Biology subjects

Takahashi, K.

Publications and source records attributed to Takahashi, K..

At least 37 records · Page 2Linked to original sources

bioRxiv2026

Optimizing Network-Level TMS-fMRI: Benchmarking a Novel TMS-Compatible "Sushi" MR Coil

Concurrent TMS-fMRI can map how stimulation affects both the targeted cortex and connected brain-wide networks, but this requires MR receive hardware that allows TMS coil placement while preserving reliable whole-brain BOLD sensitivity. We developed and benchmarked a practical TMS-compatible "Sushi" MR receive setup assembled from two flexible 18-channel body arrays. Across six experiments, we tested functional readout validity, signal quality, and active TMS-fMRI compatibility. Resting-state fMRI (n = 12) and verbal N-back task-fMRI (n = 8) were acquired with Sushi, a commercially available 2x7-channel Surface setup, and a standard 64-channel head/neck array. Functional similarity to the 64-channel reference was quantified with spatial overlap, and multi-echo combination (MEcomb) was tested as a post-acquisition signal optimization strategy. Sushi recovered subject-specific resting-state networks that more closely matched the 64-channel reference than Surface, with no significant difference from the 64-channel test-retest reference. For task-fMRI, MEcomb increased task-map similarity for Sushi, whereas setup comparisons within each pipeline were not significant. MEcomb also improved resting-state similarity and increased temporal signal-to-noise ratio (tSNR) across receive setups. In phantom measurements, TMS coil placement produced spatially graded tSNR reductions relative to the no-TMS-coil condition, strongest near the coil. In one participant, active interleaved single-pulse TMS-fMRI over two cortical sites showed no detectable pulse-locked image artifacts; whole-brain MEcomb tSNR during active TMS-fMRI was reduced by 2.6-6.9% relative to the no-coil/no-stimulation reference. Together, Sushi and MEcomb provide complementary hardware and processing tools for TMS-compatible whole-brain fMRI. This combination supports network-level functional readouts while preserving feasibility for active interleaved TMS-fMRI.

neuroscience↗

bioRxiv2025

LLM-scCurator: Data-centric feature distillation for zero-shot cell-type annotation

Zero-shot cell-type annotation with language models degrades when marker lists are dominated by biological-noise programs (e.g., ribosomal/cell-cycle/stress). We present LLM-scCurator, a data-centric, backend-agnostic framework using pre-prompt noise masking and Gini-informed distillation to recover identity markers. Across benchmarks, LLM-scCurator improves ontology-aware hierarchical accuracy from 78.4% to 86.1% (52 clusters) over naive prompting, approaching supervised/reference-transfer performance without training data. LLM-scCurator extends to spatial transcriptomics (Visium, Xenium) for reference-free discovery of fine-grained niches.

bioinformatics↗

bioRxiv2025

Chronic oral cannabidiol delays or prevents seizures in a mouse model of CLN2 disease

A growing body of literature describes the anti-inflammatory, neuroprotective, and anti-epileptic properties of the cannabis sativa constituent cannabidiol, suggesting that it might play a useful role in the treatment of neurodegenerative diseases. Late infantile neuronal ceroid lipofuscinosis (CLN2 disease) is a rare pediatric neurodegenerative disorder resulting from an inherited dysfunction of the lysosome. CLN2 disease, and its representative animal models, display neuroimmune response, neuroinflammation, neurodegeneration, and epileptic seizures, and these symptoms are all touted as potential targets of cannabidiol therapeutic benefit. Here, we treated a valid model of CLN2 disease with long-term daily cannabidiol (300mg/kg) from 1 month of age until disease end stage and evaluated epileptic seizures, lifespan, and markers of neuroimmune response. Chronic cannabidiol treatment significantly delayed or fully eliminated seizures in CLN2 mice compared to those treated with vehicle only, and the treatment led to a nearly significant extension of lifespan. These effects occurred in the absence of any therapeutic benefit to physiological markers of disease such as GFAP, CD68, and cytokine/chemokine reactivity. Taken together, we show that chronic treatment with cannabidiol confers significant anti-seizure benefit to the mouse model of CLN2 disease, and that it does not appear to do so by altering the inflammatory and neuroimmune markers traditionally used to track CLN2 disease progression.

