Biology subjects

Watanabe, Y.

Publications and source records attributed to Watanabe, Y..

At least 55 records · Page 3Linked to original sources

bioRxiv2023

Collagen of ancient bones gives an indication of endogenous DNA preservation based on the next generation sequencing technology

Ancient genome analysis has become an indispensable tool in studies of human population history and evolution after the breakthrough of whole genome sequencing technology. Meanwhile, the problem has been not resolved that ancient genome cannot be analyzed without crushing non-small pieces of precious specimens; in spite of that, in many cases, there is no DNA remaining sufficiently in the piece of samples for obtaining the whole genome sequences. In previous studies, therefore, a couple of indicators (e.g., racemization ratios) have been proposed to estimate the endogenous DNA in ancient samples. However, these studies have used polymerase chain reaction (PCR) to test whether endogenous DNA is remaining, which has been inadequate because the success or failure of PCR amplification does not necessarily reflect DNA remaining. To assess the amount of endogenous DNA, we use the ratios of reads generated by the next-generation sequencer (NGS) mapped to the human reference genome sequence. We investigate 40 Jomon remains and find a significant association between the collagen residual ratios (CRRs) in rib bones and the mapping ratios (MRs). Because the weight of bone required to measure collagen residual is much less than that required to obtain DNA needed for NGS analysis, which is always necessary for dating, the collagen in ribs can be a good indicator for successful ancient genome analyses.

genomics↗

bioRxiv2023

Hippocampal neural fluctuation between memory encoding and retrieval states during a working memory task in humans

BackgroundWorking memory (WM) is essential for everyday life, yet its neural mechanism remains unclear. Although the hippocampus plays a critical role in memory consolidation and retrieval, its role in WM tasks has yet to be fully elucidated. We hypothesized that multiunit activities in the hippocampus alter their representations depending on the memory load and phase of a WM task, particularly in relation to sharp-wave ripple complexes (SWRs), which are known as various cognitive biomarkers. MethodsWe used an open dataset of intracranial electroencephalogram (iEEG) data and multiunit activity recorded from the medial temporal lobe (MTL) of nine patients with epilepsy. The MTL includes the hippocampus, entorhinal cortex, and amygdala. During the recording, all subjects performed an eight-second Sternberg test, in which they memorized sets of four, six, or eight letters (encoding phase), waited for three seconds (maintenance phase), and recalled whether a probe letter was included (Match IN task) or not (Mismatch OUT task) (retrieval phase). We used Gaussian-process factor analysis to visualize the neural trajectories of multiunit activity in MTL regions during the task. We also detected SWRs from the iEEG data in the hippocampus. FindingsWe found that the trajectory distance between phases of the Sternberg task was larger in the hippocampus compared to the entorhinal cortex and amygdala. Additionally, the trajectory distance between the encoding and retrieval phases was memory load dependent. Moreover, a transient trajectory increase was detected during SWRs. Finally, the trajectory direction of the hippocampus fluctuated between the encoding and retrieval states, and the balance of the fluctuation was shifted to the retrieval state during SWR periods. InterpretationOur results demonstrate the involvement of the hippocampus during a WM task. Furthermore, it is suggested that SWR in the retrieval phase plays a role in memory retrieval for a WM task. Our results provide new insight into the two-stage model of memory formation.

neuroscience↗

bioRxiv2023

Intestelligence: A pharmacological neural network using intestine data

A neural network is a machine learning algorithm that can learn and make predictions by adjusting the strength of the connections between nodes. The sigmoid function is commonly used as an activation function in these nodes. This study explores the potential applicability of biological materials in the development of alternative activation functions. Inspired by the fact that acetylcholine induces intestinal contractions that follow a sigmoid function, we used pharmacological data obtained from guinea pig ilea in a layered neural network for image classification tasks. We found that the intestinal data-based neural network with the same structure as a conventional three-layer perceptron achieved an impressive classification accuracy of 85.7% {+/-} 0.6% based on the MNIST handwritten digit dataset (chance = 10%). Additionally, the neural network was trained to determine whether objects in photographs collected from the internet were digestible, achieving an accuracy of 88.5% {+/-} 0.9% (chance = 50%). Our approach highlights the potential applicability of intestine data in neural computations based on pharmacological mechanisms.

