Biology subjects

Abe, K.

Publications and source records attributed to Abe, K..

At least 55 records · Page 3Linked to original sources

bioRxiv2023

Mechanical Netrin-1-DCC Coupling Mediates Netrin-1-induced Axonal Haptotaxis

The growth cone, a motile structure located at the tip of growing axon, senses extracellular guidance cues and translates them into directional forces that drive axon outgrowth and guidance. Axon guidance directed by chemical cues on the extracellular adhesive substrate is termed haptotaxis. Recent studies reported that netrin-1 on the substrate functions as a haptotactic axon guidance cue. However, the mechanism mediating netrin-1-induced axonal haptotaxis remains unclear. Here, we demonstrate that substrate-bound netrin-1 induces axonal haptotaxis by facilitating physical interactions between the netrin-1 receptor, DCC, and the adhesive substrates. DCC serves as an adhesion receptor for netrin-1. The clutch molecule shootin1a interacted with DCC, linking it to actin filament retrograde flow at the growth cone. Speckle imaging analyses showed that DCC underwent either grip (stop) and retrograde slip on the adhesive substrate. The grip state was more prevalent on netrin-1-coated substrate compared to the control substrate polylysine, thereby transmitting larger traction force on the netrin-1-coated substrate. Furthermore, disruption of the linkage between actin flow and DCC by shootin1 knockout impaired netrin-1-induced axonal haptotaxis. These findings indicate that the directional force for netrin-1-induced haptotaxis is exerted on the substrates through the mechanical coupling between netrin-1 and DCC which occurs asymmetrically under the growth cone.

cell biology↗

bioRxiv2023

Transcription factor activity profiling reveals the role of REST and LEF1 in the recovery from depression

Psychophysiological disorders chronically impair brain functions, often accompanied by dysregulation of multiple genes, suggesting a multifaceted etiology behind the symptoms. To explore transcription factors (TFs) involved in such transcriptomic changes, we analyzed TF-activity profiles (TFAPs) from the brains of mice experienced chronic stress, and revealed alteration in TF-activity correlating with their pathophysiological phenotypes. We identified REST/NRSF and TCF/LEF associated with depressive phenotypes and discovered that neuropsychiatric drugs sertraline and lithium influence REST- and TCF/LEF-activity, both in vitro and in vivo, thereby affecting gene expression profiles. Pharmacological or genetic manipulation of REST- or TCF/LEF-activity in defeated mice impacts post-stress recovery from depressive phenotypes, with combined treatment further augmenting the outcomes. Our TFAP analysis enhances understanding of molecular mechanisms underpinning chronic diseases, aiding future therapeutic strategy development.

neuroscience↗

bioRxiv2023

A deep generative model for estimating single-cell RNA splicing and degradation rates

AO_SCPLOWBSTRACTC_SCPLOWMessenger RNA splicing and degradation are critical for gene expression regulation, the abnormality of which leads to diseases. Previous methods for estimating kinetic rates have limitations, assuming uniform rates across cells. We introduce DeepKINET, a deep generative model that estimates splicing and degradation rates at single-cell resolution from scRNA-seq data. DeepKINET outperformed existing methods on simulated and metabolic labeling datasets. Applied to forebrain and breast cancer data, it identified RNA-binding proteins responsible for kinetic rate diversity. DeepKINET also analyzed the effects of splicing factor mutations on target genes in erythroid lineage cells. DeepKINET effectively reveals cellular heterogeneity in post-transcriptional regulation.

bioinformatics↗

bioRxiv2023

Survivability and Life Support in Sealed Mini-Ecosystems with Simulated Planetary Soils

