Molecular Neurodegeneration

Progranulin (PGRN) deficiency is a common hallmark in frontotemporal dementia (FTD) patients with granulin (GRN) mutations (FTD-GRN). Previous studies by our group and others have observed that reduced PGRN perturbs lysosomal function and microglial activation, which is believed to accelerate neurodegeneration in FTD-GRN patients. Lysosomal function is intrinsically linked to lipid metabolism, and evidence suggests that GRN deficiency can alter lipid profiles in the brain. Unfortunately, studies to date have focused on whole brain or cortical extracts, limiting our ability to assess cell type-dependent changes in lipid metabolism under disease conditions. Here, we employed lipidomic analysis specifically within the pontine microglia of Grn knock-out (KO) mice, a cell population that was previously linked to disease phenotypes in this model. We observed a significant reduction in the endolysosomal lipid bis(monoacylglycero)phosphate (BMP) and the lipid metabolite phosphatidylethanolamine (PE); these microglial-specific lipid alterations mirror previous whole-brain findings, suggesting that similar changes may occur across multiple cell types in the brain. We also detected a significant increase in the myelin-composing factor galactosylceramide (GalCer), which may reflect an aberrant accumulation of myelin debris within microglia that arises due to defective lysosomal clearance. Notably, lipid perturbations were exacerbated with age within Grn KO microglia, suggesting that changes in lipid metabolism are both age- and genotype-dependent in this model. Together, our results support our hypothesis that PGRN acts as a master regulator of critical microglial processes - including lysosomal function, lipid metabolism, and the regulation of myelination - in an age-dependent manner.

BackgroundNeuromelanin (NM) is a pigment that progressively accumulates with age in catecholaminergic neurons, particularly in the substantia nigra, ventral tegmental area, and locus coeruleus. These neuronal populations are especially vulnerable to degeneration in Parkinsons disease (PD). Elevated intracellular NM levels have been linked to neurodegeneration and PD-like phenotypes in experimental models. However, the molecular mechanisms underlying NM-induced pathology remain poorly understood, as human studies cannot disentangle the specific effects of NM accumulation from those of normal aging. MethodsWe performed transcriptomic microarray analysis on laser-captured catecholaminergic neurons and regions (substantia nigra, ventral tegmental area, locus coeruleus) from NM-producing transgenic mice (tgNM) and NM-free wild-type controls across different ages, and compared them to data from postmortem human brain tissue. One of the molecular targets identified, GPNMB, was validated in mouse and human tissue, and functionally tested in vivo. ResultsWe identified region- and age-dependent transcriptional changes associated with progressive NM accumulation. NM consistently upregulated neuroinflammatory pathways with enrichment of disease-associated microglial genes, while downregulating transcription, translation, and mitochondrial functions. Locus coeruleus exhibited the earliest and strongest transcriptional alterations, whereas substantia nigra and ventral tegmental area showed a later-onset, age-progressive transcriptional dysfunction. Neuron-specific analyses revealed that many changes originated within NM-containing neurons rather than being solely glial-driven. NM-driven transcriptional profiles in mice strongly correlated with postmortem data from PD patients, underscoring their translational relevance. Among molecular targets, the glycoprotein GPNMB was consistently upregulated in NM-containing neurons and validated at RNA and protein levels in both NM-producing transgenic mice and human PD brains. Functional experiments demonstrated that GPNMB overexpression attenuated NM-linked dopaminergic neurodegeneration and improved motor performance in mice. ConclusionThis study provides a comprehensive in vivo characterization of NM-specific transcriptomic changes in catecholaminergic neurons, showing that NM accumulation drives neuroinflammatory and neurodegenerative programs. Our results support that the neuroinflammatory changes observed in tgNM mice and in human PD represent early pathological events that precede overt neurodegeneration. The disease-associated gene GPNMB emerged as a conserved NM-induced factor with protective properties, highlighting its potential as a therapeutic target in PD and aging-related neurodegeneration.

