Biology subjects

Yagi, S.

Publications and source records attributed to Yagi, S..

13 recordsLinked to original sources

bioRxiv2026

Data-driven spectroscopic dictionaries and detector-calibrated inference for photon-limited Raman hyperspectral imaging of living cells

Label-free Raman imaging of living cells is photon limited: at exposures compatible with cellular dynamics, single-pixel spectra carry about one count per channel on a dominant smooth background. We present an unmixing framework in which the decoder of a physics-constrained autoencoder is restricted to a data-driven spectroscopic dictionary: band centers,widths, and pseudo-Voigt shapes are measured from the dataset and fixed, and the network learns only nonnegative band amplitudes, a smooth B-spline background, and a per-pixel gain.First, on slit-scanning images of HeLa cells (532 nm) the dictionary yields spike-free component spectra that read as band tables, including a resonance-enhanced cytochrome-c-associated component matching literature spectra, and the most stable decomposition against the component number. Second, the dictionary and initialization calibrated at 1 s exposure perline transfer to 100 ms per line (12 s sweeps): cytochrome-c spectral identity survives a single sweep (correlation 0.92) while its map remains photon limited; the dictionary provides spectral physicality, and the transferred initialization prevents a structural collapse that global map correlations miss; in a measurement-derived phantom the dictionary estimator holds thecytochrome-c spectrum to 17-19{degrees} spectral angle at 100 ms, where classical factorizations and free decoders lose it (55-64{degrees}). Estimation on the count-equivalent detector output uses a calibrated shifted-Poisson quasi-likelihood. Third, evaluation must be time matched:correlation against a separately acquired reference saturates through slow specimen drift and acquisition mismatch rather than photon noise, and the self-consistency of learned denoisers is inflated by shared bias; time-matched self-consistency and independent cross-checks areproposed.

cell biology↗

bioRxiv2026

Prebiotic cationic amino acids support formation of an ancient protein fold

The evolutionary basis for the selection of the standard proteinogenic amino acids remains elusive, particularly for the long cationic amino acids, lysine and arginine, which were likely scarce in the prebiotic environment. In contrast, shorter cationic amino acids, such as ornithine (Orn), and 2,4-diaminobutyric acid (Dab), are thought to have been more abundant. Here, we investigated whether these prebiotic cationic amino acids can support protein tertiary structure formation, using computational protein design, biophysical and crystallographic analyses, and molecular dynamics (MD) simulations. We designed sequences for an ancient protein fold, the double-{Psi} {beta}-barrel (DPBB), with ornithine and plausible prebiotic amino acid sets. Although all the designed sequences were unfolded under standard dilute aqueous conditions, one Orn-containing variant folded in highly concentrated conditions. Crystallographic analysis revealed that both this peptide and its Dab-substituted derivative adopted the double-Z {beta}-barrel (DZBB) fold, a likely evolutionary intermediate between extant {beta}-barrel folds. Therefore, Orn and Dab might have supported the foldability of primitive proteins before the incorporation of lysine and arginine into the genetic code.

biochemistry↗

bioRxiv2026

Long-term single-particle tracking by NIR imaging using Au42 (gold) quantum needles

Single-particle tracking (SPT) over time enables direct observation of molecular transport and interactions in living cells. Fluorescence-based SPT has provided insights into intracellular processes such as endocytosis, receptor signaling, and drug delivery. Extending the observation window to several hours and beyond is critical for capturing slow intracellular dynamics, including the full course of endosomal trafficking, the long-term accumulation of particles within subcellular compartments, and transitions between transport modes that occur on hour-scale timescales. However, long-term intracellular SPT under visible-wavelength excitation remains challenging because fluorescence probes generally suffer from photobleaching and phototoxicity. While near-infrared (NIR) excitation can simultaneously mitigate these issues, generally weak emission of NIR-emitting dyes has hindered its wide application in long-term SPT. Here, we demonstrate long-term NIR SPT using atomically precise gold quantum needles, Au42(PET)32 (PET = 2-phenylethanethiolate). Continuous tracking of intracellular particles in living HEK293 cells was achieved for up to 12 h. Trajectory analysis revealed temporal transitions between directional and diffusive transport, as well as the accumulation of multiple particles within localized intracellular domains over several-hour timescales. The high photostability of Au42, combined with low phototoxicity of NIR excitation, enables visualization of intracellular transport dynamics over timescales difficult to access using conventional visible fluorescent probes. These results establish Au42-based NIR imaging as a platform for long-term, low-phototoxicity intracellular SPT and provide a framework for investigating slow intracellular dynamics in living systems.

biophysics↗

bioRxiv2026

Emergence of a potentially ancestral ATP-synthesizing activity under prebiotic amino acid constraints