Establishing a sustainable life-support system for space exploration is challenging due to the vast distances, costs, and differing environments from Earth. Using insights from the Biosphere 2 experiment, we introduced the "Ecosphere" and "Biosealed" systems in custom containers to replicate Earths ecosystems, suggesting feasible space migration through transplanting Earth-like biomes. Over four years, we gained deeper insights into these enclosed ecosystems. Moisture deficiency was a major obstacle to plant growth, which we addressed by incorporating a groundwater layer in the containers. We underscored the critical role of microorganisms in building and sustaining these ecosystems. However, temperature spikes from sunlight threatened stability. Our experiments confirmed fruit flies survival on plant-produced oxygen and photosynthetic bacteria. Interactions between plants, microbes, and simulated space soils were examined. Detailed analysis unveiled diverse microbes shaping both confined and simulated space environments. Major findings include the symbiotic relationship of plants with cyanobacteria, the potential of LED lighting in sun-limited missions, and challenges with ethylene gas and moisture. Microbial integration in rough soils holds promise for seed germination, but understanding their role in space soils is crucial. Our research offers a comprehensive foundation for future space life-support systems and underlines potential concerns about microbes affecting human health.

ecology↗

bioRxiv2023

Cross-platform comparison of highly-sensitive immunoassays for inflammatory markers in a COVID-19 cohort

A variety of commercial platforms are available for the simultaneous detection of multiple cytokines and associated proteins, often employing antibody pairs to capture and detect target proteins. In this study, we comprehensively evaluated the performance of three distinct platforms: the fluorescent bead-based Luminex assay, the proximity extension-based Olink assay, and a novel proximity ligation assay platform known as Alamar NULISAseq. These assessments were conducted on serum samples from the NIH IMPACC study, with a focus on three essential performance metrics: detectability, correlation, and differential expression. Our results reveal several key findings. Firstly, the Alamar platform demonstrated the highest overall detectability, followed by Olink and then Luminex. Secondly, the correlation of protein measurements between the Alamar and Olink platforms tended to be stronger than the correlation of either of these platforms with Luminex. Thirdly, we observed that detectability differences across the platforms often translated to differences in differential expression findings, although high detectability did not guarantee the ability to identify meaningful biological differences. Our study provides valuable insights into the comparative performance of these assays, enhancing our understanding of their strengths and limitations when assessing complex biological samples, as exemplified by the sera from this COVID-19 cohort.

immunology↗

bioRxiv2023

Ataxia-telangiectasia mutated (Atm) disruption sensitizes spatially-directed H3.3K27M/TP53 diffuse midline gliomas to radiation therapy

Diffuse midline gliomas (DMGs) are lethal brain tumors characterized by p53-inactivating mutations and oncohistone H3.3K27M mutations that rewire the cellular response to genotoxic stress, which presents therapeutic opportunities. We used RCAS/tv-a retroviruses and Cre recombinase to inactivate p53 and induce K27M in the native H3f3a allele in a lineage- and spatially-directed manner, yielding primary mouse DMGs. Genetic or pharmacologic disruption of the DNA damage response kinase Ataxia-telangiectasia mutated (ATM) enhanced the efficacy of focal brain irradiation, extending mouse survival. This finding suggests that targeting ATM will enhance the efficacy of radiation therapy for p53-mutant DMG but not p53-wildtype DMG. We used spatial in situ transcriptomics and an allelic series of primary murine DMG models with different p53 mutations to identify transactivation-independent p53 activity as a key mediator of such radiosensitivity. These studies deeply profile a genetically faithful and versatile model of a lethal brain tumor to identify resistance mechanisms for a therapeutic strategy currently in clinical trials.

cancer biology↗

bioRxiv2023

Toolbox for Gromov-Wasserstein optimal transport: Application to unsupervised alignment in neuroscience