Microglia are central regulators of neuroinflammation in Alzheimers disease (AD), yet how metabolic states modulate function remains unclear. Here we show that microglia from the APPNL-G-F mouse model revealed upregulation of glycolytic enzymes coinciding with onset of microglial activation. Surprisingly, this glycolytic shift occurred alongside reduced expression of glucose transporters, suggesting that extracellular glucose may not be the primary fuel source, implicating glycogenolysis as the potential metabolic driver. Consistent with this, significant microglial glycogen accumulation was noted in late disease, when cells exhibited features of metabolic exhaustion and functional impairment. Pharmacological inhibition of glycogenolysis blunted microglia responses to Abeta aggregates and markedly reduced Abeta uptake, confirming a functional role for glycogen metabolism in shaping microglial states. Together, these findings identify glycogen as a central regulator of microglial metabolic health and function, highlighting glycogen homeostasis as a potential therapeutic target for promoting Abeta clearance and preserving protective microglial functions in AD.

Frontotemporal lobar degeneration (FTLD) and Alzheimers disease (AD) differ in their clinical features and genetic etiologies but share progressive cognitive decline. Emerging evidence implicates lipid dysregulation in neurodegeneration, but its extent across FTLD subtypes and how it compares to AD are unclear. Here, we performed integrated lipidomic and proteomic analyses of matched frontal (disease-vulnerable) and occipital (relatively spared) post-mortem cortices from individuals with genetic and sporadic FTLD-TDP, FTLD-tau (Picks disease, PiD), AD, and controls. FTLD and AD exhibited convergent lipid alterations, including reduced levels of cardiolipins and phosphatidylethanolamines, alongside increased gangliosides, diacylglycerols, cholesterol esters, acylcarnitines, and coenzyme Q, with generally greater changes in FTLD frontal cortex. FTLD displayed additional alterations, including reductions in bis(monoacylglycerol)phosphate, ceramides, phosphatidylserines, phosphatidylinositols, and sulfatides. These lipid changes were accompanied by proteomic alterations involving lysosomal proteins, phospholipases, phospholipid remodeling enzymes, and fatty acid oxidation pathways. Although lipidomic and proteomic signatures were broadly shared across FTLD subtypes, GRN associated FTLD-TDP and PiD showed the most extensive alterations. Triglycerides were selectively reduced in PiD in association with decreased DGAT1 expression, whereas cholesterol esters were elevated across all subtypes except C9orf72 associated FTLD-TDP. These findings identify shared disruptions in lipid homeostasis and lysosomal lipid metabolism across FTLD and AD, highlighting convergent metabolic pathways underlying neurodegeneration.

Tau pathology is a central driver of neurodegeneration, yet the molecular mechanisms linking tau accumulation to neuroinflammation, metabolic failure, and white matter degeneration remain incompletely understood. Galectin-3 (Gal3) is an inflammation-associated lectin expressed by activated microglia and has been implicated in neurodegenerative disease progression. Here, we investigated whether Gal3 modulates tau-driven pathology across cellular, molecular, and systems levels. Using the P301S tauopathy mouse model with genetic deletion of Gal3, we show that Gal3 loss robustly attenuates tau pathology across vulnerable brain regions, including cortex, hippocampus, and piriform-entorhinal cortex. Gal3 deletion reduced hyperphosphorylated and pathological tau species, normalized tau kinase signaling, and restored mitochondrial and vesicular trafficking pathways disrupted by tau accumulation. Proteomic and phosphoproteomic analyses revealed widespread normalization of tau-associated immune, metabolic, and trafficking pathways, with Tau-Gal3KO mice clustering closely with wild-type controls. In parallel, Gal3 deletion markedly reduced microglial activation and Gal3-positive inflammatory signatures, preserved white matter integrity, prevented axonal degeneration, and normalized oligodendrocyte and myelin abnormalities. Functionally, Gal3 deficiency enhanced microglial myelin phagocytosis and lysosomal degradation both in vitro and in vivo, suggesting improved clearance of myelin debris under inflammatory stress. Cell-type-specific analyses further revealed restoration of mitochondrial complex I subunit expression in both excitatory neurons and parvalbumin-positive interneurons. Importantly, translational studies in human iPSC-derived neurons demonstrated that extracellular Gal3 exacerbates tau hyperphosphorylation and aggregation following tau seeding, effects that were reversed by pharmacological Gal3 inhibition. Together, these findings identify Galectin-3 as a central upstream regulator linking tau pathology to neuroinflammation, proteomic dysregulation, mitochondrial dysfunction, and white matter degeneration. Targeting Gal3 represents a promising therapeutic strategy to mitigate tau-driven neurodegenerative processes.

