Biology subjects

Ueda, M.

Publications and source records attributed to Ueda, M..

At least 19 recordsLinked to original sources

bioRxiv2026

MpILR1 Hydrolyzes Jasmonate-Amino Acid Conjugates to Activate dn-iso-OPDA Signaling in Marchantia polymorpha.

Jasmonates are essential phytohormones that coordinate defense responses and developmental programs across land plants. In angiosperms, the active jasmonate ligand jasmonoyl-L-isoleucine (JA-Ile), is produced through GH3-mediated conjugation of jasmonic acid to isoleucine and JA-Ile homeostasis is further shaped by ILR1/ILL-family amidohydrolases. In contrast, the primary bioactive jasmonate ligand in bryophytes, dinor-12-oxo-phytodienoic acid (dn-iso-OPDA), is inactivated through conjugation with amino acids, raising the question of whether these conjugates constitute a reversible hormone reservoir or an irreversible catabolic end point. Although the ILR1-like family has been characterized extensively for its role in auxin and jasmonate homeostasis in angiosperms, its function in bryophytes remains basically unexplored. Here we show that MpILR1, the sole Marchantia ortholog of the ILR1/ILL family, hydrolyzes a specific subset of dn-iso-OPDA-amino acid conjugates in vivo. Loss-of-function Mpilr1 mutants exhibit enhanced accumulation of dn-iso-OPDA conjugated to hydrophobic amino acids (Val, Leu and Ile) but not to hydrophilic residues (His, Glu and Gln), demonstrating substrate-selective hydrolysis. MpILR1 hydrolytic activity is required for full dn-iso-OPDA-mediated responses, including transcriptional activation and defense against gastropod herbivory. These findings establish MpILR1 as a key positive regulator of jasmonate signaling in Marchantia polymorpha and suggest that hormone conjugation/deconjugation is an ancient regulatory mechanism evolved during plant terrestrialization.

plant biology↗

bioRxiv2026

Apical3DTip: Elliptic Cross-section-based Reconstruction for the Embryo Initial Cell of Arabidopsis

BackgroundCell geometry plays a central role in determining division orientation and body axis formation during early embryogenesis in Arabidopsis thaliana. However, quantitative analysis of dynamic three-dimensional (3D) morphology remains challenging because live-imaging studies often rely on two-dimensional (2D) projections, while existing 3D reconstruction approaches, including mesh-based methods, often lose the original orientation information relative to the ovule and require labor-intensive mesh correction. In addition, embryo positional fluctuation caused by floating in liquid medium and continuous growth makes it difficult to analyze temporal morphological changes within a common coordinate system. ResultsWe developed a robust framework for quantitative 3D and four-dimensional (4D; 3D + time) analysis of embryo initial cell (apical cell) morphology. The method first establishes a standardized 3D coordinate system by normalizing cell orientation based on the bottom plane and the optical axis of the observation. Cell morphology is then reconstructed through ellipse-based approximation of serial cross-sections extracted from stacked imaging data, enabling accurate geometric characterization without the need for complex surface mesh reconstruction. To evaluate shape anisotropy, we quantified the apical cell shape in 3D. The framework further supports the characterization of volumetric features of subsequent division, providing a basis for quantifying 3D embryogenesis. ConclusionOur framework provides a simple and noise-reduced approach for quantitative analysis of living cell morphology in 3D. We named the integrated method of combining coordinate normalization with elliptical cross-section-based reconstruction Apical3DTip. This method enables consistent comparison of cell shapes without extensive manual corrections. The method overcomes key limitations of 2D projection-based and mesh-dependent analyses and offers a practical platform for quantifying cell shape and daughter cell shapes in 3D. More broadly, it provides a quantitative foundation for exploring the relationship between cell geometry, morphodynamics, and developmental patterning in living plant embryos.

plant biology↗

bioRxiv2026

Modular mRNA platform for in vivo antigen-specific immune tolerance

Immune-mediated diseases such as autoimmunity and allergy arise from dysregulated antigen-specific responses, yet current therapies largely rely on broad immunosuppression and rarely re-establish durable, antigen-selective tolerance. Here we introduce Tol-mRNA, a modular tolerogenic mRNA platform designed to deliver multiple immune-regulatory signals in vivo to program antigen-specific tolerance. Tol-mRNA co-encodes an antigen together with two complementary regulatory cues, PD-L1 and TGF-{beta}, enabling coordinated antigen presentation and tolerogenic signaling that robustly induces antigen-specific Tregs and suppresses pathogenic effector responses. In murine disease models, Tol-mRNA conferred therapeutic benefit across distinct immune pathologies. In experimental autoimmune encephalomyelitis, Tol-mRNA ameliorated clinical disease and reduced inflammatory immune activation. In an ovalbumin-driven food allergy model, Tol-mRNA prevented allergic symptoms and dampened type 2 inflammation. Extending these findings to humans, Tol-mRNA demonstrated potent, antigen-dependent immunosuppressive activity and a strong capacity to promote antigen-specific Treg induction in human immune settings. Collectively, these results establish Tol-mRNA as a scalable and versatile approach to reprogram antigen-specific immunity and support its development as a next-generation therapeutic modality for immune-mediated diseases.

