Biology subjects

Hatano, T.

Publications and source records attributed to Hatano, T..

10 recordsLinked to original sources

bioRxiv2025

Tegmental atrophy in isolated REM sleep behaviour disorder: Ex vivo-informed in vivo imaging

Isolated rapid eye movement (REM) sleep behaviour disorder (iRBD) is an early-stage synucleinopathy characterized by brainstem pathology. In rodents, the pontine tegmentum contains an REM sleep centre, the sublaterodorsal nucleus (SLD), which expresses corticotropin-releasing hormone binding protein (CRHBP). While the involvement of brainstem pathophysiology is thus implicated in iRBD, its solid evidence remains scarce in humans due to the difficulty in identifying small brainstem nuclei with conventional MRI technology alone. Here, we aimed to detect tegmental atrophy in iRBD with voxel-based morphometry (VBM) analysis combined with a novel human brainstem atlas. Structural MRIs from 98 patients with iRBD and 114 controls were analysed to investigate grey matter volume (GMV) using VBM. Our unique approach involved detailed assessments of the VBM results, guided by a high-resolution MRI-based atlas of the human brainstem. This brainstem atlas was founded on ex vivo MRI of 10 postmortem human specimens. We validated it with CRHBP immunostaining, which aided in identifying putative REM sleep-regulating nuclei in humans. We applied this brainstem atlas to identify atrophy in specific brainstem regions in iRBD and correlate their volumes with clinical measures, including autonomic functions. VBM revealed a focal cluster of grey matter atrophy in the dorsal pontine tegmentum of iRBD patients, including the laterodorsal tegmental nucleus, ventral part (LDTgV) and the pedunculopontine tegmental nucleus (PTg). Our atlas-based analysis confirmed the LDTgV as the site of most conspicuous atrophy, revealing a significant volume reduction in iRBD patients compared to controls with a moderate effect size (Cohens d = 0.46, Bonferroni-corrected p = 0.019). Furthermore, greater atrophy in the LDTgV and the PTg was associated with more severe autonomic dysfunction as measured by Scales for Outcomes in Parkinsons Disease-Autonomic dysfunction (SCOPA-AUT) scores (partial r = -0.237, p = 0.019 and partial r = -0.236, p = 0.019, respectively). Histological analysis confirmed that the LDTgV is selectively enriched with CRHBP-positive neurons, a putative marker for REM sleep-on neurons. We provided novel evidence for the involvement of LDTgV, the putative human homolog of the murine SLD, in iRBD. The present findings advance our understanding of the neuroanatomical basis of iRBD and will contribute to the development of early biomarkers for -synucleinopathies.

neuroscience↗

bioRxiv2025

Leveraging the salivary microbiome profile to stratify REM sleep behavior disorder and synucleinopathies

Few studies have explored noninvasive biomarkers of synucleinopathies, including Parkinsons disease (PD), and rapid-eye-movement behavior disorder (RBD), a prodromal stage for these conditions. Human oral/salivary microbiomes are altered in PD, highlighting their potential role in both PD pathogenesis and diagnosis. We analyzed 249 salivary microbiomes of controls and patients with idiopathic RBD and various synucleinopathies, including two subgroups divided based on the presence of RBD symptoms, through both cross-sectional and longitudinal studies. The microbiome composition was strikingly similar between patients with RBD and early PD exhibiting RBD symptoms. The area under the curve range for distinguishing RBD from controls and each synucleinopathy was 0.85-0.94. We further performed pseudotime trajectory analysis of the microbiome compositional space; populations with low diversity, enriched Streptococcus, and depleted Neisseria exhibited a brief pseudotime transition from RBD to early PD. Our findings suggest that early PD-like salivary dysbiosis manifests during RBD, allowing for the stratification of synucleinopathies through the innovative use of salivary microbiome profiles.

neuroscience↗

bioRxiv2024

Evolutionarily conserved principles of ESCRT-III-mediated membrane remodelling revealed by a two-subunit Asgard archaeal system

ESCRT-III proteins assemble into composite polymers that undergo stepwise changes in composition and structure to deform membranes across the tree of life. Here, using a phylogenetic analysis we demonstrate that the two ESCRT-III proteins present in our closest archaeal relatives are evolutionarily related to B-type and A-type eukaryotic paralogues, which initiate and execute membrane remodelling, respectively. This deep homology is reflected in ESCRT-III structure and function as demonstrated by the fact that ESCRT-IIIB assembles into parallel arrays on planar membranes to initiate membrane deformation, and is required to recruit ESCRT-IIIA to generate composite polymers. ESCRT-IIIA homopolymers can then remodel membranes into tubes, as a likely prelude to scission. Taken together, this analysis reveals a set of conserved principles governing ESCRT-III-dependent membrane remodelling that first emerged with the evolution of a two-component ESCRT-III system in the Asgard archaea, and which continue to underlie complex multi-component, ESCRT-III-dependent membrane remodelling in eukaryotes.