bioengineering↗

bioRxiv2023

Potentiated cholinergic and corticofugal inputs support reorganized sensory processing in the basolateral amygdala during auditory threat acquisition and retrieval

Reappraising neutral stimuli as environmental threats reflects rapid and discriminative changes in sensory processing within the basolateral amygdala (BLA). To understand how BLA inputs are also reorganized during discriminative threat learning, we performed multi-regional measurements of acetylcholine (ACh) release, single unit spiking, and functional coupling in the mouse BLA and higher-order auditory cortex (HO-AC). During threat memory recall, sounds paired with shock (CS+) elicited relatively higher firing rates in BLA units and optogenetically targeted corticoamygdalar (CAmy) units, though not in neighboring HO-AC units. Functional coupling was potentiated for descending CAmy projections prior to and during CS+ threat memory recall but ascending amygdalocortical coupling was unchanged. During threat acquisition, sound-evoked ACh release was selectively enhanced for the CS+ in BLA but not HO-AC. These findings suggest that phasic cholinergic inputs facilitate discriminative plasticity in the BLA during threat acquisition that is subsequently reinforced through potentiated auditory corticofugal inputs during memory recall.

neuroscience↗

bioRxiv2023

Exploring the genetic diversity of the Japanese Population: Insights from a Large-Scale Whole Genome Sequencing Analysis

The Japanese archipelago is a terminal location for human migration, and the contemporary Japanese people represent a unique population whose genomic diversity has been shaped by multiple migrations from Eurasia. Through high-coverage whole-genome sequencing (WGS) analysis of 9,850 samples from the National Center Biobank Network, we analyzed the genomic characteristics that define the genetic makeup of the modern Japanese population from a population genetics perspective. The dataset comprised populations from the Ryukyu Islands and other parts of the Japanese archipelago (Hondo). Low frequency detrimental or pathogenic variants were found in these populations. The Hondo population underwent two episodes of population decline during the Jomon period, corresponding to the Late Neolithic, and the Edo period, corresponding to the Early Modern era, while the Ryukyu population experienced a population decline during the shell midden period of the Late Neolithic in this region. Genes related to alcohol and lipid metabolism were affected by positive natural selection. Two genes related to alcohol metabolism were found to be 12,500 years out of phase with the time when they began to be affected by positive natural selection; this finding indicates that the genomic diversity of Japanese people has been shaped by events closely related to agriculture and food production. Author summaryThe human population in the Japanese archipelago exhibits significant genetic diversity, with the Ryukyu Islands and other parts of the archipelago (Hondo) having undergone distinct evolutionary paths that have contributed to the genetic divergence of the populations in each region. In this study, whole genome sequencing of healthy individuals from national research hospital biobanks was utilized to investigate the genetic diversity of the Japanese population. Haplotypes were inferred from the genomic data, and a thorough population genetic analysis was conducted. The results indicated not only genetic differentiation between Hondo and the Ryukyu Islands, but also marked differences in past population size. In addition, gene genealogies were inferred from the haplotypes, and the patterns were scrutinized for evidence of natural selection. This analysis revealed unique traces of natural selection in East Asian populations, many of which were believed to be linked to dietary changes brought about by agriculture and food production.

genomics↗

bioRxiv2023

Physicochemical properties of the vacuolar membrane and cellular factors determine formation of vacuolar invaginations