immunology↗

bioRxiv2026

FUT8 mediated core fucosylation modulates growth related functions of LRP1 in liver cancer cells

Fucosylation is a cancer-associated glycosylation change, and core fucosylation of N-glycans catalyzed by the 1,6-fucosyltransferase FUT8, has been closely linked to tumor progression, metastasis, drug resistance, and poor prognosis. However, the core-fucosylated proteins that directly support hepatocarcinogenesis are not fully defined. In a KRAS-G12D-driven mouse liver cancer model, we observed increased fucosylation and FUT8 upregulation, and glycoproteomic analysis of fucose-enriched fractions identified low-density lipoprotein receptor-related protein 1 (LRP1) as a prominent core-fucosylated protein in tumor tissue. In immortalized mouse hepatocytes, genetic or pharmacological inhibition of FUT8 markedly increased Lrp1 mRNA and protein, indicating that loss of core fucosylation is accompanied by robust upregulation of LRP1. In human HepG2 cells, LRP1 knockout suppressed cell proliferation and markedly altered colony morphology, leading to compact rounded clusters instead of the typical polygonal pattern. It also reduced EGFR protein and further inhibited proliferation of HepG2 cell. These findings identify LRP1 as a FUT8-dependent core-fucosylated receptor in experimental hepatocarcinogenesis and suggest that the FUT8-LRP1 axis contributes to the maintenance of proliferative signaling in hepatoma cells.

cancer biology↗

bioRxiv2026

Ca2+ oscillations promote microtubule-band turnover and support tip growth in Arabidopsis zygotes

The zygote is the origin of development, and in most angiosperms, it divides asymmetrically to establish the apical-basal axis. In Arabidopsis thaliana, the zygote undergoes tip growth-like polar elongation, using a subapical transverse microtubule band (MT band). Because canonical tip-growing cells rely on longitudinal actin filaments (F-actin), it remains unclear whether and how the zygote employs conserved tip-growth mechanisms. Here, using quantitative live-cell imaging, pharmacological perturbations, and mechanical simulations, we found that oscillatory Ca{superscript 2} waves, a hallmark of tip growth, are coupled to zygote elongation through a bidirectional feedback loop, as in other tip-growing cells. However, Ca{superscript 2} waves were dispensable for overall F-actin alignment but promoted MT band turnover. Our study provides a model showing that the zygote uses a conserved tip-growth module, in which Ca{superscript 2} oscillations and cell elongation reinforce each other, but redirects the target cytoskeleton to the MT band, enabling zygote-specific tip growth for axis formation.

plant biology↗

bioRxiv2026

Two-step polar plastid migration via F-actin and microtubules ensures unequal inheritance during asymmetric division of Arabidopsis zygote

The zygote is the origin of development, and in most angiosperms, it divides asymmetrically to establish the apical-basal axis. In Arabidopsis thaliana, various organelles in the zygote undergo polar migration along actin filaments (F-actin), resulting in unequal inheritance, but the behavior of plastids, essential precursors of chloroplasts, has remained unclear. Here, using quantitative live-cell imaging, we reveal that plastids undergo two-step polar migration: they first move apically together with the nucleus along F-actin, and when nuclear migration slows, they switch to microtubule (MT)-dependent migration to move further apically. This results in unequal plastid inheritance by the apical cell. Although these plastids are amyloplasts containing starch granules, starch is dispensable for migration, unlike the gravity response. Instead, a fertilization-activated MAP kinase pathway is required for polar plastid migration. Our results demonstrate that the zygote possesses a spatiotemporal regulatory mechanism that ensures unequal plastid inheritance at the onset of plant ontogeny.

plant biology↗

bioRxiv2025

HDA19-mediated deacetylation of histone H3.3 lysine 27 and 36 regulates plant sensitivity to salt stress