neuroscience↗

bioRxiv2025

Osteochondrogenic Transdifferentiation of Vascular Smooth Muscle Cells and Microenvironmental Dynamics in Medial Arterial Calcification

BackgroundVascular calcification, particularly medial arterial calcification (MAC), considerably affects cardiovascular mortality. Current treatments are limited because of the unclear molecular mechanisms of MAC. This study aimed to establish an MAC mouse model using O-ring-induced transverse aortic constriction (OTAC) and to identify critical genes, pathways, and cellular interactions involved in MAC formation by combining lineage tracing technology and single-cell RNsA sequencing (scRNA-seq). MethodsWe developed an OTAC mouse model to mimic MAC. Adult C57BL/6J male mice underwent OTAC, and subsequent analyses were performed at various intervals. Histological and immunohistochemical evaluations were conducted to observe changes in vascular smooth muscle cells (VSMCs). Lineage tracing was used to confirm the origin of osteochondrogenic cells in the tunica media. Additionally, scRNA-seq was performed to capture the dynamic changes in VSMCs and other cell types involved in MAC development. ResultsOsteochondrogenic cells and the subsequent MAC in OTAC mice were observed using histological evaluations. Immunohistochemistry showed time-dependent decreases in -SMA expression and increases in SOX9 and RUNX2 expression in VSMCs. Lineage tracing using Myh11CreERT2;ROSA26-EGFP mice confirmed that osteochondrogenic cells originated from contractile VSMCs. scRNA-seq confirmed osteochondrogenic transdifferentiation of VSMCs and highlighted dynamic changes in surrounding cells involved in calcification-related and inflammatory processes. ConclusionThe OTAC method enabled a comprehensive understanding of the microenvironmental dynamics during MAC progression at the single-cell level. This powerful model provides a robust platform for future therapeutic intervention research. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/684711v1_ufig1.gif" ALT="Figure 1"> View larger version (58K): org.highwire.dtl.DTLVardef@c64634org.highwire.dtl.DTLVardef@12b0b9aorg.highwire.dtl.DTLVardef@106c5b4org.highwire.dtl.DTLVardef@19dd4f4_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2025

TCP3-mediated regulation of cell expansion in Arabidopsis thaliana

(1) Cell expansion is crucial for organ morphogenesis in multicellular organisms. Apoplast acidification triggers plant cell expansion. Plant hormones and transcription factors such as TEOSINTE BRANCHED, CYCLOIDEA, and PROLIFERATING CELL NUCLEAR ANTIGEN BINDING FACTORs (TCPs) control cell expansion. However, details regarding the regulatory mechanism of cell expansion for organ morphogenesis remain unclear. (2) In this study, we used molecular, biochemical, cellular, genetic, atomic force microscopy, and tensile testing analyses and showed that miR319-targeted TCPs integrated cell expansion with organ morphogenesis in Arabidopsis thaliana. (3) We found that TCPs directly induce the expression of genes encoding cell wall loosening proteins and SMALL AUXIN UP RNAs (SAURs), activators of plasma membrane-localized H+-ATPases. TCP-mediated activation of plasma membrane- localized H+-ATPases stimulates apoplast acidification, reduce cell stiffness, promote cell expansion, and thus exaggerate elongation of the hypocotyl. Ectopic expression of a SAUR gene in sextuple tcp mutant plants substantially recovered the hypocotyl morphology of the tcp mutant, providing genetic evidence of TCP-mediated SAUR regulation. (4) Collectively, our data show that TCPs regulate apoplast acidification for cell expansion.

plant biology↗

bioRxiv2025

Alphavirus replicon particle expressing IL-12 reprograms tumor-associated macrophages and neutrophils and induces anti-tumor immunity