Vacuoles change their morphology in response to stress. In yeast exposed to chronically high temperatures, vacuolar membranes get deformed and invaginations are formed. We show that phase-separation of vacuolar membrane occurred after heat stress leading to the formation of the invagination. In addition, Hfl1, a vacuolar membrane-localized Atg8-binding protein, was found to suppress the excess vacuolar invaginations after heat stress. At that time, Hfl1 formed foci at the neck of the invaginations in wild-type cells, whereas it was efficiently degraded in the vacuole in the atg8{Delta} mutant. Genetic analysis showed that the endosomal sorting complex required for transport (ESCRT) machinery was necessary to form the invaginations irrespective of Atg8 or Hfl1. In contrast, a combined mutation with the vacuole BAR domain protein Ivy1 led to vacuoles in hfl1{Delta}ivy1{Delta} and atg8{Delta}ivy1{Delta} mutants having constitutively invaginated structures; moreover, these mutants showed stress-sensitive phenotypes. Our findings suggest that vacuolar invaginations result from the combination of changes in the physiochemical properties of the vacuolar membrane and other cellular factors. Summary statementVacuolar invaginations occur through a combination of the changes of physiochemical properties of the vacuolar membrane and other cellular factors in yeast. Particularly, Hfl1 suppresses the excess invaginations.

cell biology↗

bioRxiv2022

Neural signatures of auditory hypersensitivity following acoustic trauma

Throughout the brain, neurons exhibit a remarkable capacity to maintain stable firing rates despite large perturbations in afferent activity levels. As an exception, homeostatic regulation of neural activity often fails in the adult auditory system after hearing loss. Cochlear deafferentation caused by aging or noise exposure triggers widespread neural hyperactivity, particularly in the auditory cortex (ACtx), which underlies perceptual disorders including tinnitus and hyperacusis. Here, we show that mice with noise-induced damage of the high-frequency cochlear base were behaviorally hypersensitive to spared mid-frequency tones and to direct optogenetic stimulation of auditory thalamocortical neurons. Chronic 2-photon calcium imaging from ACtx pyramidal neurons (PyrNs) revealed an initial stage of diffuse hyperactivity, hypercorrelation, and hyperresponsivity that consolidated around deafferented map regions three or more days after acoustic trauma. Deafferented PyrN ensembles displayed hypersensitive decoding of spared mid-frequency tones, mirroring behavioral hypersensitivity. At the level of individual PyrNs, some exhibited stable, homeostatic gain control after acoustic trauma, while others showed non-homeostatic excess gain. Interestingly, factors such as baseline spontaneous activity levels and sound level encoding could account for 40% of the variability in PyrN gain regulation after acoustic trauma. These findings suggest that non-homeostatic regulation of cortical sound intensity coding following sensorineural loss may underlie the well-established clinical phenomenon of loudness hypersensitivity. Further, while cortical gain changes are triggered by reduced bottom-up afferent input, their subsequent stabilization is also shaped by their local circuit milieu, where baseline response features can identify neurons with the greatest propensity for developing pathological hyperactivity following sensorineural hearing loss.

neuroscience↗

bioRxiv2022

Lassa virus glycoprotein nanoparticles elicit a neutralizing antibody that defines a new site of vulnerability

Lassa virus is endemic in large parts of West Africa and causes a hemorrhagic fever. Recent years have seen several serious outbreaks of Lassa fever with high mortality rates. A vaccine to curtail infection is urgently needed. The development of a recombinant protein vaccine has been hampered by the instability of soluble Lassa virus glycoprotein complex (GPC) trimers, which disassemble into monomeric subunits after expression. Here we use two-component protein nanoparticles to stabilize GPC in a trimeric conformation and present twenty prefusion GPC trimers on the surface of an icosahedral nanoparticle. Cryo-EM studies of assembled GPC nanoparticles demonstrated a well-ordered structure and yielded a high-resolution structure of an unliganded GPC. These nanoparticles induced potent humoral immune responses in rabbits and protective immunity against a lethal Lassa virus challenge in guinea pigs. We isolated a neutralizing antibody which was mapped to the putative receptor-binding site, revealing a novel site of vulnerability on GPC.