BackgroundAggregated alpha-synuclein (syn) phosphorylated at serine 129 (PS129) accumulates in synucleinopathies, with the olfactory bulb (OB) being severely affected. Non-aggregated physiological PS129 is abundant in the mammalian OB, where it likely modulates syn-protein interactions. The impact of aggregation on the PS129 interactome in the OB remains unclear. We hypothesized that syn aggregation alters the PS129 interactome, shifting canonical synaptic partners (e.g., SNARE proteins) toward the aggregate-associated network. To test this hypothesis, we mapped PS129 interactions in the OB of PFF-injected WT and SNCAA53T/A53T (M83) mice using biotinylation by antibody recognition (BAR) and pretreated with calf-intestine alkaline phosphatase (CIAP) to distinguish physiological from aggregate-associated PS129 interactomes. Seeding and spread were assessed by immunohistochemistry and in situ seeding immunodetection assay (isSID). ResultsFollowing OB-PFF injections, CIAP-resistant aggregates and seeds were detected throughout the neuroaxis of M83 mice (e.g., OB to brainstem) but less so in WT mice. isSID seeding was concentrated near CIAP-resistant aggregates, but the overlap was only partial. BAR-PS129 identified 2,309 proteins in M83 OBs and 990 proteins in WT OBs. Of these, 357 proteins in M83 mice and 247 proteins in WT mice were associated with the CIAP-resistant, aggregate-enriched PS129 fraction. In both models, the CIAP-resistant interactome largely overlapped with the broader PS129 interactome, suggesting that seeded aggregation primarily affects existing PS129 interactions rather than directing the formation of new pathological ones. A conserved 107-protein CIAP-resistant signature shared between M83, and WT mice was enriched for axon-glia adhesion, myelin-associated, axonal/cytoskeletal, proteostatic, and synaptic vesicle-related proteins. ConclusionAggregated PS129 engages a subset of PS129 networks enriched at axon-glial interfaces. CIAP-resistant aggregates were partially associated with seed competency, indicating that CIAP resistance and seed competency are related but not equivalent. These results provide molecular details of syn seeding and spread from the OB.