Cancers evade anti-tumor immunity by the infiltration of immunosuppressive myeloid cells such as macrophages and neutrophils into the tumor microenvironment (TME). The limited efficacy of current immunotherapy is attributed to the presence of these immunosuppressive cells. Here, we describe a novel immunotherapy approach, using a single-cycle alphavirus replicon particle (VRP) that carries a self-amplifying RNA encoding interleukin-12 (IL-12). Leveraging the natural tropism of alphavirus to myeloid cells, single cell transcriptomic analyses of mouse tumor tissues demonstrated that intratumoral (i.t.) injection of the vector effectively reprogrammed tumor-associated macrophages and neutrophils toward a pro-inflammatory, anti-tumor phenotype via interferon signaling, further enhanced by IL-12 expression. The treatment also induced cytotoxic NK and T cell responses, resulting in a significant reduction of tumor growth and effectively preventing nodal and distant metastases in solid tumor mouse models. By targeting tumor-supporting myeloid cells to transform the TME into an immune-active state, our myeloid modulation approach has the potential to enhance the efficacy of current immunotherapies.

cancer biology↗

bioRxiv2025

Conformational analysis of liganded human hemoglobin by cryo electron microscopy

The long-standing debate on the preferred conformation of liganded hemoglobin (Hb) in solution has yet to be completely resolved. While some studies have used lyophilized human hemoglobin for structural studies by cryo-EM, we recently presented the first cryo-EM analysis of freshly prepared human and crocodilian Hbs. Further three-dimensional (3D) classification analysis of these datasets reveals distinct structural characteristics. CO-bound adult human Hb (CO-HbA) shows a mixture of conformations, with the R2 conformation most populated, R strongly represented, and other intermediate states present in sufficient quantity to produce maps. CO-bound crocodile Hb showed the R conformation and, unexpectedly, a smaller population of molecules in a T-like conformation. The amino acid substitution Glu {beta}39 [->] Arg, unique to crocodilian Hbs, appears to favour the R conformation over R2.

molecular biology↗

bioRxiv2025

Sleep stage-specific effects of 0.75 Hz phase-synchronized rTMS and tACS on delta frequency activity during sleep

Slow oscillatory activity during non-rapid-eye-movement (NREM) sleep plays a crucial role in both physical health and cognitive functions. Enhancing slow oscillatory activity during sleep has the potential to benefit these domains, yet an optimal stimulation protocol has not been established. This study aimed to investigate whether repetitive transcranial magnetic stimulation (rTMS), synchronized with the trough phase of 0.75 Hz transcranial alternating current stimulation (tACS) can modulate EEG activity in the delta frequency range during sleep and enhance cognitive functions. Healthy adults participated in a within-subject, counterbalanced study design comparing real and sham stimulation conditions. Combined rTMS and tACS was applied over the bilateral prefrontal cortex before sleep. We evaluated (1) power spectral density and functional connectivity within the delta frequency range during resting state and sleep, (2) retention of declarative memory learned before sleep, and (3) sleep parameters including spindle activity, sleep stage ratios, sleep onset latency and sleep efficiency. The combined rTMS and tACS protocol significantly increased delta oscillatory activity during the N3 sleep stage compared to sham. Functional connectivity, as measured by global efficiency, was enhanced during the N2 sleep stage. However, the stimulation did not improve declarative memory retention, spindle activity or other sleep parameters. These findings demonstrate the potential of combined rTMS and tACS as a non-invasive method to enhance delta oscillatory activity during sleep. While the stimulation did not improve memory performance, its ability to modulate delta activity during sleep suggests potential clinical applications for addressing pathological alterations in slow wave activity during sleep. State of SignificanceSlow oscillatory activity during non-rapid eye movement (NREM) sleep is relevant for declarative memory consolidation and overall sleep quality. However, noninvasive brain-stimulation methods to selectively modulate these oscillations remain elusive. In this study, we show that pre-sleep application of repetitive transcranial magnetic stimulation (rTMS) synchronized to the trough of 0.75 Hz transcranial alternating current stimulation (tACS) significantly enhances delta oscillatory activity during N3 sleep and increases global functional connectivity during N2 sleep in healthy humans without sleep disturbances without altering sleep structure. Our findings indicate that rTMS synchronized to the trough of the tACS exerts sleep-stage-specific effects on delta oscillatory activity. This stimulation protocol represents a promising approach for restoring pathological slow-wave activity during sleep in clinical populations.