BackgroundUnderstanding how sensory stimuli are represented across different brains, species, and artificial neural networks is a critical topic in neuroscience. Traditional methods for comparing these representations typically rely on supervised alignment, which assumes direct correspondence between stimuli representations across brains or models. However, it has limitations when this assumption is not valid, or when validating the assumption itself is the goal of the research. New methodTo address the limitations of supervised alignment, we propose an unsupervised alignment method based on Gromov-Wasserstein optimal transport (GWOT). GWOT optimally identifies correspondences between representations by leveraging internal relationships without external labels, revealing intricate structural correspondences such as one-to-one, group-to-group, and shifted mappings. ResultsWe provide a comprehensive methodological guide and introduce a toolbox called GWTune for using GWOT in neuroscience. Our results show that GWOT can reveal detailed structural distinctions that supervised methods may overlook. We also demonstrate successful unsupervised alignment in key data domains, including behavioral data, neural activity recordings, and artificial neural network models, demonstrating its flexibility and broad applicability. Comparison with existing methodsUnlike traditional supervised alignment methods such as Representational Similarity Analysis, which assume direct correspondence between stimuli, GWOT provides a nuanced approach that can handle different types of structural correspondence, including fine-grained and coarse correspondences. Our method would provide richer insights into the similarity or difference of representations by revealing finer structural differences. ConclusionWe anticipate that our work will significantly broaden the accessibility and application of unsupervised alignment in neuroscience, offering novel perspectives on complex representational structures. By providing a user-friendly toolbox and a detailed tutorial, we aim to facilitate the adoption of unsupervised alignment techniques, enabling researchers to achieve a deeper understanding of cross-brain and cross-species representation analysis.

neuroscience↗

bioRxiv2023

Enhanced Aversive Signals During Classical Conditioning in Dopamine Axons in Medial Prefrontal Cortex

Midbrain dopamine neurons impact neural processing in the prefrontal cortex (PFC) through mesocortical projections. However, the signals conveyed by dopamine projections to the PFC remain unclear, particularly at the single-axon level. Here, we investigated dopaminergic axonal activity in the medial PFC (mPFC) during reward and aversive processing. By optimizing microprism-mediated two-photon calcium imaging of dopamine axon terminals, we found diverse activity in dopamine axons responsive to both reward and aversive stimuli. Some axons exhibited a preference for reward, while others favored aversive stimuli, and there was a strong bias for the latter at the population level. Long-term longitudinal imaging revealed that the preference was maintained in reward- and aversive-preferring axons throughout classical conditioning in which rewarding and aversive stimuli were paired with preceding auditory cues. However, as mice learned to discriminate reward or aversive cues, a cue activity preference gradually developed only in aversive-preferring axons. We inferred the trial-by-trial cue discrimination based on machine learning using anticipatory licking or facial expressions, and found that successful discrimination was accompanied by sharper selectivity for the aversive cue in aversive-preferring axons. Our findings indicate that a group of mesocortical dopamine axons encodes aversive-related signals, which are modulated by both classical conditioning across days and trial-by-trial discrimination within a day. Impact statementTwo-photon calcium imaging revealed that many mesocortical dopamine axons show enhanced selectivity for aversive cue processing during classical conditioning.

neuroscience↗

bioRxiv2023

Machine-learning-based motion tracking revealed the inverse correlation between adhesivity and surface motility of the leptospirosis spirochete

Bacterial motility is generally a critical virulence factor for pathogenic species, and thus studies on bacterial motility are significant given that they elucidate the mechanisms of infection and disease. Although fluorescent labeling has been the mainstream approach to detecting individual bacteria in a population or in host tissues, it contains problems related to protein expression stability and interference with bacterial physiology. Here, we applied machine learning to microscopic image analysis to achieve label-free motion tracking of the zoonotic bacterium Leptospira interrogans on cultured animal cells. The label-free method allowed us to measure various leptospiral strains isolated from human and animal patients, and natural reservoirs, showing that fast mobility on kidney cells tends to result in severe symptom. We also analyzed the surface locomotion of mutant strains lacking outer membrane proteins (OMPs) and revealed that the loss of OMPs reduced adherence and facilitated motility on cultured kidney cells. The behavior of clinical isolates and OMP-deficient mutants on cultured cells showed the inverse correlation between adhesion and mobility, which could affect infection outcomes. Thus, our computer vision technique eliminated the restriction on available bacterial strains and provided information that could help in understanding the mechanisms underlying motility-dependent bacterial pathogenicity.

biophysics↗

bioRxiv2023

Development of two mouse strains conditionally expressing bright luciferases with distinct emission spectra as new tools for in vivo imaging

