteins.

neuroscience↗

bioRxiv2021

Investigating crosstalk among PTMs provides novel insight into the structural basis underlying the differential effects of Nt17 PTMs on mutant Httex1 aggregation

Post-translational modifications (PTMs) within the first 17 amino acids (Nt17) of the Huntingtin protein (Htt) have been shown to inhibit the aggregation and attenuate the toxicity of mutant Htt proteins in vitro and in various models of Huntingtons disease. Our groups previous studies suggested that the Nt17 PTM code is a combinatorial code that involves a complex interplay between different PTMs. Here, we expand on these studies by investigating the effect of methionine 8 oxidation (oxM8) and crosstalk between this PTM and either lysine 6 acetylation (AcK6) or threonine 3 phosphorylation (pT3) on the aggregation of mutant Httex1. We show that M8 oxidation delays but does not inhibit the aggregation and has no effect on the final morphologies of mutant Httex1 aggregates. This delay in aggregation kinetics could be attributed to the transient accumulation of oligomeric aggregates, which disappear upon the formation of Httex1 oxM8 fibrils. Interestingly, the presence of both oxM8 and AcK6 resulted in dramatic inhibition of Httex1 fibrillization, whereas the presence of oxM8 did not influence the aggregation inhibitory effect of pT3. To gain insight into the structural basis underlying these proteins aggregation properties, we investigated the impact of each PTM and the combination of these PTMs on the conformational properties of the Nt17 peptide by circular dichroism spectroscopy and molecular dynamics simulation. These studies show that M8 oxidation decreases the helicity of the Nt17 in the presence or absence of PTMs and provides novel insight into the structural basis underlying the effects of different PTMs on mutant Httex1 aggregation. PTMs that lower the mutant Httex1 aggregation rate (oxM8, AcK6/oxM8, pT3, pT3/oxM8, and phosphorylation at Serine 13) result in stabilization and increased population of a short N-terminal helix (first eight residues) in Nt17 or decreased abundance of other helical forms, including long helix and short C-terminal helix. PTMs that did not alter the aggregation of mutant Httex1 exhibit a similar distribution of helical conformation as the unmodified peptides. These results show that the relative abundance of N- vs. C-terminal helical conformations and long helices, rather than the overall helicity of Nt17, better explains the effect of different Nt17 PTMs on mutant Httex1; thus, explaining the lack of correlation between the effect of PTMs on the overall helicity of Nt17 and mutant Httex1 aggregation in vitro. Taken together, our results provide novel structural insight into the differential effects of single PTMs and crosstalk between different PTMs in regulating mutant Httex1 aggregation. TOC Figure O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=133 SRC="FIGDIR/small/432155v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@15dbf84org.highwire.dtl.DTLVardef@1c8a50corg.highwire.dtl.DTLVardef@1cbe0c6org.highwire.dtl.DTLVardef@120e0a_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2021

Towards deciphering the Nt17 code: How the sequence and conformation of the first 17 amino acids in Huntingtin regulate the aggregation, cellular properties, and neurotoxicity of mutant Httex1

Converging evidence points to the N-terminal domain comprising the first 17 amino acids of the Huntingtin protein (Nt17) as a key regulator of its aggregation, cellular properties and toxicity. In this study, we further investigated the interplay between Nt17 and the polyQ domain repeat length in regulating the aggregation and inclusion formation of exon 1 of the Huntingtin protein (Httex1). In addition, we investigated the effect of removing Nt17 or modulating its local structure on the membrane interactions, neuronal uptake, and toxicity of monomeric or fibrillar Httex1. Our results show that the polyQ and Nt17 domains synergistically modulate the aggregation propensity of Httex1 and that the Nt17 domain plays an important role in shaping the surface properties of mutant Httex1 fibrils and regulating their poly-Q-dependent growth, lateral association and neuronal uptake. Removal of Nt17 or disruption of its transient helical conformations slowed the aggregation of monomeric Httex1 in vitro, reduced inclusion formation in cells, enhanced the neuronal uptake and nuclear accumulation of monomeric Httex1 proteins, and was sufficient to prevent cell death induced by Httex1 72Q overexpression. Finally, we demonstrate that the uptake of Httex1 fibrils into primary neurons and the resulting toxicity are strongly influenced by mutations and phosphorylation events that influence the local helical propensity of Nt17. Altogether, our results demonstrate that the Nt17 domain serves as one of the key master regulators of Htt aggregation, internalization, and toxicity and represents an attractive target for inhibiting Htt aggregate formation, inclusion formation, and neuronal toxicity. HighlightsO_LIThe Nt17 and polyQ domains synergistically promote Httex1 aggregation. C_LIO_LIThe Nt17 domain is a key determinant of the lateral association and morphology of fibrils. C_LIO_LIThe Nt17 domain and conformation regulate the nuclear/cytoplasmic distribution and toxicity of Httex1. C_LIO_LINt17 conformation is a key determinant of Httex1 fibril membrane interaction and cellular uptake. C_LIO_LINt17 serves as one of the master regulators of Httex1 aggregation, cellular uptake and toxicity. C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=147 SRC="FIGDIR/small/431207v2_ufig1.gif" ALT="Figure 1"> View larger version (16K): org.highwire.dtl.DTLVardef@7afd68org.highwire.dtl.DTLVardef@92c67aorg.highwire.dtl.DTLVardef@24823dorg.highwire.dtl.DTLVardef@e2a835_HPS_FORMAT_FIGEXP M_FIG The Nt17 domain: A master switch of Httex1 aggregation, uptake, subcellular localization and neurotoxicity. In this paper, we showed that 1) the Nt17 and polyQ domains synergistically promote Httex1 aggregation; 2) the Nt17 domain is a key determinant of the lateral association and morphology of fibrils in vitro, 3) Nt17 conformation is a key determinant of Httex1 fibril membrane interaction and cellular uptake in primary neurons; 4) the Nt17 domain and conformation regulate the nuclear/cytoplasmic distribution and toxicity of Httex1 in primary neurons. The figure was created with Biorender and https://www.vectorstock.com/royalty-free-vector/icon-on-and-off-toggle-switch-button-white-design-vector-30148026 C_FIG