BackgroundPhospholipase C gamma-2 (PLC{gamma}2) catalyzes the hydrolysis of the membrane phosphatidylinositol-4,5-bisphosphate (PIP2) to form diacylglycerol (DAG) and inositol trisphosphate (IP3), feeding into diverse downstream signaling pathways. PLCG2 polymorphisms have been associated with reduced and/or increased risk of Alzheimers disease (AD) and related dementias, longevity, autoinflammation, and immune disorders. In the brain, PLC{gamma}2 is expressed in microglia, and other neuroimmune and vascular interface populations, yet its role in brain homeostasis remains incompletely defined. MethodsWe analyzed the brains of three-month-old Plcg2 wild-type (WT), heterozygous (Het KO) and homozygous knockout (Homo KO) littermate mice modeling human PLCG2 loss-of-function risk alleles linked to AD risk using a multiomic approach that included lipidomics, metabolomics, proteomics, and transcriptomics, together with immunofluorescence, as well as flow-cytometric profiling of peripheral and brain-draining immune compartments. ResultsPlcg2 deficiency substantially impaired early survival and produced splenomegaly without increasing total spleen cellularity, instead shifting spleen composition toward myeloid/innate-enriched cells and away from B cells, with expansion of age-associated B-cell (ABC-like) subsets and parallel reductions in CD4 and CD8 regulatory T cells in spleen and cervical lymph nodes. Brain lipidomics revealed selective depletion of PIP2, despite very low bulk PLC{gamma}2 protein abundance relative to other PLC family members. PLC{gamma}2 loss led to significant reductions in myelin-enriched lipid classes and myelin/paranode-associated proteins, accompanied by compensatory upregulation of oligodendrocyte/myelin genes, and modest shifts in microglial, lysosomal, complement, and oxidative metabolism pathways by NanoString and DIA-MS. Targeted acylcarnitine profiling demonstrated reprogramming of brain oxidative metabolism, with increased short-, medium-, and long-chain acylcarnitines and enrichment of mitochondrial matrix fatty-acid and amino-acid catabolic enzymes in Homo KO brains. ConclusionsLoss of PLC{gamma}2 installs a coordinated program that compromises systemic immune tolerance and subtly erodes central myelin and phosphoinositide homeostasis while enhancing brain oxidative metabolism, effects that extend beyond microglial phagocytic signaling and may underlie increased vulnerability to AD pathology and aging, providing a mechanistic framework for how PLCG2 variation may link systemic immune regulation, white-matter integrity, and neurodegenerative risk. LimitationsBecause constitutive Plcg2 Homo KO mice display high early mortality and intestinal vascular abnormalities, observed phenotypes may reflect developmental compensation and may not fully recapitulate protective human PLCG2 variants.

INTRODUCTION: Parkinson's disease (PD) presents with motor and non-motor symptoms, including dementia, but the severity and rate of cognitive decline are heterogeneous and difficult to predict clinically. METHODS: We quantified baseline serum proteins with the high-throughput SomaScan(R) assay in 834 PD individuals and performed Cox regression to identify proteins associated with subsequent development of dementia. Candidate biomarker proteins were replicated in 371 individuals from an independent cohort and meta-analysed. RESULTS: Protein targets significantly associated with progression to dementia were predominantly involved in synaptic plasticity, protein degradation/lysosomal function and extracellular matrix organisation. Mendelian Randomisation further revealed that changes in the Nogo receptor RTN4R may be causally associated with the development of Lewy body dementia. DISCUSSION: We identified several proteins predicting progression to dementia in PD, indicating changes in blood proteome that precede the development of clinical symptoms by several years, providing a window of opportunity to identify at-risk individuals early on.

Immunoproteasome induction is prominent in Alzheimers disease (AD), but whether it protects proteostasis or amplifies neuroinflammation remains unresolved. Here, we generated immunoproteasome-deficient PS19 tauopathy and APP/human tau double-knock-in mice by crossing each disease model with L7M1 mice lacking two immunoproteasome catalytic subunits. Immunoproteasome deficiency increased phospho-tau burden, exacerbated amyloid-{beta} pathology and heightened microglial reactivity without suppressing constitutive 26S proteasome activity. In primary microglia and longitudinal two-photon imaging, immunoproteasome-deficient microglia engaged and engulfed tau aggregate-bearing material but failed to resolve internalized cargo, revealing a post-engulfment degradative checkpoint. Single-nucleus transcriptomics identified a remodeled P2ry12low/Trem2high microglial state with impaired phagolysosomal and mitochondrial programs. Reanalysis of human single-nucleus transcriptomic datasets showed that reduced microglial immunoproteasome expression was associated with cargo-processing gene-program changes similar to those observed in immunoproteasome-deficient mouse microglia. Together, these findings identify immunoproteasome biogenesis as a protective glial stress response that supports microglial aggregate clearance in AD.