neuroscience↗

bioRxiv2025

Imaging traumatic brain injuries in mice with potassium channel PET tracer 3F4AP

ObjectiveTraumatic brain injury (TBI) can lead to secondary injury, including axon and myelin damage, which contributes to long-term neurological deficits. The PET tracer [18F]3F4AP, a fluorinated derivative of the FDA-approved drug 4-aminopyridine, selectively binds to voltage-gated potassium (KV) channels, offering a novel approach to assess TBI-related node of Ranvier disruption and demyelination. This study evaluates [18F]3F4AP PET in penetrating and non-penetrating TBI models. MethodsEither controlled cortical impact (CCI, penetrating) or concussive (non-penetrating) TBI models were used to induce TBI in mice. Dynamic PET imaging with [18F]3F4AP was performed at time points of 0, 3, 7, 14, and/or 31 days post-injury (dpi), with quantitative analyses comparing tracer uptake in injured versus control regions. Luxol fast blue (LFB) staining was conducted to evaluate histological myelin loss. ResultsIn the CCI model, [18F]3F4AP PET imaging demonstrated a 34% increase in tracer uptake at the injury site at 7 dpi, correlating with histological evidence of demyelination. Tracer uptake gradually declined over time, reflecting potential remyelination. The concussive TBI model showed a smaller and more diffuse increase in uptake at 7 dpi compared to CCI. Conclusion[18F]3F4AP PET imaging effectively detects demyelination following TBI, with very high sensitivity in penetrating injuries. These findings highlight the potential of [18F]3F4AP as a valuable imaging biomarker for assessing TBI progression and/or therapeutic response. Further studies are warranted to explore its clinical applicability and comparison with other imaging modalities.

neuroscience↗

bioRxiv2025

A polymeric, PDMS-based large-scale skull replacement suitable for optical and mechanical access for long-term neuronal imaging, electrophysiology, and optogenetics

Many techniques to record and manipulate neuronal activity across large portions of the vertebrate brain, such as widefield and two-photon calcium imaging, electrophysiology, and optogenetics, are now available. However, few effective approaches enable both optical and mechanical access to the brain. In this work, we offer an in-depth guide for synthesizing, implanting, and using polydimethylsiloxane (PDMS) windows as skull replacements for chronic optical neuronal imaging. Furthermore, we provide instructions to perform viral injections and multi-site silicon probe implantation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/648966v1_ufig1.gif" ALT="Figure 1"> View larger version (71K): org.highwire.dtl.DTLVardef@1900a2dorg.highwire.dtl.DTLVardef@18c436corg.highwire.dtl.DTLVardef@19e6a4corg.highwire.dtl.DTLVardef@154c228_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO C_FIG

neuroscience↗

bioRxiv2025

Phase-synchronized 40 Hz transcranial electric and magnetic stimulation boosts gamma oscillations and working memory