How amino acid composition shapes enzymatic function remains a central question in molecular evolution. Early proteins were likely composed of a limited set of amino acids, yet the catalytic properties of proteins under such compositional constraints are still poorly understood. Here we show that restricting amino acid repertoires can replace native enzymatic activity with an alternative phosphoryl-transfer reaction that disproportionates ADP into ATP and AMP. Using a reconstructed ancestral nucleoside diphosphate kinase (NDK) variant composed of a restricted amino acid set, we demonstrate this reaction, which is not observed in extant NDKs but is chemically analogous to that catalyzed by adenylate kinase. Although modest in catalytic rate, X-ray crystallography, molecular dynamics, and mutational analyses reveal a distinct active-site organization in which aspartate and arginine residues cooperatively coordinate Mg2+ and support phosphoryl transfer through a noncanonical mechanism. Lysine can substitute for arginine under these constraints while retaining activity. Together, these findings show that restricting amino acid diversity can remodel active sites and promote alternative phosphoryl-transfer reactions, illustrating how limitations in amino acid availability could influence catalytic functions in early enzyme evolution. Significant StatementThe first enzymes arose on early Earth, where only a limited set of amino acids was likely available. Whether such limitations reduce catalytic efficiency or reshape enzyme function is still unclear. We reconstructed a model of an ancestral enzyme using a simplified amino acid set approximating those available on early Earth and found that restricting amino acid diversity replaced its original catalytic activity with an alternative function that produces ATP from two ADP molecules. Structural and mutational analyses indicate that this shift involves a reorganization of the active site. These results provide experimental evidence that limiting amino acid diversity can reshape catalytic function and may have influenced the emergence of catalytic activities during enzyme evolution.

evolutionary biology↗

bioRxiv2023

Missing-link fold reveals the evolutionary pathway between RNA polymerase and ribosomal proteins

Numerous molecular machines are required to drive the central dogma of molecular biology. However, the means by which such numerous proteins emerged in the early evolutionary stage of life remains enigmatic. Many of them possess small {beta}-barrel folds with different topologies, represented by DPBB conserved in DNA and RNA polymerases, and RIFT, OB, and SH3 in ribosomal proteins. Here, we discovered that the previously reconstructed ancient DPBB sequence could also adopt a novel {beta}-barrel fold named DZBB, which shares similarities with RIFT and OB. Indeed, DZBB could be transformed into them through simple engineering experiments. Furthermore, the OB designs could be converted into SH3 by circular-permutation. These results indicate that these {beta}-barrels diversified quickly from a common ancestor at the beginning of the central dogma evolution.

biochemistry↗

bioRxiv2022

High Estradiol reduces adult neurogenesis but strengthens functional connectivity within the hippocampus during spatial pattern separation in adult female rats

Adult neurogenesis in the dentate gyrus plays an important role for pattern separation, the process of separating similar inputs and forming distinct neural representations. Estradiol modulates neurogenesis and hippocampus function, but to date no examination of estradiols effects on pattern separation have been conducted. Here, we examined estrogenic regulation of adult neurogenesis and functional connectivity in the hippocampus after the spatial pattern separation task in female rats. Ovariectomized Sprague-Dawley rats received daily injections of vehicle, 0.32{micro}g (Low) or 5 {micro}g (High) of estradiol benzoate until the end of experiment. A single bromodeoxyuridine (BrdU) was injected one day after initiation of hormone or vehicle treatment and rats were tested in the delayed nonmatching to position spatial pattern separation task in the 8-arm radial maze for 14 days beginning two weeks after BrdU injection. Rats were perfused 90 minutes after the final trial and brain sections were immunohistochemically stained for BrdU/neuronal nuclei (NeuN) (new neurons), Ki67 (cell proliferation), and the immediate early gene, zif268 (activation). Results showed that only high estradiol reduced the density of BrdU/NeuN-ir cells and had significant inter-regional correlations of zif268-ir cell density in the hippocampus following pattern separation. Estradiol treatment did not influence pattern separation performance or strategy use. These results show that higher doses of estradiol can reduce neurogenesis but at the same time increases correlations of activity of neurons within the hippocampus during spatial pattern separation.

neuroscience↗

bioRxiv2022

Estrogens dynamically regulate neurogenesis in the dentate gyrus of adult female rats