Aging is a major risk factor for neurological disease, yet the molecular architecture of human brain aging remains poorly defined. Here, we analyzed more than 10,000 cerebrospinal fluid (CSF) proteomes across multiple cohorts and proteomic platforms to develop a 249-protein CSF aging clock that accurately predicted chronological age and generalized across independent datasets. CSF brain-age acceleration was increased across diverse neurological diseases, associated with blood-brain barrier (BBB) dysfunction, and predictive of longitudinal cognitive decline, neuroimaging progression and dementia conversion. A simplified 30-protein panel retained similar prognostic performance. Biologically, the clock resolved two opposing programs: pro-aging activation of immune, vascular/BBB, extracellular matrix and coagulation pathways, marked by CHI3L1, CD14, VWF, LRG1 and LTBP2, and collapse of anti-aging neuronal-maintenance programs, marked by NPTX2, COL1A2, NID1, CDH8 and PENK. Brain-wide single-cell and regional mapping linked these programs to disease-vulnerable compartments. These findings establish a CSF-based molecular framework for quantifying biological brain aging and predicting neurological disease progression.

Lysosomes make key contributions to the maintenance of cellular proteostasis, and their functional compromise has been linked to aging and neurodegenerative disease. A defining characteristic of lysosomes is their relative acidity compared to other subcellular compartments, a quality that enables the efficient breakdown of macromolecules. Evidence suggests that neuronal lysosomal pH becomes dysregulated with aging and neurodegenerative disease, yet the mechanisms by which lysosomal pH is maintained remain incompletely understood. To better understand neuronal lysosomal pH regulation, we conducted a genome-wide CRISPRi-based screen in iPSC-derived iNeurons for modifiers of lysosomal pH. We validated several previously known regulators of lysosomal pH and identified novel pathways capable of modifying lysosomal pH, including protein UFMylation and mitochondrial homeostasis. We demonstrate that loss of the lysosomal cationic amino acid exporter, PQLC2, prevents lysosomal acidification in a manner independent of amino acid transport. A novel, tauopathy-associated mutation in PQLC2 impairs lysosomal acidification and drives tau accumulation. Together, this study reveals novel genes that modify lysosomal pH and highlights potential new targets for ameliorating age-related lysosome dysfunction.

Amyotrophic lateral sclerosis (ALS) is a fatal, rapidly progressive neurodegenerative disease of motor neurons for which therapeutics are limited. Improved biomarkers are imperative to improve patient care and therapeutic development. Here, we employed 35-plex isobaric tandem mass tag labeling based on isobutyl-proline reporter group (TMTpro) to perform unbiased proteomic analysis of cerebrospinal fluid (CSF) and plasma from control (n= 28, n= 31) and sporadic ALS (sALS) (n= 39, n= 41), from the Target ALS Global Natural History Study (TALS GNHS). We identified 2,875 proteins in CSF and 1,118 proteins in plasma and identified known and novel differentially expressed proteins (DEPs) between controls and sALS, some of which were orthogonally validated using immunoassay. Comparison of TMTpro-MS and Olink proximity extension assay proteomics revealed common and non-overlapping differentially expressed proteins illustrating strengths unique to each platform. This initial cross-sectional proteomic study of biofluids from the TALS GNHS, with unrestricted availability of study results to the research community, highlights the potential of this resource as a potent platform for ALS biomarker discovery.

Tauopathies are characterized by the accumulation and spread of pathogenic tau aggregates throughout the brain, a process that is increasingly recognized to involve not only neurons but also microglia. However, whether pathogenic MAPT directly alters microglial degradative capacity remains poorly understood. Here, using isogenic human induced pluripotent stem cell-derived microglia carrying the pathogenic MAPT IVS10+16 mutation, we identify tau as a regulator of microglial lysosomal function. MAPT IVS10+16 microglia exhibited coordinated suppression of lysosomal and autophagic pathways, reduced lysosomal protease abundance and activity, and impaired autophagosome-lysosome fusion. Mutant microglia also showed reduced uptake of extracellular tau aggregates, reduced tau accumulation in acidic compartments, and a blunted lysosomal response to proteopathic stress. Conversely, genetic loss of MAPT increased lysosomal degradative capacity and accumulation of extracellular tau aggregates within acidic compartments, supporting a cell-intrinsic role for endogenous tau in regulating microglial degradative function. Pharmacologic enhancement of the autophagy lysosome pathway in MAPT IVS10+16 microglia increased proteolytic activity and improved tau handling. Together, these findings reveal a reciprocal relationship between tau and microglial lysosome function and identify degradative capacity as a modifiable component of the microglial response to tau pathology.