Gamma oscillations play a crucial role in core cognitive functions such as memory processes. Enhancing gamma oscillatory activity, which is reduced in Alzheimers Disease, may have therapeutic potential, but effective interventions remain to be determined. Previous studies have shown that phase-synchronized electric and magnetic stimulation boosts brain oscillatory activities at theta, alpha, and delta frequency bands in different ways. The high-frequency gamma frequency band remains to be investigated. This study applies novel noninvasive brain stimulation techniques, namely phase-locked 40-Hz intermittent theta-burst stimulation (iTBS) and transcranial alternating current stimulation (tACS), and explores gamma oscillation changes in the brain. Thirty healthy young participants randomly underwent 40-Hz tACS (1), 40-Hz iTBS (2), two combined interventions (phase-locked iTBS to tACS peak sine wave or tACS trough sine wave) (3-4), and a sham condition (5). The target regions were the left and right dorsolateral prefrontal cortex and were stimulated by simultaneous tACS and iTBS. Gamma oscillatory activities (for 2 hours after intervention) were monitored following each intervention. Our results show that all stimulation protocols enhanced 40-Hz oscillatory power. The iTBS-tACS Peak shows the most significant and stable increase in gamma oscillatory activities (up to 2 hours), followed by 40-Hz tACS and 40-Hz iTBS. 40-Hz tACS and 40-Hz iTBS had the strongest acute effects (up to 30 min) on induced gamma oscillations, while 40-Hz tACS most consistently induced gamma oscillations for up to 2 hours in overall resting EEG data. Phase-synchronizing iTBS with tACS at 40 Hz and the 40 Hz tACS alone targeting the dorsolateral prefrontal cortex, may be a viable approach for inducing and stabilizing gamma oscillatory activity, particularly in conditions where endogenous gamma oscillatory is attenuated, such as Alzheimers Disease. Registration IDs:101017716 (CORDIS - European Union-funded project)

neuroscience↗

bioRxiv2025

Membrane vesicles of Shewanella oneidensis MR-1 enhance denitrification growth in a species-selective manner

Denitrification, a fundamental bacterial respiratory process that occurs in anoxic environments, plays a pivotal role in energy synthesis and the global nitrogen cycle. Although the significance of this process is well-recognized, its regulation within polymicrobial communities remains poorly understood, particularly concerning interspecies interactions. In this study, we investigated the role that bacterial membrane vesicles (MV) play in modulating denitrification across bacterial species. MV is known to carry specific biomolecules such as secondary metabolites, proteins, and nucleic acids, therefore considered to be a secretion pathway. We found that MV produced by Shewanella oneidensis enhanced denitrification in a species-specific manner. Bacteria with highly hydrophobic surfaces tended to respond to denitrification enhancement, suggesting that the MV-bacteria attachment process is the key to generating species specificity. Transcriptome analysis and isotopic metabolite tracking indicated that the MV influenced denitrifying activities, rather than the transcription of denitrification-related genes. We further demonstrated that c-type cytochromes in MV act as key components that enhance denitrification. These insights expand our understanding of bacterial ecology, highlighting the role of membrane vesicles in facilitating respiratory competition and cooperation in polymicrobial communities.

microbiology↗

bioRxiv2025

Bayesian inference of fitness landscapes via tree-structured branching processes

MotivationThe complex dynamics of cancer evolution, driven by mutation and selection, underlies the molecular heterogeneity observed in tumors. The evolutionary histories of tumors of different patients can be encoded as mutation trees and reconstructed in high resolution from single-cell sequencing data, offering crucial insights for studying fitness effects of and epistasis among mutations. Existing models, however, either fail to separate mutation and selection or neglect the evolutionary histories encoded by the tumor phylogenetic trees. ResultsWe introduce FiTree, a tree-structured multi-type branching process model with epistatic fitness parameterization and a Bayesian inference scheme to learn fitness landscapes from single-cell tumor mutation trees. Through simulations, we demonstrate that FiTree outperforms state-of-the-art methods in inferring the fitness landscape underlying tumor evolution. Applying FiTree to a single-cell acute myeloid leukemia dataset, we identify epistatic fitness effects consistent with known biological findings and quantify uncertainty in predicting future mutational events. The new model unifies probabilistic graphical models of cancer progression with population genetics, offering a principled framework for understanding tumor evolution and informing therapeutic strategies. Availability and implementationThe Python package FiTree and the analysis workflows are available at https://github.com/cbg-ethz/FiTree.

bioinformatics↗

bioRxiv2024

Rapid clonal selection within early hematopoietic cell compartments presages outcome to ivosidenib combination therapy