immunology↗

bioRxiv2022

The interferon stimulated gene-encoded protein HELZ2 inhibits human LINE-1 retrotransposition and LINE-1 RNA-mediated type I interferon induction

Some interferon stimulated genes (ISGs) encode proteins that inhibit LINE-1 (L1) retrotransposition. Here, we used immunoprecipitation followed by liquid chromatography-tandem mass spectrometry to identify proteins that associate with the L1 ORF1-encoded protein (ORF1p) in ribonucleoprotein particles. Three ISG proteins that interact with ORF1p inhibit retrotransposition: HECT and RLD domain containing E3 ubiquitin-protein ligase 5 (HERC5); 2-5-oligoadenylate synthetase-like (OASL); and helicase with zinc finger 2 (HELZ2). HERC5 destabilizes ORF1p, but does not affect its cellular localization. OASL impairs ORF1p cytoplasmic foci formation. HELZ2 recognizes sequences and/or structures within the L1 5UTR to reduce L1 RNA, ORF1p, and ORF1p cytoplasmic foci levels. Overexpression of WT or reverse transcriptase-deficient L1s led to a modest induction of IFN- expression, which was abrogated upon HELZ2 overexpression. Notably, IFN- expression was enhanced upon overexpression of an ORF1p RNA binding mutant, suggesting ORF1p binding might protect L1 RNA from "triggering" IFN- induction. Thus, ISG proteins can inhibit retrotransposition by different mechanisms.

molecular biology↗

bioRxiv2021

Revisiting PFA-mediated tissue fixation chemistry: FixEL enables trapping of small molecules in the brain to visualize their distribution dynamics

Various small molecules have been used as functional probes for tissue imaging in medical diagnosis and pharmaceutical drugs for disease treatment. The spatial distribution, target selectivity, and diffusion/extrusion kinetics of small molecules in structurally complicated specimens are critical for function. However, robust methods for precisely evaluating these parameters in the brain have been limited. Herein we report a new method termed "Fixation-driven chemical crosslinking of exogenous ligands (FixEL)" which traps and images exogenously administered molecules-of-interest (MOI) in complex tissues. This method relies on proteins-MOI interactions, and chemical crosslinking of amine-tethered MOI with paraformaldehyde used for perfusion fixation. FixEL is used to obtain images of the distribution of the small molecules and their dynamics, which addresses selective/nonselective binding to proteins, time-dependent localization changes, and diffusion/retention kinetics of MOI such as PET tracer derivatives or drug-like small molecules. Clear imaging of a nanobody distributed in the whole brain was also achieved with high spatial resolution using 2D/3D mode.

neuroscience↗

bioRxiv2021

A functional topography within the cholinergic basal forebrain for processing sensory cues associated with reward and punishment

Basal forebrain cholinergic neurons (BFCNs) project throughout the cortex to regulate arousal, stimulus salience, plasticity, and learning. Although often treated as a monolithic structure, the basal forebrain features distinct connectivity along its anteroposterior axis that could impart regional differences in BFCN processing. Here, we performed simultaneous bulk calcium imaging from anterior and posterior BFCNs over a one-month period of variable reinforcement learning in mice. BFCNs in both regions showed equivalently weak responses to unconditioned visual stimuli and anticipated rewards. Anterior BFCNs in the horizontal limb of the diagonal band were more responsive to reward omission, more accurately classified behavioral outcomes, and more closely tracked fluctuations in pupil-indexed global brain state. Posterior BFCNs in globus pallidus and substantia innominata were more responsive to unconditioned auditory stimuli, orofacial movements, aversive reinforcement, and showed robust associative plasticity for punishment-predicting cues. These results identify a functional topography that diversifies cholinergic modulatory signals broadcast to downstream brain regions.