Tauopathies encompass diverse neurodegenerative diseases unified by aberrant patterns of tau deposition in brain. Although most appear sporadic, some are linked to genetic etiologies that offer unique mechanistic insights. Here we report that X-linked Dystonia-Parkinsonism (XDP), caused by a non-coding retrotransposon-associated repeat insertion in TAF1, involves a significant imbalance of tau isoforms and the accumulation of hyperphosphorylated, four-repeat tau in the brain. In striatal tissue, both misfolded tau accumulation, predominantly in astrocytes, and MAPT exon 10 inclusion correlated with repeat length within the causal insertion. Transcriptomic profiling across brain regions revealed dysregulation of known tau-related pathways. Levels of phosphorylated tau181, glial fibrillary acidic protein, and neurofilament light chain were elevated in patient plasma and discriminated XDP from controls. These findings implicate defective tau proteostasis as a key pathogenic mechanism and position XDP as a genetic model for uncovering cellular drivers that may disrupt tau in other more common neurodegenerative diseases.

BackgroundParkinsons disease (PD) is driven by -synuclein (-syn) aggregation and affects vulnerable dopaminergic and GABAergic neurons, and its incidence rises dramatically with age. In our -syn mouse model, motor impairment required both syn oligomers and the aging milieu, and pharmacological inhibition of the age hyperactivated Rho GTPase CDC42 with CASIN fully restored motor function, yet the underlying transcriptional pathways mediating this rescue remain to be elucidated. MethodsWe used an inducible -syn oligomer PD mouse model across three age groups (6, 16, and 24 months) with four conditions per group: -syn non-induced (OFF), induced (ON), and each with CASIN treatment (OFF-CASIN, ON-CASIN). Brain tissue from one hemisphere (0 to -5 mm Bregma) was sequenced using 10x Genomics 3 Chromium, with 3-4 mice per condition of both male and female mice. ResultssnRNA-seq demonstrated that CASIN robustly reverted PD-related transcriptional alterations at 24 months whereas aging-related changes were strongest at 16 months. Network and pathway analyses identified CASINs mode of action on two major downstream signaling cascades--MAPK and PI3K/AKT-- in the context of aging and MAPK signaling in PD. ConclusionConvergent gene-, transcription factor-, pathway-, and network-level evidence points to EGFR-PI3K-MAPK signaling as the axis through which CASIN may restore mitochondrial and synaptic function in PD and aging

Background: Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by death of upper and lower motor neurons, usually presented with clinical heterogeneity. Fluid biomarker development remains dominated by neurofilament light chain (NEFL), a marker of neuroaxonal injury. NEFL is however unspecific to ALS and its phenotypes and there is currently a lack of biomarkers that capture ALS heterogeneity such as onset site and ALS-frontotemporal spectrum disorder (ALS-FTSD). Therefore, we investigated whether plasma proteomics could reveal pathway-level signatures that stratify and explain ALS heterogeneity. Methods: We profiled ~5,400 plasma proteins (Olink Explore HT) in 299 patients with ALS and 50 age- and sex comparable healthy controls. We used two complementary analytic frameworks: (i) differential protein abundance analysis to identify altered proteins in ALS and across clinical subgroups, and (ii) weighted gene correlation network analysis (WGCNA) to identify coordinated protein modules and relate them to ALS diagnosis and to ALS-specific clinical traits (site of onset, ALS-FTSD, ALS functional rating scale-revised (ALSFRS-R) score, and plasma NEFL). Results: Differential abundance analysis identified 56 proteins altered in ALS versus controls, of which 40 were increased. WGCNA identified 11 co-expression modules, with ALS samples having the strongest correlation to a protein module (n=51) highly enriched for muscle-related proteins. Out of the 40 proteins that had increased expression levels, 29 overlapped with the muscle-enriched protein module, indicating that muscle related proteins are the dominant circulating proteomic signature in ALS. This signal extended to clinical stratification: spinal-onset patients showed a strong positive association with the muscle-module. Further, differential abundance analysis of spinal- versus bulbar-onset ALS identified changes that mapped predominantly to the same module, supporting a molecular signature of onset phenotype. In contrast, cognitive status (ALS-FTSD) mapped to distinct modules enriched for extracellular matrix/cell-adhesion pathways, consistent with a separable biological axis of disease heterogeneity. Although multiple modules correlated with NEFL, trait-specific signatures were not fully explained by neuroaxonal injury. Notably, the muscle-enriched module increased with higher NEFL and lower ALSFRS-R, supporting its interpretation as a severity-linked, muscle-involvement proxy. Conclusions: Large-scale plasma proteomics reveals that heterogeneity in ALS reflects underlying biological structures. We identified a dominant muscle-associated protein network that distinguished ALS patients from controls and correlated with disease onset phenotype and severity, alongside distinct protein networks linked to ALS-FTSD. By integrating differential protein abundance with network-based analysis, we defined pathway-level biomarker signatures that extend beyond NEFL, enabling biologically informed patient stratification and improved therapeutic monitoring.

