A proteomics analysis of 5xFAD mouse brain regions reveals the lysosome-associated protein Arl8b as a candidate biomarker for Alzheimer's disease

BACKGROUND: Alzheimer's disease (AD) is characterized by the intra- and extracellular accumulation of amyloid-β (Aβ) peptides. How Aβ aggregates perturb the proteome in brains of patients and AD transgenic mouse models, remains largely unclear. State-of-the-art mass spectrometry (MS) methods can comprehensively detect proteomic alterations, providing relevant insights unobtainable with transcriptomics investigations. Analyses of the relationship between progressive Aβ aggregation and protein abundance changes in brains of 5xFAD transgenic mice have not been reported previously. METHODS: We quantified progressive Aβ aggregation in hippocampus and cortex of 5xFAD mice and controls with immunohistochemistry and membrane filter assays. Protein changes in different mouse tissues were analyzed by MS-based proteomics using label-free quantification; resulting MS data were processed using an established pipeline. Results were contrasted with existing proteomic data sets from postmortem AD patient brains. Finally, abundance changes in the candidate marker Arl8b were validated in cerebrospinal fluid (CSF) from AD patients and controls using ELISAs. RESULTS: Experiments revealed faster accumulation of Aβ42 peptides in hippocampus than in cortex of 5xFAD mice, with more protein abundance changes in hippocampus, indicating that Aβ42 aggregate deposition is associated with brain region-specific proteome perturbations. Generating time-resolved data sets, we defined Aβ aggregate-correlated and anticorrelated proteome changes, a fraction of which was conserved in postmortem AD patient brain tissue, suggesting that proteome changes in 5xFAD mice mimic disease-relevant changes in human AD. We detected a positive correlation between Aβ42 aggregate deposition in the hippocampus of 5xFAD mice and the abundance of the lysosome-associated small GTPase Arl8b, which accumulated together with axonal lysosomal membranes in close proximity of extracellular Aβ plaques in 5xFAD brains. Abnormal aggregation of Arl8b was observed in human AD brain tissue. Arl8b protein levels were significantly increased in CSF of AD patients. CONCLUSIONS: We report a comprehensive biochemical and proteomic investigation of hippocampal and cortical brain tissue derived from 5xFAD transgenic mice, providing a valuable resource to the neuroscientific community. We identified Arl8b, with significant abundance changes in 5xFAD and AD patient brains. Arl8b might enable the measurement of progressive lysosome accumulation in AD patients and have clinical utility as a candidate biomarker

Mutant huntingtin impairs neurodevelopment in human brain organoids through CHCHD2-mediated neurometabolic failure

27 p.-7 fig.Expansion of the glutamine tract (poly-Q) in the protein huntingtin (HTT) causes the neurodegenerative disorder Huntington's disease (HD). Emerging evidence suggests that mutant HTT (mHTT) disrupts brain development. To gain mechanistic insights into the neurodevelopmental impact of human mHTT, we engineered male induced pluripotent stem cells to introduce a biallelic or monoallelic mutant 70Q expansion or to remove the poly-Q tract of HTT. The introduction of a 70Q mutation caused aberrant development of cerebral organoids with loss of neural progenitor organization. The early neurodevelopmental signature of mHTT highlighted the dysregulation of the protein coiled-coil-helix-coiled-coil-helix domain containing 2 (CHCHD2), a transcription factor involved in mitochondrial integrated stress response. CHCHD2 repression was associated with abnormal mitochondrial morpho-dynamics that was reverted upon overexpression of CHCHD2. Removing the poly-Q tract from HTT normalized CHCHD2 levels and corrected key mitochondrial defects. Hence, mHTT-mediated disruption of human neurodevelopment is paralleled by aberrant neurometabolic programming mediated by dysregulation of CHCHD2, which could then serve as an early interventional target for HD.We acknowledge support from the Deutsche Forschungsgemeinschaft (DFG)(PR1527/5-1 to A.P., RTG 2155 ProMoAge to H.O. and L.A.M.K., SFB167 B07 to J.P., RU2795: “Synapses under stress”: PR-1527/6-1 to A.P. and AN-1440/4-1 to R.A.), the Berlin Institute of Health (BIH) (to S.D., J.P., R.K., and A.P.),the Bundesministerium für Bildung und Forschung (BMBF) (AZ. 031L0211 and 01GM2002A to A.P. and 01EE2303B to J.P.), the Medical Faculty of Heinrich Heine University (FoKo grant to A.P. and S.C.), the European Commission’s Horizon Europe Program (SIMPATHIC #101080249 to A.P.),the National Science Centre, Poland (NCN grant No. 20 16/22/M/NZ2/00548 and 2017/27/B/NZ1/02401 to P.L.), the UK Dementia Research Institute programme grant (to J.P.), the Instituto de Salud Carlos III (ISCIII) grant PI20-00057 (to C.U.), the Berlin School of Integrative Oncology through the GSSP program of the German Academy of Exchange Service (DAAD) and the Joachim Herz Foundation through the Add-on Fellowship program (to T.M.P.), and the Studienstiftung des deutschen Volkes (to Se.Li.). We acknowledge the Center for Advanced Imaging (CAi) at Heinrich Heine University Düsseldorf for providing access to the Perki- nElmer Operetta CLS (DFG grant number INST 208/760-1 FUGG) and Olympus FV3000 microscope.Peer reviewe