neuroscience↗

bioRxiv2020

Site-specific phosphorylation of Huntingtin exon 1 recombinant proteins enabled by the discovery of novel kinases

Posttranslational modifications (PTMs) within the first 17 amino acids (Nt17) of exon1 of the Huntingtin protein (Httex1) play important roles in modulating its cellular properties and functions in health and disease. In particular, phosphorylation of threonine and serine residues (T3, S13, and/or S16) has been shown to inhibit Htt aggregation in vitro and inclusion formation in cellular and animal models of Huntingtons disease (HD). In this manuscript, we describe a new and simple methodology for producing milligram quantities of highly pure wild type or mutant Httex1 proteins that are site-specifically phosphorylated at T3 or at both S13 and S16. This advance was enabled by 1) the discovery and validation of novel kinases that efficiently phosphorylate Httex1 at S13 and S16 (TBK1), at T3 (GCK) or T3 and S13 (TNIK and HGK); and, 2) the development of an efficient methodology for producing recombinant native Httex1 proteins using a SUMO-fusion expression and purification strategy. As proof of concept, we demonstrate how this method can be applied to produce Httex1 proteins that are both site- specifically phosphorylated and fluorescently labeled or isotopically labeled. Together, these advances should increase access to these valuable tools and expand the range of methods and experimental approaches that can be used to elucidate the mechanisms by which phosphorylation influences Httex1 structure, aggregation, interactome and function(s) in health and disease.

biochemistry↗

bioRxiv2020

Phosphorylation of the overlooked tyrosine 310 regulates the structure, aggregation, and microtubule- and lipid-binding properties of Tau

The microtubule-associated protein Tau is implicated in the pathogenesis of several neurodegenerative disorders, including Alzheimers disease. Increasing evidence suggests that post-translational modifications play critical roles in regulating Tau normal functions and its pathogenic properties in Tauopathies. Very little is known about how phosphorylation of tyrosine residues influences the structure, aggregation, and microtubule- and lipid-binding properties of Tau. In this work, we aimed to address this knowledge gap and determine the relative contribution of phosphorylation of one or several of the five tyrosine residues in Tau (Y18, Y29, Y197, Y310 and Y394) to the regulation of its biophysical, aggregation and functional properties. Towards this goal, we used a combination of site-specific mutagenesis and in vitro phosphorylation by c-Abl kinase to generate Tau species phosphorylated at all tyrosine residues, all tyrosine residues except Y310 or Y394 (pTau-Y310F, pTau-Y394F) and Tau phosphorylated only at Y310 or Y394 (4F\pY310 or 4F\pY394). Our results show that phosphorylation at all five tyrosine residues, multiple N-terminal tyrosine residues (Y18, Y29 and Y197) or site-specific phosphorylation at residue Y310, itself located in the microtubule-binding and aggregation-prone domain of Tau, was sufficient to abolish Tau aggregation and inhibit its microtubule- and lipid-binding properties. NMR studies demonstrated that these effects were mediated by a local decrease in {beta}-sheet propensity of the PHF6 domain. Our findings underscore the unique role of Y310 phosphorylation in the regulation of Tau aggregation, microtubule and lipid interactions and highlight the importance of conducting further studies to elucidate its role in the regulation of Tau normal functions and its pathogenic properties.