In vivo bioluminescence imaging (BLI) has been an invaluable noninvasive method to visualize molecular and cellular behaviors in laboratory animals. Bioluminescent reporter mice possessing luciferases for general use have been limited to a classical luciferase, Luc2, from Photinus pyralis, and have been extremely powerful for various in vivo studies. However, applicability of reporter mice for in vivo BLI could be further accelerated by increasing light intensity using other luciferases and/or improving the biodistribution of their substrates in animal body. Here, we created two Cre-dependent reporter mice incorporating luciferases: oFluc derived from Pyrocoeli matsumurai and Akaluc, both of which had been reported previously to be brighter than Luc2 when using appropriate substrates; we then tested their bioluminescence in neural tissues and other organs in living mice. When expressed throughout the body, both luciferases emitted an intense yellow (oFluc) or far-red (Akaluc) light easily visible to the naked eye. Moreover, oFluc and Akaluc were similarly bright in the pancreas for in vivo BLI. However, Akaluc was superior to oFluc for brain imaging, because its substrate, AkaLumine-HCl, was distributed to the brain more efficiently than the oFluc substrate, D-luciferin. We also demonstrated that the light produced by oFluc and Akaluc was sufficiently spectrally distinct for dual-color imaging in a single living mouse. Taken together, these novel bioluminescent reporter mice are an ideal source of cells with bright bioluminescence and may facilitate the in vivo BLI of various tissues/organs for preclinical and biomedical research in combination with a wide variety of Cre-driver mice.

bioengineering↗

bioRxiv2022

Cleavage of α-1,4-Glycosidic Linkages by the Glycosylphosphatidylinositol-Anchored α-Amylase AgtA Decreases the Molecular Weight of Cell Wall α-1,3-Glucan in Aspergillus oryzae

Aspergillus fungi contain -1,3-glucan with a low proportion of -1,4-glucan as a major cell wall polysaccharide. Glycosylphosphatidylinositol (GPI)-anchored -amylases are conserved in Aspergillus fungi. The GPI-anchored -amylase AmyD in Aspergillus nidulans has been reported to directly suppress the biosynthesis of cell wall -1,3-glucan but not to degrade it in vivo. However, the detailed mechanism of cell wall -1,3-glucan biosynthesis regulation by AmyD remains unclear. Here we focused on AoAgtA, which is encoded by the Aspergillus oryzae agtA gene, an ortholog of the A. nidulans amyD gene. Similar to findings in A. nidulans, agtA overexpression in A. oryzae grown in submerged culture decreased the amount of cell wall -1,3-glucan and led to the formation of smaller hyphal pellets in comparison with the wild-type strain. We analyzed the enzymatic properties of recombinant (r)AoAgtA produced in Pichia pastoris and found that it degraded soluble starch, but not linear bacterial -1,3-glucan. Furthermore, rAoAgtA cleaved 3--maltotetraosylglucose with a structure similar to the predicted boundary structure between the -1,3-glucan main chain and a short spacer composed of -1,4-linked glucose residues in cell wall -1,3-glucan. Interestingly, rAoAgtA randomly cleaved only the -1,4-glycosidic bonds of 3--maltotetraosylglucose, indicating that AoAgtA may cleave the spacer in cell wall -1,3-glucan. Consistent with this hypothesis, heterologous overexpression of agtA in A. nidulans decreased the molecular weight (MW) of cell wall -1,3-glucan. These in vitro and in vivo properties of AoAgtA suggest that GPI-anchored -amylases can degrade the spacer -1,4-glycosidic linkages in cell wall -1,3-glucan before its insolubilization, and this spacer cleavage decreases the MW of cell wall -1,3-glucan in vivo.

microbiology↗

bioRxiv2022

Single-molecule tracking of Nanog and Oct4 in mouse embryonic stem cells

Nanog and Oct4 are core transcription factors in a gene regulatory network that regulate hundreds of target genes for pluripotency maintenance in mouse embryonic stem cells. To understand their function as a gatekeeper or a pioneer factor at the molecular scale, we quantified the residence time on target loci, fluctuation at the loci, and interaction clustering by visualizing single molecules of Nanog and Oct4 in a living nucleus during the pluripotency loss. Interestingly, Nanog interacted longer with its target loci in the lower Nanog expression state or at the onset of differentiation, indicating the possibility of a new feedback mechanism to maintain Nanog expression. The interaction time of Nanog and Oct4 corresponded to their fluctuation and interaction clustering, which depended on their expression or differentiation state, respectively. The DNA viscoelasticity near the Oct4 target locus remained flexible during the differentiation, reflecting its role as a pioneer factor. Based on these results, we propose a new feedback mechanism for pluripotency maintenance in which Nanog function is prolonged, corresponding to chromatin condensation, and Oct4 reopens the condensation.