Acquired resistance to targeted non-intensive therapies is common in myeloid malignancies. Yet, key questions remain as to how rapidly resistant clones are selected by treatment and in which hematopoietic cell compartments clonal selection occurs. To address this gap, we studied clonal responses to ivosidenib + venetoclax {+/-} azacitidine combination therapy in 8 patients with IDH1-mutant myeloid malignancy. Whilst all 8 patients initially responded to treatment, 6 relapsed and 2 remained in sustained remission for > 4 years. To study longitudinal clonal dynamics through hematopoietic differentiation, we performed high-sensitivity single-cell genotyping in index-sorted sequential patient samples. In all patients who relapsed, therapy-resistant clones were selected rapidly, within 1-3 treatment cycles, at times when hematopoiesis was still largely sustained by either normal or pre-leukemic cells. Selection of therapy-resistant clones preceded overt treatment failure by months or even years. Relapse was associated either with clones harboring newly-detected myeloid driver mutations or expansion of minor pre-existing clones that had reduced fitness prior to treatment. In both cases, resistant clones were selected within immature cell populations previously shown to contain leukemic stem cell (LSC) potential, preceding malignant expansion of these compartments by immunophenotyping. In contrast, in both patients remaining in remission, leukemic clones were eradicated and rapidly replaced by clonal and wild-type hematopoiesis. These observations suggest that, in patients treated with non-intensive ivosidenib combination therapy, rapid clonal selection occurs in populations with LSC potential, where failure to eliminate either genetically evolved or persistent leukemic clones ultimately leads to relapse. Key points- Rapid selection of leukemic clones occurs within small populations with LSC potential, months or years prior to relapse. - Rapid eradication of leukemic clones leads to sustained remission in the context of ivosidenib combination therapy.

cancer biology↗

bioRxiv2024

Identification of novel wraparound transcripts in JC polyomavirus

JC polyomavirus (JCPyV) is a ubiquitous pathogen causing progressive multifocal leukoencephalopathy (PML) in immunocompromised individuals. Polyomaviruses (PyVs) utilize complex transcriptional regulation mechanisms to express diverse mRNAs from their compact, double-stranded circular DNA genomes. A recent study employed next-generation sequencing (NGS) to provide a detailed transcriptome atlas of PyVs, including BK polyomavirus and simian virus 40. However, detailed information regarding the JCPyV transcriptome remains limited. Here, we conducted a comprehensive analysis combining short-read and long-read NGS methods to unveil the detailed transcriptome atlas of JCPyV. RNA isolated from IMR-32 and 293 cells transfected with the circular JCPyV genome was analyzed by NGS, leading to the discovery of 39 novel viral transcripts in addition to 12 previously annotated ones. Among these novel transcripts, we identified characteristic wraparound transcripts, conserved across PyVs, which are constructed from long primary transcripts generated through continuous, multiple rounds of circular transcription of the viral genome. These wraparound transcripts included both late transcripts harboring leader-to-leader repeated sequences and SuperT transcripts containing multiple LxCxE motifs. Notably, wraparound transcripts, including a SuperT transcript, were also detected in brain tissues from PML patients. Collectively, this study significantly expands our understanding of the JCPyV transcriptome, revealing the expression of wraparound transcripts in PML lesions. These findings provide valuable insights into the molecular basis of JCPyV gene expression and PML pathogenesis, potentially facilitating the development of effective countermeasures against PML. Author SummaryJC polyomavirus (JCPyV) was first identified in 1971 from the brain tissue of a patient with progressive multifocal leukoencephalopathy (PML), a life-threatening neurological disease. Despite over 50 years since the pathogens identification, the prognosis for PML remains extremely poor due to the lack of effective therapeutics. Elucidating the JCPyV transcriptome can expand our understanding of viral gene expression and PML pathogenesis, potentially providing targets for novel therapeutic strategies. Here, we employed a combination of three next-generation sequencing technologies to explore novel viral transcripts, creating a detailed transcriptome atlas of JCPyV. We identified 39 novel viral transcripts in addition to the 12 previously annotated ones in cultured cells transfected with the JCPyV genome. These novel transcripts included wraparound transcripts, which arise from continuous, multiple rounds of circular transcription of the viral genome. Wraparound transcripts were characterized by the presence of tandem repeats of approximately 100 nucleotides. Notably, these characteristic wraparound transcripts were also detected in brain tissues from PML patients. Our study provides new insights into JCPyV gene expression, potentially contributing to the development of effective therapeutics.

