Biology subjects

Takatori, S.

Publications and source records attributed to Takatori, S..

8 recordsLinked to original sources

bioRxiv2026

Subtype-specific secretion of extracellular vesicles by LRRK2 and Rab GTPases under lysosomal stress

LRRK2, the Parkinsons disease-associated kinase, phosphorylates a subset of Rab GTPases and regulates membrane dynamics. We previously reported that lysosomal stress activates LRRK2 and thereby induces the exocytic secretion of lysosomal contents, but the detailed secretion mechanism remained unclear. Here we found that, under lysosomal stress, endolysosomal luminal and membrane components were secreted with extracellular vesicles (EVs) via LRRK2. Bis(monoacylglycerol)phosphate, an endolysosomal lipid and a urinary marker of LRRK2 activity, was similarly secreted via LRRK2, whereas CD9-positive EVs were not involved. Further dissection of the secreted EVs revealed that Alix-positive EVs were secreted via Rab8a as well as the ESCRT component VPS4, whereas LAMP1/cathepsin B-positive EVs were secreted via Rab10/Rab35, and SNARE proteins syntaxin 2 and VAMP8 regulated the secretion of both EV subtypes. These findings suggest a distinctive stress-induced secretory mechanism whereby LRRK2 facilitates the secretion of multiple EV subtypes by controlling Rab GTPases involved in each pathway.

cell biology↗

bioRxiv2026

Cryo-EM structure of pro-aggregant P301L/S320F double-mutant tau filaments formed in mouse brains following peripheral AAV delivery

Tauopathies are characterized by the accumulation of abnormally phosphorylated tau filaments, with disease-specific folds revealed by cryo-electron microscopy (cryo-EM). Here, we delivered a recombinant human tau carrying the P301L/S320F double mutation into AppNL-G-F/MAPT double knock-in mice using a blood-brain-barrier-permeable AAV (AAV-PHP.eB), enabling rapid induction of tau aggregation in vivo. The mutant tau was systemically administered via retro-orbital injection, providing a minimally invasive approach for widespread neuronal transduction. Phosphorylated tau aggregation, as well as seeding activity, were observed without the need for exogenous seeds. Our cryo-EM analysis resolved a novel filament fold incorporating residues 279-329, in which the P301L and S320F mutations introduced stabilizing interactions that reinforced filament assembly. This conformation was distinct from previously reported folds, including Alzheimer and Pick tau filaments. Moreover, the filaments adopted a more compact architecture than those observed in patient-derived samples or other model mice. This finding demonstrates that P301L/S320F double-mutant tau adopts a structurally unique, highly aggregation-prone fold, and that this system provides a rapid platform for modeling tau filament formation.

neuroscience↗

bioRxiv2026

Aging disrupts tissue homeostasis and constrains blastema-mediated regeneration in the Cladonema medusa

Regenerative capacity varies widely across animals, yet aging is often accompanied by declining tissue homeostasis and regenerative potential. In many species, regeneration of complex structures relies on epimorphic programs that form a blastema, coordinating cell proliferation and pattern reorganization after injury. Although aging-associated regeneration defects have been documented in several bilaterian models, how aging shapes regeneration in early-branching metazoans with robust regenerative abilities remains unclear. Here, we investigate aging and regeneration in the medusa stage of Cladonema pacificum, a cnidarian that retains high regenerative capacity within a finite lifespan. We show that aging in Cladonema medusae is accompanied by progressive morphological and functional deterioration, including shrinkage of the umbrella and manubrium, tentacle shortening, and reduced reproductive output, along with increased DNA damage. At the cellular level, aging is associated with depletion of differentiated cell populations, including nematocytes and neurons, as well as reductions in stem cell-associated populations and proliferative activity in the tentacle bulb. Consistent with these changes, tentacle regeneration is markedly impaired in aged medusae and is characterized by delayed wound closure, defective blastema formation, and incomplete functional recovery. Together, our findings indicate that aging disrupts both tissue homeostasis and blastema-mediated regeneration in Cladonema medusae, establishing a tractable model for studying aging-regeneration interactions and supporting the view that aging imposes broadly shared constraints on regenerative systems across metazoans.