biochemistry↗

bioRxiv2019

TBK1 regulates autophagic clearance of soluble mutant huntingtin and inhibits aggregation/toxicity in different models of Huntington's disease

Phosphorylation of the N-terminal domain of the Huntingtin (HTT) protein (at T3, S13, and S16) has emerged as a key regulator of HTT stability, clearance, localization, aggregation and toxicity. Herein, we report the discovery and validation of a kinase, TANK-binding kinase 1 (TBK1), that specifically and efficiently phosphorylates both wild-type and mutant full-length or N-terminal fragments of HTT in vitro (S13/S16) and in cell/ neuronal cultures (S13). We show that overexpression of TBK1 in mammalian cells, primary neurons and a Caenorhabditis elegans model of Huntingtons Disease (HD) increases mutant HTTex1 phosphorylation, lowers its levels, increases its nuclear localization and significantly reduces its aggregation and cytotoxicity. Our mechanistic studies demonstrate that the TBK1-mediated neuroprotective effects are due to phosphorylation-dependent inhibition of mutant HTTex1 aggregation and an increase in autophagic flux. These findings suggest that upregulation and/or activation of TBK1 represents a viable strategy for the treatment of HD. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=148 SRC="FIGDIR/small/869586v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@1368abeorg.highwire.dtl.DTLVardef@1cecff4org.highwire.dtl.DTLVardef@b6b7c5org.highwire.dtl.DTLVardef@1daf53d_HPS_FORMAT_FIGEXP M_FIG Graphical abstract C_FIG

neuroscience↗

bioRxiv2019

A simple, versatile and robust centrifugation-based filtration protocol for the isolation and quantification of α-synuclein monomers, oligomers and fibrils: towards improving experimental reproducibility in α-synuclein research

Increasing evidence suggests that the process of alpha-synuclein (aSyn) aggregation from monomers into amyloid fibrils via oligomeric intermediates plays an essential role in the pathogenesis of different synucleinopathies, including Parkinsons disease (PD), multiple system atrophy and dementia with Lewy bodies. However, the nature of the toxic species and the mechanisms by which they contribute to neurotoxicity and disease progression remain elusive. Over the past two decades, significant efforts and resources have been invested in studies aimed at identifying the putative toxic species along the pathway of aSyn fibrillization, and to develop small molecule drugs or antibodies that target toxic aSyn oligomeric intermediates. Although this approach has helped to advance the field and provide insights into the biological properties and toxicity of different aSyn species, many of the fundamental questions regarding the role of aSyn aggregation in PD remain unanswered, and no therapeutic compounds targeting aSyn oligomers have passed clinical trials. Several factors have contributed to this slow progress, including the complexity of the aggregation pathways and the heterogeneity and dynamic nature of aSyn aggregates. In the majority of experiment, the aSyn samples used contain mixtures of aSyn species that exist in an equilibrium and their ratio changes upon modifying experimental conditions. The failure to quantitatively account for the distribution of different aSyn species in different studies has contributed not only to experimental irreproducibility but also to misinterpretation of results and misdirection of valuable resources. Towards addressing these challenges and improving experimental reproducibility in Parkinsons research, we describe here a simple centrifugation-based filtration protocol for the isolation, quantification and assessment of the distribution of of aSyn monomers, oligomers and fibrils, in heterogeneous aSyn samples of increasing complexity. The protocol is simple, does not require any special instrumentation and can be performed rapidly on multiple samples using small volumes. Here, we present and discuss several examples that illustrate the applications of this protocol and how it could contribute to improving the reproducibility of experiments aimed at elucidating the structural basis of aSyn aggregation, seeding activity, toxicity and pathology spreading. This protocol is applicable, with slight modifications, to other amyloid-forming proteins.\n\nTable of Content Figure\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=84 SRC=\"FIGDIR/small/772160v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (33K):\norg.highwire.dtl.DTLVardef@13fb942org.highwire.dtl.DTLVardef@9bc3adorg.highwire.dtl.DTLVardef@4b9c66org.highwire.dtl.DTLVardef@9054f_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2019

Extent of N-terminus exposure by altered long-range interactions of monomeric alpha-synuclein determines its aggregation propensity