Mutant huntingtin impairs neurodevelopment in human brain organoids through CHCHD2-mediated neurometabolic failure

Expansion of the glutamine tract (poly-Q) in the protein huntingtin (HTT) causes the neurodegenerative disorder Huntington’s disease (HD). Emerging evidence suggests that mutant HTT (mHTT) disrupts brain development. To gain mechanistic insights into the neurodevelopmental impact of human mHTT, we engineered male induced pluripotent stem cells to introduce a biallelic or monoallelic mutant 70Q expansion or to remove the poly-Q tract of HTT. The introduction of a 70Q mutation caused aberrant development of cerebral organoids with loss of neural progenitor organization. The early neurodevelopmental signature of mHTT highlighted the dysregulation of the protein coiled-coil-helix-coiled-coil-helix domain containing 2 (CHCHD2), a transcription factor involved in mitochondrial integrated stress response. CHCHD2 repression was associated with abnormal mitochondrial morpho-dynamics that was reverted upon overexpression of CHCHD2. Removing the poly-Q tract from HTT normalized CHCHD2 levels and corrected key mitochondrial defects. Hence, mHTT-mediated disruption of human neurodevelopment is paralleled by aberrant neurometabolic programming mediated by dysregulation of CHCHD2, which could then serve as an early interventional target for HD

The Anti-amyloid Compound DO1 Decreases Plaque Pathology and Neuroinflammation-Related Expression Changes in 5xFAD Transgenic Mice

Self-propagating amyloid-β (Aβ) aggregates or seeds possibly drive pathogenesis of Alzheimer's disease (AD). Small molecules targeting such structures might act therapeutically in vivo. Here, a fluorescence polarization assay was established that enables the detection of compound effects on both seeded and spontaneous Aβ42 aggregation. In a focused screen of anti-amyloid compounds, we identified Disperse Orange 1 (DO1) ([4-((4-nitrophenyl)diazenyl)-N-phenylaniline]), a small molecule that potently delays both seeded and non-seeded Aβ42 polymerization at substoichiometric concentrations. Mechanistic studies revealed that DO1 disrupts preformed fibrillar assemblies of synthetic Aβ42 peptides and decreases the seeding activity of Aβ aggregates from brain extracts of AD transgenic mice. DO1 also reduced the size and abundance of diffuse Aβ plaques and decreased neuroinflammation-related gene expression changes in brains of 5xFAD transgenic mice. Finally, improved nesting behavior was observed upon treatment with the compound. Together, our evidence supports targeting of self-propagating Aβ structures with small molecules as a valid therapeutic strategy

Additional file 1 of A proteomics analysis of 5xFAD mouse brain regions reveals the lysosome-associated protein Arl8b as a candidate biomarker for Alzheimer’s disease

Additional file 1: Table S1. Commercial antibodies used in this study. Table S2. Characteristics of AD and HD patients and corresponding controls. Table S3. Spearman correlation analysis of Arl8b protein level measurements and AD biomarker levels