developmental biology↗

bioRxiv2025

Inhibition of NF-κB signaling pathway in astrocytes facilitates amyloid-β clearance by kallikrein-related peptidase 7

Alzheimer disease (AD) is characterized by the deposition of amyloid-{beta} peptide (A{beta}). Decreased A{beta} clearance is observed in sporadic AD patients, suggesting that enhancing A{beta} clearance is a potential therapeutic approach for AD. We identified kallikrein-related peptidase 7 (KLK7) as an astrocyte-derived A{beta}-degrading protease, and its mRNA expression is reduced in AD brains. Memantine, an N-methyl-D-aspartate (NMDA) receptor antagonist, upregulates KLK7 expression in astrocytes; however, the regulatory mechanism remains unclear. Here, we show that the NMDA receptor signaling negatively regulates KLK7 mRNA expression via nuclear factor-{kappa}B (NF-{kappa}B). Inhibition of NF-{kappa}B signaling pathway in astrocytes increases KLK7 expression and promotes A{beta} degradation. Moreover, the mRNA expression level of the NF-{kappa}B family is elevated in AD brains and shows a negative correlation with KLK7 mRNA expression. Finally, the injection of an NF-{kappa}B inhibitor significantly upregulates Klk7 expression and reduces A{beta} levels in vivo. These findings suggest that the NMDA receptor-NF-{kappa}B signaling axis in astrocytes negatively regulates KLK7 expression and modulates KLK7-mediated A{beta} clearance.

neuroscience↗

bioRxiv2025

Aggregation of HAPLN2, a component of the perinodal extracellular matrix, is a hallmark of physiological brain aging in mice

Protein aggregation is a hallmark of neurodegenerative diseases and is also observed in the brains of elderly individuals without such conditions, suggesting that aging drives the accumulation of protein aggregates. However, the comprehensive understanding of age-dependent protein aggregates involved in brain aging remains unclear. Here, we investigated proteins that become sarkosyl-insoluble with age and identified hyaluronan and proteoglycan link protein 2 (HAPLN2), a hyaluronic acid-binding protein of the extracellular matrix at the nodes of Ranvier, as an age-dependent aggregating protein in mouse brains. Elevated hyaluronic acid levels and impaired microglial function reduced the clearance of HAPLN2, leading to its accumulation. HAPLN2 oligomers induced microglial inflammatory responses both in vitro and in vivo. Furthermore, age-associated HAPLN2 aggregation was also observed in the human cerebellum. These findings suggest that HAPLN2 aggregation results from age-related decline in brain homeostasis and may exacerbate the brain environment by activating microglia. This study provides new insights into the mechanisms underlying cerebellar aging and highlights the role of HAPLN2 in age-associated changes in the brain. Author SummaryTo identify age-dependent protein aggregates unrelated to disease, we analyzed the sarkosyl-insoluble proteome of young and aged mouse brains. We discovered that hyaluronan and proteoglycan link protein 2 (HAPLN2), a hyaluronic acid-binding protein existing in the nodal extracellular matrix, accumulated with age. Age-dependently accumulated HAPLN2 formed large protein aggregates that cannot be solubilized by the anionic detergent sarkosyl or by hyaluronidase digestion. In addition, HAPLN2 formed irregularly shaped puncta that were mislocalized from the nodes of Ranvier in the cerebellar white matter of not only aged mice but also aged human brains. Oligomers of full-length HAPLN2 specifically induced microglial activation in vitro and in vivo. Our findings suggest that the accumulation of HAPLN2 aggregates is a new hallmark of brain aging and a possible factor contributing to brain inflammation.

biochemistry↗

bioRxiv2024

Selective agonism of GPR34 stimulates microglial uptake and clearance of amyloid β fibrils