As an intrinsically disordered protein, monomeric alpha synuclein (aSyn) constantly reconfigures and probes the conformational space. Long-range interactions across the protein maintain its solubility and mediate this dynamic flexibility, but also provide residual structure. Certain conformations lead to aggregation prone and non-aggregation prone intermediates, but identifying these within the dynamic ensemble of monomeric conformations is difficult. Herein, we used the biologically relevant calcium ion to investigate the conformation of monomeric aSyn in relation to its aggregation propensity. By using calcium to perturb the conformational ensemble, we observe differences in structure and intra-molecular dynamics between two aSyn C-terminal variants, D121A and pS129, and the aSyn familial disease mutants, A30P, E46K, H50Q, G51D, A53T and A53E, compared to wild-type (WT) aSyn. We observe that the more exposed the N-terminus and the beginning of the NAC region are, the more aggregation prone monomeric aSyn conformations become. N-terminus exposure occurs upon release of C-terminus interactions when calcium binds, but the level of exposure is specific to the aSyn mutation present. There was no correlation between single charge alterations, calcium affinity, or the number of ions bound on aSyns aggregation propensity, indicating that sequence or post-translation modification (PTM)-specific conformational differences between the N- and C-termini and the specific local environment mediate aggregation propensity instead. Understanding aggregation prone conformations of monomeric aSyn and the environmental conditions they form under will allow us to design new therapeutics targeted to the monomeric protein, to stabilise aSyn in non-aggregation prone conformations, by either preserving long-range interactions between the N- and C-termini or by protecting the N-terminus from exposure.

biophysics↗

bioRxiv2018

The making of a Lewy body: the role of alpha-synuclein post-fibrillization modifications in regulating the formation and the maturation of pathological inclusions.

Although converging evidence point to alpha-synuclein (a-syn) aggregation and Lewy body (LB) formation as central events in Parkinson's disease (PD), the molecular mechanisms that regulate these processes and their role in disease pathogenesis remain poorly understood. Herein, we applied an integrative biochemical, structural and imaging approach to elucidate the sequence, molecular and cellular mechanisms that regulate LB formation in primary neurons. Our results establish that post-fibrillization C-terminal truncation mediated by calpains 1 and 2 and potentially other enzymes, plays critical roles in regulating a-syn seeding, fibrillization and orchestrates many of the events associated with LB formation and maturation. These findings combined with the abundance of a-syn truncated species in LBs and pathological a-syn aggregates have significant implications for ongoing efforts to develop therapeutic strategies based on targeting the C-terminus of a-syn or proteolytic processing of this region.

neuroscience↗

bioRxiv2018

Chronic corticosterone enhancement aggravates alpha-synuclein brain spreading pathology and substantia nigra neurodegeneration in mice

Chronic stress and associated heightened glucocorticoid levels are risk factors for depression, a common non-motor symptom in Parkinsons disease (PD). However, how heightened glucocorticoids influence PD neuropathology [alpha-synuclein (-Syn) containing Lewy pathology and neurodegeneration] and disease progression is unclear. To address this knowledge gap, we investigated the impact of chronic corticosterone administration on -Syn pathology, neurodegeneration, behavior and mitochondrial function in a mouse model of -Syn pathology spreading after intracerebral injection of -Syn preformed fibrils (PFFs). Our results demonstrate that heightened corticosterone aggravates neurodegeneration and -Syn pathology spreading, intriguingly to specific brain regions, such as the entorhinal cortex. Corticosterone-treatment abolished distinct physiological adaptations after PFF-injection and induced differential physiological and behavioral consequences. Taken together, our work points to elevated glucocorticoids as a risk factor for the development of the neuropathological hallmarks of PD. Strategies aimed at reducing glucocorticoid levels might slow down pathology spreading and disease progression in synucleinopathy.

neuroscience↗

bioRxiv2018

N-terminal phosphorylation of Huntingtin: A molecular switch for regulating Htt aggregation, helical conformation, internalization and nuclear targeting.

Phosphorylation of exon1 of the Huntingtin protein (Httex1) has been shown to play important roles in regulating the structure, toxicity and cellular properties of N-terminal fragments and the full-length Huntingtin protein. Here, we investigated and compared the effect of bona fide phosphorylation at S13 and/or S16 on the structure, aggregation, membrane binding, and subcellular properties of mutant Httex1-Q18A. We show that serine phosphorylation at either S13 or S16 strongly disrupts the amphipathic -helix of the N-terminus, inhibits the aggregation of mutant Httex1 and prompts the internalization and nuclear targeting of Httex1 preformed aggregates. In synthetic peptides phosphorylation at S13 and/or S16 strongly disrupted the amphipathic -helix of the N-terminal 17 residues (Nt17) of Httex1 and Nt17 membrane binding. Our studies on peptides bearing a different combination of phosphorylation sites within Nt17 revealed a novel phosphorylation-dependent switch for regulating the structure of Httex1 involving crosstalk between phosphorylation at T3 and S13 or S16. Together, our results provide novel insights into the role of phosphorylation in regulating Httex1 structure and function in health and disease and underscore the critical importance of identifying enzymes responsible for regulating Htt phosphorylation and their potential as therapeutic targets for the treatment of Huntingtons disease.

neuroscience↗