microbiology↗

bioRxiv2024

Orthogonal spectral and temporal envelope representation in auditory cortex

Speech perception relies on two fundamental acoustic components: spectral and temporal. While spectral information is known to be represented in the auditory cortex through tonotopy, how temporal features are organized has remained unclear. Here, by varying rise-ramp steepness and frequencies, we reveal that the steepness of the temporal envelope--a critical cue for phonemes discrimination and sound source perception--is systematically mapped in the mouse auditory cortex. Using widefield calcium imaging, we discovered that the envelope steepness is represented orthogonally to the tonotopic axis, forming a two-dimensional cortical map that mirrors the dual structure of sounds. This organization was observed in primary-like auditory regions but not in higher-order-like areas, indicating distinct auditory processing streams. These findings uncover a principle of cortical organization, suggesting that the auditory cortex encodes sound along two independent axes and thereby provides a neural basis for parallel processing for complex sounds such as speech and natural acoustic environments.

neuroscience↗

bioRxiv2024

Replication-dependent histone (Repli-Histo) labeling dissects the physical properties of euchromatin/heterochromatin in living human cells.

A string of nucleosomes, where genomic DNA is wrapped around histones, is organized in the cell as chromatin. Chromatin in the cell varies greatly, from euchromatin to heterochromatin, in its genome functions. It is important to understand how heterochromatin is physically different from euchromatin. However, their specific labeling methods in living cells are limited. To address this, we have developed replication-dependent histone labeling (Repli-Histo labeling) to label nucleosomes in euchromatin and heterochromatin based on DNA replication timing. We investigated local nucleosome motion in the four chromatin classes from euchromatin to heterochromatin of living human and mouse cells. We found that more euchromatic regions (earlier replicated regions) show greater nucleosome motion. Notably, the motion profile in each chromatin class persists throughout interphase. Genome chromatin is essentially replicated from regions with greater nucleosome motions, even though the replication timing program is perturbed. Our findings, combined with computational modeling, suggest that earlier replicated regions have more accessibility and local chromatin motion can be a major determinant of genome-wide replication timing.

cell biology↗

bioRxiv2024

Plasma membrane H+-ATPase activation increases global transcript levels and promotes the shoot growth of light-grown Arabidopsis seedlings.

(1) Plant cell growth require the elongation of cells mediated by cell wall remodelling and turgor pressure changes. The plasma membrane (PM) H+-ATPase facilitates both cell wall remodelling and turgor pressure changes, by acidifying the apoplast of cells, referred to as acid growth. The acid growth theory is mostly established on the auxin-induced activation of PM H+-ATPase in non-photosynthetic tissues. However, how PM H+-ATPase affect the growth in photosynthetic tissues of Arabidopsis remains unclear. (2) Here, a combination of transcriptomics and cis-regulatory element analysis was conducted to identify the impact of PM H+-ATPase on global transcript levels and the molecular mechanism downstream of the PM H+-ATPase. (3) The PM H+-ATPase activation increased transcript levels globally, especially cell wall modification-related genes. The transcript level changes were in PM H+-ATPase-dependent manner. Involvement of Ca2+ was suggested as CAMTA motif was enriched in the promoter of PM H+-ATPase-induced genes and cytosolic Ca2+ elevated upon PM H+-ATPase activation. (4) PM H+-ATPase activation in photosynthetic tissues promote the expression of cell wall modification enzymes and shoot growth, adding a novel perspective of photosynthesis-dependent PM H+-ATPase activation in photosynthetic tissues to the acid growth theory that has primarily based on findings from non-photosynthetic tissues.

plant biology↗