neuroscience↗

bioRxiv2021

Site-specific steric control of SARS-CoV-2 spike glycosylation

A central tenet in the design of vaccines is the display of native-like antigens in the elicitation of protective immunity. The abundance of N-linked glycans across the SARS-CoV-2 spike protein is a potential source of heterogeneity between the many different vaccine candidates under investigation. Here, we investigate the glycosylation of recombinant SARS-CoV-2 spike proteins from five different laboratories and compare them against infectious virus S protein. We find patterns which are conserved across all samples and this can be associated with site-specific stalling of glycan maturation which act as a highly sensitive reporter of protein structure. Molecular dynamics (MD) simulations of a fully glycosylated spike support s a model of steric restrictions that shape enzymatic processing of the glycans. These results suggest that recombinant spike-based SARS-CoV-2 immunogen glycosylation reproducibly recapitulates signatures of viral glycosylation.

biochemistry↗

bioRxiv2021

The Cell Surface Receptors Ror1/2 Control Cardiac Myofibroblast Differentiation

BackgroundA hallmark of heart failure is cardiac fibrosis, which results from the injury-induced differentiation response of resident fibroblasts to myofibroblasts that deposit extracellular matrix. During myofibroblast differentiation, fibroblasts progress through polarization stages of early pro-inflammation, intermediate proliferation, and late maturation, but the regulators of this progression are poorly understood. Planar cell polarity receptors, receptor tyrosine kinase like orphan receptor 1 and 2 (Ror1/2), can function to promote cell differentiation and transformation. In this study, we investigated the role of the Ror1/2 in a model of heart failure with emphasis on myofibroblast differentiation. Methods and ResultsThe role of Ror1/2 during cardiac myofibroblast differentiation was studied in cell culture models of primary murine cardiac fibroblast activation and in knockout mouse models that underwent transverse aortic constriction (TAC) surgery to induce cardiac injury by pressure overload. Expression of Ror1 and Ror2 were robustly and exclusively induced in fibroblasts in hearts after TAC surgery, and both were rapidly upregulated after early activation of primary murine cardiac fibroblasts in culture. Cultured fibroblasts isolated from Ror1/2-KO mice displayed a pro-inflammatory phenotype indicative of impaired myofibroblast differentiation. Although the combined ablation of Ror1/2 in mice did not result in a detectable baseline phenotype, TAC surgery led to the death of all mice by day 6 that was associated with myocardial hyper-inflammation and vascular leakage. ConclusionsTogether, these results show that Ror1/2 are essential for the progression of myofibroblast differentiation and for the adaptive remodeling of the heart in response to pressure overload.

cell biology↗

bioRxiv2021

Mutational effects of ubiquitously present gamma radiation on Arabidopsis thaliana: insight into radiosensitivity in the reproductive stage

Earth has always been exposed to ionizing radiation from natural sources, and man-made sources have added to this radiation. In order to assess mutational effects of ubiquitously present radiation on plants, we performed a whole-genome resequencing analysis of mutations induced by chronic irradiation throughout the life-cycle of Arabidopsis thaliana under controlled conditions. Resequencing data from 12 M1 lines and 36 M2 progeny derived under gamma-irradiation conditions ranging from 0.0 to 2.0 Gy/d were obtained to identify de novo mutations, including single base substitutions (SBSs) and small insertions/deletions (INDELs). The relationship between de novo mutation frequency and a low-to-middling dose of radiation was assessed by statistical modeling. The increasing of de novo mutations in response to doses of irradiation fit the negative binomial model, accounting for the high variability of mutation frequency observed. Among the different types of mutations, SBSs were more prevalent than INDELs, with deletions being more frequent than insertions. Furthermore, we observed that the mutational effects of chronic radiation are more intensive during the reproductive stage. These outcomes could provide valuable insights into practical strategies for environmental radioprotection of plants on Earth and in space.

genetics↗

bioRxiv2021

Native-like SARS-CoV-2 spike glycoprotein expressed by ChAdOx1 nCoV-19/AZD1222 vaccine