biophysics↗

bioRxiv2022

Complete sequencing of the mouse pseudoautosomal region, the most rapidly evolving 'chromosome'

The pseudoautosomal region (PAR) of mammalian sex chromosomes is a small region of sequence identity that allows pairing, crossover, recombination, and proper chromosome segregation during male meiosis. The structure of the mouse PAR is largely unknown. Here, we developed a new assembly method to robustly resolve repetitive sequences and employed highly accurate long-read sequencing data to reveal the entire PAR sequence. The PAR of the widely-used inbred strain C57BL/6J is [~]700 kb, comprising 10 protein- coding genes in a mass of complex repetitive sequences. A large segmental duplication exhibiting copy-number polymorphisms even among C57BL/6J littermates is present. High GC-content exons and short introns are common properties of PAR genes and are the consequence of maintaining gene function, while PAR is rapidly evolving. Elucidating the mouse PAR sequence completes the mouse euchromatic genome sequencing and enables the exploration of the function and evolution of the PAR using modern molecular genetic approaches.

genomics↗

bioRxiv2021

A glycosylphosphatidylinositol-anchored α-amylase encoded by amyD contributes to a decrease in the molecular mass of cell wall α-1,3-glucan in Aspergillus nidulans

-1,3-Glucan is one of the main polysaccharides in the cell wall of Aspergillus nidulans. We previously revealed that it plays a role in hyphal aggregation in liquid culture, and that its molecular mass (MM) in an agsA-overexpressing (agsAOE) strain was larger than that in an agsB-overexpressing (agsBOE) strain. The mechanism that regulates the MM of -1,3-glucan is poorly understood. Although the gene amyD, which encodes glycosyl-phosphatidylinositol (GPI)-anchored -amylase (AmyD), is involved in the biosynthesis of -1,3-glucan in A. nidulans, how it regulates this biosynthesis remains unclear. Here we constructed strains with disrupted amyD ({Delta}amyD) or overexpressed amyD (amyDOE) in the genetic background of the ABPU1 (wild-type), agsAOE, or agsBOE strain, and characterized the chemical structure of -1,3-glucans in the cell wall of each strain, focusing on their MM. The MM of -1,3-glucan from the agsBOE amyDOE strain was smaller than that in the parental agsBOE strain. In addition, the MM of -1,3-glucan from the agsAOE {Delta}amyD strain was greater than that in the agsAOE strain. These results suggest that AmyD is involved in decreasing the MM of -1,3-glucan. We also found that the C-terminal GPI-anchoring region is important for these functions.

microbiology↗

bioRxiv2021

A Novel Ex Vivo Peritoneal Model to Investigate Mechanisms of Peritoneal Metastasis in Gastric Adenocarcinoma

Peritoneal metastases (PM) portend limited survival in patients with Gastric Adenocarcinoma (GCa), and strategies to prevent and/or more effectively treat PM are needed. Existing models are limited in recapitulating key elements of the peritoneal metastatic cascade. To explore the underlying cellular and molecular mechanisms of PM, we have developed an ex vivo human peritoneal explant model. Fresh peritoneal tissue samples were obtained from patients undergoing abdominal surgery and suspended, mesothelial layer down but without direct contact, above a monolayer of red-fluorescent stained AGS human GCa cells for 24hrs, then washed and cultured for a further 3 days. Implantation and invasion of GCa cells within the explant were examined using real-time confocal fluorescence microscopy. Superficial implantation of AGS GCa cells within the mesothelial surface was readily detected, and colonies expanded over 3 days. To investigate the sensitivity of the model to altered GCa cellular implantation, we stably transfected AGS cells with E-Cadherin, restoring the E-Cadherin that they otherwise lack. This markedly suppressed implantation and invasion of AGS cells into the submesothelial mesenchymal layer. Here we show that this ex vivo human peritoneal explant model is responsive to manipulation of genetic factors that regulate peritoneal implantation and invasion by GCa cells, with reproducible results.

cancer biology↗

bioRxiv2021

scMM: Mixture-of-experts multimodal deep generative model for single-cell multiomics data analysis.