Microglia, the primary immune cells of the central nervous system, play a crucial role in maintaining brain homeostasis through phagocytosis of various substrates, including amyloid-{beta} (A{beta}) fibrils, a hallmark of Alzheimer disease (AD) pathology. However, the molecular mechanisms regulating microglial A{beta} uptake remain poorly understood. Here, we identified GPR34, a Gi/o-coupled receptor highly expressed in microglia, as a novel regulator of fibrillar A{beta} phagocytosis. Treatment with a selective GPR34 agonist, M1, specifically enhanced uptake of A{beta} fibrils, but not its monomer or oligomer, in both mouse and human microglia. Mechanistically, M1 reduced intracellular cAMP levels, which inversely correlated with A{beta} uptake activity. Importantly, a single intrahippocampal injection of M1 in an AD mouse model significantly increased microglial A{beta} uptake in vivo. Furthermore, single-nucleus RNA-sequencing analysis of Japanese AD patient samples revealed a significant reduction of GPR34 expression in microglia from AD patients compared to controls. We also observed an age-dependent decline in microglial GPR34 expression in both human and mouse datasets, suggesting a potential contribution of GPR34 downregulation to age-related A{beta} accumulation and AD risk. Collectively, our findings identify GPR34 as a promising target for modulating microglial A{beta} clearance and highlight the therapeutic potential of GPR34 agonists in AD. Significance statementAlzheimer disease (AD) is characterized by amyloid-{beta} (A{beta}) accumulation in the brain. Microglia, the brains immune cells, play a crucial role in the metabolism of A{beta}. We discovered that activating the microglial receptor GPR34 with a selective agonist enhances the phagocytosis of A{beta} fibrils, a key pathogenic form of A{beta}. Importantly, GPR34 expression decreases with aging and AD progression, potentially contributing to impaired A{beta} clearance. Our findings highlight GPR34 as a promising therapeutic target for AD, as boosting its activity could promote A{beta} clearance and slow disease progression. This study provides valuable insights into microglial function in AD and offers a novel strategy for developing disease-modifying therapies.

neuroscience↗

bioRxiv2021

BORCS6 is involved in the enlargement of lung lamellar bodies in Lrrk2 knockout mice

Leucine-rich repeat kinase 2 (LRRK2) has been implicated in the pathogenesis of Parkinson disease. It has been shown that Lrrk2 knockout (KO) rodents have enlarged lamellar bodies (LBs) in their alveolar epithelial type II cells, although the underlying mechanisms remain unclear. Here we performed proteomic analyses on LBs isolated from Lrrk2 KO mice and found that the LB proteome is substantially different in Lrrk2 KO mice compared with wild-type mice. In Lrrk2 KO LBs, several Rab proteins were increased, and subunit proteins of BLOC-1-related complex (BORC) were decreased. The amount of surfactant protein C was significantly decreased in the bronchoalveolar lavage fluid obtained from Lrrk2 KO mice, suggesting that LB exocytosis is impaired in Lrrk2 KO mice. We also found that the enlargement of LBs is recapitulated in A549 cells upon KO of LRRK2 or by treating cells with LRRK2 inhibitors. Using this model, we show that KO of BORCS6, a BORC subunit gene, but not other BORC genes, causes LB enlargement. Our findings implicate the LRRK2-BORCS6 pathway in the maintenance of LB morphology.

cell biology↗

bioRxiv2020

Molecular height measurement by cell surface optical profilometry (CSOP)

The physical dimensions of proteins and glycans on cell surfaces can critically affect cell function, for example by preventing close contact between cells and limiting receptor accessibility. However, high-resolution measurements of molecular heights on native cell membranes have been difficult to obtain. Here we present a simple and rapid method that achieves nanometer height resolution by localizing fluorophores at the tip and base of cell surface molecules and determining their separation by radially averaging across many molecules. We use this method, which we call cell surface optical profilometry (CSOP), to quantify height of key multi-domain proteins on a model macrophage and cancer cell, as well as to capture average protein and glycan heights on native cell membranes. We show that average height of a protein is significantly smaller than its contour length due to thermally driven bending and rotation on the membrane and that height strongly depends on local surface and solution conditions. We find that average height increases with cell surface molecular crowding, while it decreases with solution crowding by solutes, both of which we confirm with molecular dynamics simulations. We also use experiments and simulations to determine the height of an epitope based on the location of an antibody, which allows CSOP to profile various proteins and glycans on a native cell surface using antibodies and lectins. This versatile method for profiling cell surfaces has the potential to advance understanding of the molecular landscape of cells and its role in cell function.

biophysics↗