Vaccine development against the SARS-CoV-2 virus focuses on the principal target of the neutralizing immune response, the spike (S) glycoprotein. Adenovirus-vectored vaccines offer an effective platform for the delivery of viral antigen, but it is important for the generation of neutralizing antibodies that they produce appropriately processed and assembled viral antigen that mimics that observed on the SARS-CoV-2 virus. Here, we describe the structure, conformation and glycosylation of the S protein derived from the adenovirus-vectored ChAdOx1 nCoV-19/AZD1222 vaccine. We demonstrate native-like post-translational processing and assembly, and reveal the expression of S proteins on the surface of cells adopting the trimeric prefusion conformation. The data presented here confirms the use of ChAdOx1 adenovirus vectors as a leading platform technology for SARS-CoV-2 vaccines.

microbiology↗

bioRxiv2020

Impaired phosphatidylethanolamine metabolism activates a reversible stress response that detects and resolves mutant mitochondrial precursors

Phosphatidylethanolamine made in mitochondria has long been recognized as an important precursor for phosphatidylcholine production that occurs in the endoplasmic reticulum (ER). Recently, the strict mitochondrial localization of the enzyme that makes PE in the mitochondrion, phosphatidylserine decarboxylase 1 (Psd1), was questioned. Since a dual localization of Psd1 to the ER would have far-reaching implications, we initiated our study to independently re-assess the subcellular distribution of Psd1. Our results support the unavoidable conclusion that the vast majority, if not all, of functional Psd1 resides in the mitochondrion. Through our efforts, we discovered that mutant forms of Psd1 that impair a self-processing step needed for it to become functional are dually localized to the ER when expressed in a PE-limiting environment. We conclude that severely impaired cellular PE metabolism provokes an ER-assisted adaptive response that is capable of identifying and resolving nonfunctional mitochondrial precursors.

cell biology↗

bioRxiv2020

Comprehensive analysis of Japanese population history by detecting ancestry-marker polymorphisms without using ancestral genomic information

Modern Japanese have two major ancestral populations: the indigenous Jomon hunter gatherers and continental East Asian farmers. To figure out the formation process of current Japanese population, we developed a reference-free detection method of variants derived from ancestral populations using a summary statistic, the ancestry-marker index (AMI). We confirmed by computer simulations that AMI can detect ancestry-derived variants even in an admixed population of recently diverged source populations with high accuracy, which cannot be achieved by the most widely used statistics, S*, for identifying archaic ancestry. We applied the AMI to modern Japanese samples and identified 208,648 single nucleotide polymorphisms (SNPs) that were likely derived from the Jomon people (Jomon-derived variants). The analysis of Jomon-derived variants in 10,842 modern Japanese individuals recruited from all over Japan revealed that the admixture proportions of the Jomon people varied between prefectures, probably due to the differences of population sizes of immigrants in the final Jomon to the Yayoi period. The estimated allele frequencies of genome-wide SNPs in the ancestral populations of modern Japanese suggested their phenotypic characteristics possibly for adaptation to their respective livelihoods; higher triglycerides and blood sugar for the Jomon ancestry and higher C-reactive protein and eosinophil counts for continental ancestry. According to our findings, we propose a formation model of modern Japanese population; regional variations in admixture proportions of the Jomon people and continental East Asians formed genotypic and phenotypic gradations of current Japanese archipelago populations.

evolutionary biology↗

bioRxiv2020

Two-component spike nanoparticle vaccine protects macaques from SARS-CoV-2 infection

The SARS-CoV-2 pandemic is continuing to disrupt personal lives, global healthcare systems and economies. Hence, there is an urgent need for a vaccine that prevents viral infection, transmission and disease. Here, we present a two-component protein-based nanoparticle vaccine that displays multiple copies of the SARS-CoV-2 spike protein. Immunization studies show that this vaccine induces potent neutralizing antibody responses in mice, rabbits and cynomolgus macaques. The vaccine-induced immunity protected macaques against a high dose challenge, resulting in strongly reduced viral infection and replication in upper and lower airways. These nanoparticles are a promising vaccine candidate to curtail the SARS-CoV-2 pandemic.

immunology↗