AO_SCPLOWBSTRACTC_SCPLOWThe recent development in single-cell multiomics analysis has enabled simultaneous detection of multiple traits at the single-cell level, thus providing deeper insights into the cellular phenotypes and functions in diverse tissues. However, currently, it is challenging to infer the joint representations and learn relationships among multiple modalities from complex multimodal single-cell data. Herein, we present scMM, a novel deep generative model-based framework for the extraction of interpretable joint representations and cross-modal generation. scMM addresses the complexity of data by leveraging a mixture-of-experts multimodal variational autoencoder. The pseudocell generation strategy of scMM compensates for the limited interpretability of deep learning models and discovered multimodal regulatory programs associated with latent dimensions. Analysis of recently produced datasets validated that scMM facilitates high-resolution clustering with rich interpretability. Furthermore, we show that cross-modal generation by scMM leads to more precise prediction and data integration compared with the state-of-the-art and conventional approaches.

bioinformatics↗

bioRxiv2020

Single-cell transcriptomics, scRNA-Seq and C1 CAGE discovered distinct phases of pluripotency during naïve-to-primed conversion in mice

BackgroundTwo types of mammalian pluripotent stem cells (PSC), i.e. naive and primed possess distinct cellular characteristics. It is largely unknown how these differences are generated during naive-to-primed transition process. We have established a robust in vitro transition system using a Wnt inhibitor for the first time and analyzed dynamic changes in cellular status via single-cell RNA-sequencing and C1 CAGE analyses. ResultsAnalysis of known marker genes suggested that the cell transition process progresses as expected. However, cluster analyses revealed a sudden increase in expression profile diversities three and four days after induction of the transition. These expression diversities can be reconciled by the presence of two subpopulations with distinct transcription profiles emerging at these time points. One of the subpopulations appears transiently, and surprisingly these cells showed a global downregulation of gene expression. Moreover, initiation of random X chromosome inactivation (XCI) coincides with the appearance of these transient cells. The other subpopulation can be maintained as a stem cell line and possesses expression profiles more similar to those of primed epiblast stem cells (EpiSC) than embryonic stem cells (ESC). However, there are important differences in gene expression related to epithelial-mesenchymal transition (EMT), suggesting that this subpopulation may represent a novel pluripotent state that has an intermediate cellular phenotype between ESC and EpiSC. ConclusionsThese findings should contribute to our understanding of the establishment and maintenance of distinct differentiation statuses of mammalian PSCs and provide new insights into the pluripotency spectrum in general.

genomics↗

bioRxiv2020

Implications of back-and-forth motion and powerful propulsion for spirochetal invasion

The spirochete Leptospira spp. can move in liquid and on a solid surface using two periplasmic flagella (PFs), and its motility is an essential virulence factor for the pathogenic species. Mammals are infected with the spirochete through the wounded dermis, which implies the importance of behaviors on the boundary with such viscoelastic milieu; however, the leptospiral pathogenicity involving motility remains unclear. We used a glass chamber containing a gel area adjoining the leptospiral suspension to resemble host dermis exposed to contaminated water and analyzed the motility of individual cells at the liquid-gel border. Insertion of one end of the cell body to the gel increased switching of the swimming direction. Moreover, the swimming force of Leptospira was also measured by trapping single cells using an optical tweezer. It was found that they can generate [~]17 pN of force, which is [~]30 times of the swimming force of Escherichia coli. The force-speed relationship suggested the load-dependent force enhancement and showed that the power (the work per unit time) for the propulsion is [~]3.1x10-16 W, which is two-order of magnitudes larger than the propulsive power of E. coli. The powerful and efficient propulsion of Leptospira using back-and-forth movements could facilitate their invasion.

microbiology↗