evolutionary biology↗

bioRxiv2024

Structure and physiological investigation of arginylated actin

Actin is an evolutionarily conserved cytoskeletal protein with crucial roles in cell polarity, division, migration, and muscle contraction. Actin function is regulated in part by posttranslational modifications. One such modification in non-muscle cells is arginylation, in which an arginine residue is added to the N-terminus of {beta}-actin. What is the structure of arginylated {beta}-actin (R-{beta}-actin), are its interactions with other proteins altered and what phenotypes result when R-{beta}-actin is the sole actin isoform present in the cell? Here we report the 4.2 [A] structure of ADP-bound human R-{beta}-actin filaments, the overall structure of which is nearly identical to the filaments made of non-arginylated actin. In vitro functional assays using isoform-pure actins with defined post-translational modifications reveal that the interaction between myosin-II and actin is altered upon actin arginylation, due to frequent detachment of myosin-II from R-actin filaments. In vivo, we find that replacement of the only actin gene in Schizosaccharomyces pombe with a synthetic gene encoding R-Sp-actin reduces Arp2/3-based actin patches while thickening the formin-induced actin. Furthermore, consistent with altered interactions between myosin-II and R-actin filaments, the assembly and constriction of cytokinetic actomyosin ring are perturbed in the R-Sp-actin cells. Thus, despite the overall structural similarity of arginylated and non-arginylated actin filaments, actin arginylation affects actin filament assortment into distinct subcellular structures and its interaction with myosin II.

biochemistry↗

bioRxiv2023

Connection between drug-mediated neurotransmission and salivary microbiome

Salivary microbiome alterations associated with cognitive function have been reported in patients with neurodegenerative diseases1-3. Gut microorganisms can modulate therapeutic efficacy via drug metabolism4-7. Additionally, several drugs against diabetes and inflammatory bowel disease can lead to microbial dysbiosis8-10. However, the effect of central nervous system (CNS) drug use on the microbiome remains unknown. Here, we show that the usage of anti-dementia drugs, including donepezil and memantine, more largely affects the salivary microbiome composition than the gut microbiome composition. We observed salivary microbiome diversity reduction in patients with neurodegenerative diseases who received N-methyl-D-aspartate receptor inhibitor drugs. Furthermore, the use of acetylcholine-modulating drugs contributed to the salivary microbiome composition, suggesting that the salivary microbiome responds to changes in CNS drug-induced cerebral acetylcholine and glutamate levels. Multivariate analysis adjusted with or without the use of anti-dementia drugs demonstrated that the difference in the salivary microbiome correlated with cognitive function. We show the unique salivary microbiome structure of CNS drug users, suggesting the possibility of monitoring pharmacokinetics using the salivary microbiome. Our results also provide evidence of the presence of the microbiome-oral-brain axis and will accelerate the elucidation of the interplay between the salivary microbiome and neurodegeneration.

microbiology↗

bioRxiv2022

Structural Mechanisms of Actin Isoforms

Actin isoforms organize into distinct networks that are essential for the normal function of eukaryotic cells. Despite a high level of sequence and structure conservation, subtle changes in their design principles determine the interaction with myosin motors and actin-binding proteins. The functional diversity is further increased by posttranslational modifications (PTMs). Therefore, identifying how the structure of actin isoforms relates to function is important for our understanding of normal cytoskeletal physiology. Here, we report the high-resolution structures of filamentous skeletal -actin (3.37[A]), cardiac -actin (3.07[A]), {beta}-actin (2.99[A]), and {gamma}-actin (3.38[A]) in the Mg2+{middle dot}ADP state with their native PTMs. The structures revealed isoform-specific conformations of the N-terminus that shifts closer to the filament surface upon myosin binding, thereby establishing isoform-specific interfaces. Retropropagated structural changes further show that myosin binding modulates actin filament structure. Further, our structures enabled us to reveal the location of disease-causing mutations and to analyze them with respect to known locations of PTMs. Collectively, the previously unknown structures of single-isotype, posttranslationally modified bare cardiac -actin, {beta}-actin, and {gamma}-actin reveal general principles, similarities, and differences between isoforms. They complement the repertoire of known actin structures and allow for a comprehensive understanding of in vitro and in vivo functions of actin isoforms.

biochemistry↗

bioRxiv2022

mNeonGreen-tagged fusion proteins and nanobodies reveal localization of tropomyosin to patches, cables, and contractile actomyosin rings in live yeast cells