Estrone and estradiol differentially modulate neuroplasticity and cognition, but how they influence maturation pathways of new neurons in the adult hippocampus is not known. The present study assessed the effects of estrone and estradiol on various aspects of neurogenesis in the dentate gyrus (DG) of ovariectomized young adult Sprague-Dawley rats using daily subcutaneous injections of 17{beta}-estradiol or estrone. Rats were injected with a DNA synthesis marker, 5-bromo-2-deoxyuridine (BrdU), and were perfused one, two, or three weeks after BrdU injection and treatment. Immunofluorescent labelling for GFAP/Sox2 was used to examine the density of neural stem/progenitor cells and Ki67 for cell proliferation. Double-immunofluorescent labelling of BrdU with doublecortin (DCX) or NeuN was used to examine the attrition and maturation of adult-born neurons over time. Length of time post ovariectomy was associated with a reduction in neural stem/progenitor cells in the DG. Estrogens had effects on different components of neurogenesis after one week of exposure but not after prolonged treatment. Estradiol enhanced, whereas estrone reduced, cell proliferation after one week but not after longer exposure to hormones. Both estrogens increased the density of BrdU/DCX-ir cells after one week of exposure but showed greater attrition of new neurons between one and two weeks after exposure, suggesting that the effects of estrogens on neurogenesis were not sustained. These results demonstrate that estrogens modulate several aspects of adult hippocampal neurogenesis differently in the short term, but may lose their ability to influence neurogenesis after long-term exposure.

neuroscience↗

bioRxiv2022

Sex differences in contextual pattern separation, activation of new neurons, and functional connectivity within the limbic system

Sex differences in the structure and function of hippocampus exist. Here, we examined sex differences in contextual pattern separation, functional connectivity, and activation of new neurons during fear memory. Two-month-old male and female Sprague-Dawley rats were injected with the DNA synthesis markers, iododeoxyuridine (IdU) and chlorodeoxyuridine (CldU) three weeks and four weeks before perfusion, respectively. One week after CldU injection, the rats underwent a context discrimination task in which rats were placed in context A (shock) and context A (no shock) every day for 12 days. On the test day, rats were placed in the shock context (context A) to measure fear memory and expression of zif268, an immediate early gene across 15 different limbic and reward regions. We found that females, but not males, showed contextual discrimination during the last days of training. On the test day, both sexes displayed similar levels of freezing, indicating equivalent fear memory for context A. Despite similar fear memory, males showed more positive correlations of zif268 activation between the limbic regions and the striatum, whereas females showed more negative correlations among these regions. Females showed greater activation of the frontal cortex, dorsal CA1, and 3-week-old adult-born dentate granular cells compared to males. These results highlight the importance of studying sex differences in fear memory and the contribution of adult neurogenesis to the neuronal network and may contribute to differences in susceptibility to fear related disorders such as posttraumatic stress disorder.

neuroscience↗

bioRxiv2021

Seven amino acid types suffice to reconstruct the core fold of RNA polymerase

AbstractThe extant complex proteins must have evolved from ancient short and simple ancestors. Nevertheless, how such prototype proteins emerged on the primitive earth remains enigmatic. The double-psi beta-barrel (DPBB) is one of the oldest protein folds and conserved in various fundamental enzymes, such as the core domain of RNA polymerase. Here, by reverse engineering a modern DPBB domain, we reconstructed its evolutionary pathway started by "interlacing homo- dimerization" of a half-size peptide, followed by gene duplication and fusion. Furthermore, by simplifying the amino acid repertoire of the peptide, we successfully created the DPBB fold with only seven amino acid types (Ala, Asp, Glu, Gly, Lys, Arg, and Val), which can be coded by only GNN and ARR (R = A or G) codons in the modern translation system. Thus, the DPBB fold could have been materialized by the early translation system and genetic code.

biochemistry↗

bioRxiv2020

Sex Differences in Risk-based Decision Making

Decision-making is a complex process essential to daily adaptation in many species. Risk is an inherent aspect of decision-making and it is influenced by gonadal hormones. Testosterone and 17{beta}-estradiol may modulate decision making and impact the mesocorticolimbic dopamine pathway. Here, we explored sex differences, the effect of gonadal hormones and the dopamine agonist amphetamine on risk-based decision making. Intact or gonadectomised (GDX) male and female rats underwent to a probabilistic discounting task. High and low doses of testosterone propionate (1.0 or 0.2 mg) and 17{beta}-estradiol benzoate (0.3 g) were administered to assess acute effects on risk-based decision making. After 3-days of washout period, intact and GDX rats received high or low (0.5 or 0.125 mg/kg) doses of amphetamine and re-tested in the probabilistic discounting task. Under baseline conditions, males made more risky choices during probability discounting compared to female rats, particularly in the lower probability blocks, but GDX did not influence risky choice. The high, but not the low dose, of testosterone modestly reduced risky decision making in GDX male rats. Conversely, 17{beta}-estradiol had no significant effect on risky choice regardless of GDX status in either sex. Lastly, a higher dose of amphetamine increased risky decision making in both intact males and females, but had no effect in GDX rats. These findings demonstrated sex differences in risk-based decision making, with males showing a stronger bias towards larger, uncertain rewards. GDX status influenced the effects of amphetamine, suggesting different dopaminergic regulation in risk-based choices among males and females.