INTRODUCTIONMicroglia regulate amyloid plaque-associated microenvironments that contribute to downstream tau pathology in Alzheimers disease (AD). Variants within the MS4A locus are strongly associated with AD risk and resilience and are linked to microglial biology; however, the functional role of MS4A4A in plaque-associated tau pathology remains poorly understood. METHODSSingle-nucleus RNA sequencing (snRNA-seq) was performed on hippocampi from non-transgenic, Ms4a4a knockout (4A-KO), 5xFAD, and 5xFAD 4A-KO mice at 6 months of age. To assess plaque-associated tau pathology, AD-derived tau aggregates were injected into the hippocampus of 5xFAD and 5xFAD 4A-KO mice at 6 months, and histological analyses were performed 3 months later. RESULTSAmyloid pathology was the dominant driver of microglial state transitions, while Ms4a4a loss selectively remodeled activated microglial transcriptional programs enriched for interferon, lysosomal, autophagic, and proteostatic pathways. Activated microglia from 5xFAD 4A-KO mice exhibited altered expression of genes linked to immune signaling and protein handling. Following AD-tau inoculation, Ms4a4a loss did not significantly alter overall phospho-tau burden but selectively reduced dense-core plaque-associated neuritic plaque tau (NP-tau), particularly in the contralateral hemisphere. This phenotype was strongest surrounding X-34-positive fibrillar plaques and occurred without major changes in plaque-associated microgliosis. DISCUSSIONThese findings identify Ms4a4a as a regulator of plaque-associated microglial programs linked to NP-tau accumulation in the amyloid-bearing brain. More broadly, this work supports a model in which AD resilience-associated microglial pathways selectively shape plaque-associated microenvironments that promote downstream tau pathology.

Post-translational modifications (PTMs) in APP and MAPT contribute to plaques and tangles in Alzheimers disease (AD). Yet broader proteome-wide PTMs in the AD brain are relatively unexplored. Therefore, this study highlights associations between PTMs, quantified by mass spectrometry in prefrontal cortex tissue, and Alzheimers disease neuropathology and cognition. Leveraging PTMs quantified from prefrontal cortices in 101 Rush Memory and Aging Project participants. We assessed associations with post-mortem amyloid-{beta} and tau burden, global cognition, and cognitive decline. First, APP and MAPT PTM associations were assessed on these outcomes given their known relevance in AD, followed by assessment of protein-wide effects of PTMs. Then, kinase enrichment analysis was performed on each outcome to assess which kinases might contribute to the results. We observed a novel association of APP-K687 acetylation, a known mutation hotspot driving pathology, with amyloid-{beta} load ({beta}=0.44, P=3.9e-8), while confirming known MAPT PTMs with tangle burden. Further, we identified 20+ novel PTMs associations with AD neuropathology, including ENO2-K256 ubiquitination ({beta}=0.353, P=1.13e-6), PSMD13-K31 ubiquitination ({beta}=0.568, P=1.34e-6), and PLXND1-K1826 ubiquitination ({beta}=0.577, P=7.08e-8) for tangle burden and SYP-K23 ubiquitination ({beta}=1.50, P=4.7e-8), TMEFF2-C80 cysteine oxidation ({beta}=1.64, P=1.1e-8), and STX1B-T121 phosphorylation ({beta}=0.898, P=3.3e-7) for amyloid-{beta} load. Further, kinase enrichment analyses highlight the complexity of disease-related proteome changes with some kinases like CDK5 showing expected over-enrichment (amyloid z=3.44, P=3.0e-4; tau z=4.98, P=3.3e-7) but others like PKC family kinases showing divergent enrichment between amyloid (z=8.98-11.55, P<1.0e-18) and tau (z=-2.83--3.88, P<0.006). This study provides an atlas of brain PTMs within crucial proteins like MAPT and APP and at the proteome-wide level, that impact AD neuropathology and clinical presentation. Further, we explored what kinases might be driving phosphorylation results, emphasizing the complex proteome changes which impact AD. In sum, these results highlight robust post-translational alterations in the AD brain and provide novel targets for future mechanistic studies.