Tropomyosins are structurally conserved -helical coiled-coil dimeric proteins that bind along the length of filamentous actin (F-actin) in fungi and animals. Tropomyosins play essential roles in the stability of actin filaments in non-muscle cells and are essential for calcium regulation of myosin II contractility in the muscle. Despite the crucial role of tropomyosin in actin cytoskeletal regulation, in vivo investigations of tropomyosin are limited, mainly due to the suboptimal live cell imaging tools currently available in many organisms. Here, we report mNeon-Green (mNG) tagged tropomyosin, with native promoter and linker length configuration, that clearly reports tropomyosin localization and dynamics in Schizosaccharomyces pombe (Cdc8), Schizosaccharomyces japonicus (Cdc8), and Saccharomyces cerevisiae (Tpm1 and Tpm2), in vivo and in isolated S. pombe cell division apparatuses. We extended this approach to also visualize the mammalian TPM2 isoform. Finally, we generated a camelid-nanobody against S. pombe Cdc8, which mimics the localization of mNG-Cdc8 in vivo without significantly influencing cell growth and dynamics of actin cytoskeleton. Using these tools, we report the presence of tropomyosin in previously unappreciated patch-like structures in fission and budding yeasts, show flow of tropomyosin (F-actin) cables to the cytokinetic actomyosin ring, and identify rearrangements of the actin cytoskeleton during mating. These powerful tools and strategies will aid better analyses of tropomyosin and actin cables in vivo.

cell biology↗

bioRxiv2021

Characterisation of the Ubiquitin-ESCRT pathway in Asgard archaea sheds new light on origins of membrane trafficking in eukaryotes

The ESCRT machinery performs a critical role in membrane remodelling events in all eukaryotic cells, including in membrane trafficking, membrane repair, cytokinetic abscission, in viral egress, and in the generation of extracellular vesicles. While the machinery is complex in modern day eukaryotes, where it comprises dozens of proteins, the system has simpler and more ancient origins. Indeed, homologues of ESCRT-III and the Vps4 ATPase, the proteins that execute the final membrane scission reaction, play analogous roles in cytokinesis and potentially in extracellular vesicle formation in TACK archaea where ESCRT-I and II homologues seem to be absent. Here, we explore the phylogeny, structure, and biochemistry of homologues of the ESCRT machinery and the associated ubiquitylation system found in genome assemblies of the recently discovered Asgard archaea. In these closest living prokaryotic relatives of eukaryotes, we provide evidence for the ESCRT-I and II sub-complexes being involved in the ubiquitin-directed recruitment of ESCRT-III,_as it is in eukaryotes. This analysis suggests a pre-eukaryotic origin for the Ub-coupled ESCRT system and a likely path of ESCRT evolution via a series of gene duplication and diversification events.

cell biology↗

bioRxiv2020

N-terminal modification of actin by acetylation and arginylation determines the architecture and assembly rate of linear and branched actin networks

The great diversity in actin network architectures and dynamics is exploited by cells to drive fundamental biological processes, including cell migration, endocytosis and cell division. While it is known that this versatility is the result of the many actin-remodeling activities of actin-binding proteins, recent work implicates post-translational modification of the actin N-terminus by either acetylation or arginylation itself as an equally important regulatory mechanism. However, the molecular mechanisms by which acetylation and arginylation alter the properties of actin are not well understood. Here, we directly compare how processing, and modification of the N-terminus of actin affects its intrinsic polymerization dynamics and its remodeling by actin-binding proteins that are essential for cell migration. We find that in comparison to acetylated actin, arginylated actin reduces intrinsic as well as formin-mediated elongation and Arp2/3-mediated nucleation. By contrast, there are no significant differences in Cofilin-mediated severing. Taken together, these results suggest that cells can employ the differently modified actins to precisely regulate actin dynamics. In addition, unprocessed, or non-acetylated actin show very different effects on formin-mediated-elongation, Arp2/3-mediated nucleation, and severing by Cofilin. Altogether, this study shows that the nature of the N-terminus of actin can induce distinct actin network dynamics, which can be differentially used by cells to locally finetune actin dynamics at distinct cellular locations, such as at the leading edge.

biochemistry↗

bioRxiv2019

Pick-ya actin: A method to purify actin isoforms with bespoke key post-translational modifications

Actin is one of the most abundant eukaryotic cytoskeletal polymer-forming proteins, which in the filamentous form regulates a number of physiological processes, ranging from cell division and migration to development and tissue function. Actins are differentially post-translationally modified (PTMs) in different organisms, which include Met, Ala, Asp, and Glu N-acetylation, N-arginylation, and the 73th His residue (His-73) methylation, with different organisms displaying a distinct signature of PTMs. Currently methods are not available to produce actin isoforms with organism specific PTM profile. Here we report Pick-ya actin, a method to express actin isoforms from any eukaryote with its own key characteristic PTM pattern. We achieve this using a synthetic biology strategy in a yeast strain that expresses 1. actin isoforms with the desired N-end via ubiquitin fusion and 2. mammalian enzymes that promote acetylation and methylation. Pick-ya actin should greatly facilitate biochemical, structural, and physiological studies of the actin cytoskeleton and its PTMs.

cell biology↗