neuroscience↗

bioRxiv2019

Sex differences in maturation and attrition rate of adult born neurons in the hippocampus of rats

Sex differences exist in the regulation of adult neurogenesis in the hippocampus in response to hormones and cognitive training. Here we investigated the trajectory and maturation rate of adult-born neurons in the dentate gyrus (DG) of male and female rats. Sprague-Dawley rats were perfused two hours, 24 hours, one, two or three weeks after BrdU injection, a DNA synthesis marker that labels dividing progenitor cells and their progeny. Adult-born neurons (BrdU/NeuN-ir) matured faster in males compared to females. Males had a greater density of neural stem cells (Sox2-ir) in the dorsal, but not in the ventral, DG and had higher levels of cell proliferation (Ki67-ir) than non-proestrous females. However, males showed a greater reduction in neurogenesis between one and two weeks after mitosis, whereas females showed similar levels of neurogenesis throughout the weeks. The faster maturation and greater attrition of new neurons in males compared to females suggests greater potential for neurogenesis to respond to external stimuli in males and emphasizes the importance of studying sex on adult hippocampal neurogenesis. Significance StatementPreviously studies examining the characteristics of adult-born neurons in the dentate gyrus have used almost exclusively male subjects. Researchers have assumed the two sexes have a similar maturation and attrition of new neurons in the dentate gyrus of adults. However, this study highlights notable sex differences in the attrition, maturation rate and potential of neurogenesis in the adult hippocampus that has significant implications for the field of neuroplasticity. These findings are important in understanding the relevance of sex differences in the regulation of neurogenesis in the hippocampus in response to stimuli or experience and may have consequences for our understanding of diseases that involve neurodegeneration of the hippocampus, particularly those that involve sex differences, such as Alzheimers disease and depression.

neuroscience↗

bioRxiv2018

Experimental Zika Virus Infection in the Pregnant Common Marmoset Induces Spontaneous Fetal Loss and Neurodevelopmental Abnormalities

During its most recent outbreak across the Americas, Zika virus (ZIKV) was surprisingly shown to cause fetal loss and congenital malformations in acutely and chronically infected pregnant women. However, understanding the underlying pathogenesis of ZIKV congenital disease has been hampered by a lack of relevant in vivo experimental models. Here we present a candidate New World monkey model of ZIKV infection in pregnant marmosets that faithfully recapitulates human disease. ZIKV inoculation at the human-equivalent of early gestation caused an asymptomatic seroconversion, induction of type I/II interferon-associated genes and proinflammatory cytokines, and persistent viremia and viruria. Spontaneous pregnancy loss was observed 16-18 days post-infection, with extensive active placental viral replication and fetal neurocellular disorganization similar to that seen in humans. These findings underscore the key role of the placenta as a conduit for fetal infection, and demonstrate the utility of marmosets as a highly relevant model for studying congenital ZIKV disease and pregnancy loss.

microbiology↗

bioRxiv2017

Experimental Zika Virus Infection in a New World Monkey Model Reproduces Key Features of the Human Disease

Human infections by Zika virus (ZIKV), a mosquito-borne flavivirus, are associated with a current widespread outbreak in the Americas, and have been associated with neurological complications and adverse fetal outcomes such as microcephaly in pregnant women. A suitable non-human primate model is urgently needed. To evaluate ZIKV infectivity, pathogenesis, and persistence, we inoculated 4 marmosets with ZIKV and followed them by clinical monitoring and serial sampling of body fluids for up to 11 weeks. We found that marmosets experimentally infected with ZIKV reproduced key features of the human disease, including (1) asymptomatic infection, (2) brief period of detectable virus in serum (<1 week), (3) detection in other body fluids (urine, saliva, semen, and stool) for at least 2 weeks following acute infection, and (4) persistence in lymph nodes, but not other tissues, at 1 month post-infection. ZIKV-positive saliva and serum samples, but not urine, were found to be infectious in cell culture. By day 6 post-inoculation, most marmosets exhibited detectable neutralizing antibody responses concurrent with activation of NK cell and B cell subsets and an increase in circulating cytokines associated with type II interferon signaling, Transcriptome profiling revealed enrichment of immune responses to active viral infection, with up-regulation of both type I and II interferon signaling pathways, anduncovered potential host biomarkers. These results suggest that a New World monkey model of acute ZIKV infection mimics the human disease, and is likely to be useful for testing of drug and vaccine candidates.

microbiology↗