In the pathologically and clinically heterogeneous neurodegenerative disorder amyotrophic lateral sclerosis (ALS), objective biochemical predictors of survival are essential to handle complexity in clinical trials, enrich clinical decision-making and interrogate the biology of disease progression. In this longitudinal study, we performed high-depth proximity extension assay proteomics using 1,095 samples of serum (N=851) and CSF (N=244) from 426 people with ALS, with orthogonal replication in an external cohort of 349 people with ALS. Age- and sex-adjusted Cox analysis identified 57 proteins in serum, including neurofilament light chain (NEFL) and peripherin, as well as five proteins in CSF, including tropomyosin 3 (TPM3) that were associated with survival (FDR-adjusted p[&ge;]0.05). Penalised Cox regression identified a panel of 9 serum proteins - including NEFL, peripherin, TNF receptor superfamily member 27 (EDA2R) and calcitonin - that reflect the extent of disease as well as the progression rate, improving survival prediction compared with models using clinical parameters and NEFL. Joint modelling identified associations between the longitudinal trajectories of serum EDA2R and calcitonin with survival, highlighting their potential role in measuring disease progression. This work indicates the utility of multiple proteins reflecting diverse biological pathways in refining survival stratification and highlights systemic factors in ALS progression.

Astrocytes are among the earliest cells to exhibit dysfunction in Alzheimers disease (AD), developing profound calcium signalling deficits before amyloid plaques have formed, yet the underlying mechanisms remain unknown. Lysosomal dysfunction is a hallmark of AD, but whether it initiates this early functional impairment or arises as a consequence of established pathology remains unresolved. Here, we find that astrocytic cytosolic calcium activity is suppressed prior to amyloid plaque deposition and is accompanied by reduced lysosomal acidification in vivo. Using a lysosome-targeted calcium indicator selectively expressed in astrocytes, we directly visualise lysosomal calcium dynamics in vivo and reveal a profound early loss of lysosomal calcium release, identifying lysosomal failure as an initiating event in astrocyte dysfunction in AD. Reduced expression of the lysosomal calcium channel TRPML1 provides the mechanistic basis for this deficit. Astrocyte-specific restoration of TRPML1 expression rescues lysosomal homeostasis and cytosolic calcium signalling and prevents astrocyte reactivity and morphological hypertrophy. Strikingly, early TRPML1 restoration prevents both the initial calcium hypoactivity observed before plaque formation and the later hyperactivity that characterises post-plaque disease, demonstrating that lysosomal calcium homeostasis stabilises astrocyte function across the disease trajectory. TRPML1 restoration also reduces amyloid plaque burden, indicating that astrocytic lysosomal competence directly shapes disease pathology. These findings identify lysosomal calcium failure as an early organelle-level mechanism linking amyloid stress to astrocyte dysfunction in AD, and position TRPML1-mediated lysosomal calcium signalling as a tractable target for limiting disease progression.