Plants survive extreme environments through rapid chromatin reprogramming, yet the epigenetic marks that confer stress resilience remain poorly understood. Histone deacetylase HDA19 is a key epigenetic regulator in Arabidopsis, and hda19-deficient mutants display tolerance to multiple abiotic stresses, including drought, heat, and salinity. Using lysine acetylome profiling, we identified a non-canonical K27/K36 di-acetylation mark on histone H3.3, among nine H3 variants, as a specific substrate of HDA19. Under salinity stress, this mark decreased in wild-type plants but increased in hda19 mutants, while other known H3 modifications were similarly affected in both genotypes. Mimicking constitutive di-acetylation of H3.3 K27/K36 through lysine-to-glutamine substitutions promoted accumulation of stress-responsive late embryogenesis abundant (LEA) proteins and conferred salinity tolerance in seedlings, phenocopying hda19 mutants. Furthermore, generating the lea7-1/lea29-1/rab18-1 triple mutant abolished hda19-dependent salinity tolerance, confirming the LEA proteins role downstream of HDA19. Our findings demonstrate that H3.3 K27/K36 di-acetylation, modulated by HDA19, drives LEA protein accumulation and enables plants to withstand environmental stress, revealing a previously unknown core mechanism of plant stress resilience. Significance statementHistone acetylation is crucial for regulating chromatin states and transcription, yet the specific histone codes controlled by histone deacetylases and acetyltransferases during plant stress acclimation remain unclear. Arabidopsis mutants lacking histone deacetylase 19 show enhanced tolerance to drought, heat, and salinity. Using acetylome profiling and genetic analyses, we identify a previously unknown di-acetylation of histone H3.3 at lysines 27 and 36 as a key epigenetic switch activating late embryogenesis abundant proteins essential for salt tolerance in seedlings. This discovery reveals a novel mechanism linking histone H3.3 K27/K36 di-acetylation to stress-resilient protein accumulation, offering new insights into epigenetic control of plant stress responses and informing strategies to improve crop resilience under climate change.

plant biology↗

bioRxiv2025

Transient let-7/miR-98 reprograms primed pluripotency to induce naivehuman PSCs

Naive human pluripotent stem cells (hPSCs) are valuable for modeling early development and enabling regenerative applications, yet resetting from the primed state remains inefficient and mechanistically unclear. We identify an unexpected role for miR-98-5p, a let-7 family microRNA (miRNA) typically linked to differentiation, in promoting naive induction. Using a synthetic mRNA screen containing diverse miRNA-responsive elements, we uncovered miR-98-5p as a potent enhancer of resetting. Single-cell RNA sequencing and targeted perturbations show that miR-98-5p directly represses NR6A1 while transiently activating BMP2/4 signaling, triggering network transitions. These changes destabilize the primed gene-regulatory network and enable acquisition of a naive state. Leveraging this mechanism, we establish a feeder-free, miRNA-based, editing-free protocol that boosts resetting efficiency from <10% to [~]80%. To our knowledge, this is the first demonstration that a let-7 miRNA can drive naive induction by coupling direct target repression with activation of pro-naive signaling, suggesting a generalizable reprogramming principle of transient network destabilization.

developmental biology↗

bioRxiv2025

(3R, 7S)-11-hydroxy-jasmonic acid is a major oxidative shunt product of jasmonate catabolism in Arabidopsis thaliana

Jasmonoyl-L-isoleucine (JA-Ile) is a pivotal oxylipin plant hormone that regulates numerous physiological processes and stress responses. Because JA-Ile is synthesized from jasmonic acid (JA), the cellular level of JA is tightly controlled. Although the oxidative catabolism of JA to 12-hydroxy-JA (12-OH-JA) is well-characterized, the biosynthesis, accumulation dynamics, and biological functions of 11-hydroxy-JA (11-OH-JA) remain poorly understood. Here, we conducted a comprehensive investigation of 11-OH-JA in Arabidopsis thaliana, encompassing its chemical synthesis, accumulation patterns, biological activity, and biosynthetic pathways. Using chemically synthesized 11-OH-JA of naturally occurring (3R,7S)-stereochemistry, we identified 11-OH-JA as the predominant hydroxylated JA derivative that accumulates following wounding. Enzymatic assays and in silico docking studies revealed that JOX1/2/3/4 exclusively catalyze the conversion of JA to 11-OH-JA, whereas 12-OH-JA is produced by distinct pathways. Notably, the quadruple mutant joxQ completely abolished 11-OH-JA accumulation without affecting 12-OH-JA levels. Furthermore, 11-OH-JA exhibited no binding affinity for the COI1-JAZ co-receptor, indicating that it acts as a biologically inactive shunt product of JA catabolism. Our findings establish JOX-mediated 11-hydroxylation of JA as a primary inactivation pathway, separate from 12-hydroxylation. Parallel, non-redundant inactivation routes deliver layered signal termination of costly defense signals. This study provides new insights into JA catabolism, highlighting 11-OH-JA as a crucial factor in the attenuation of jasmonate signaling, and thereby redefining the framework of JA turnover in A. thaliana.

plant biology↗

bioRxiv2025

mRNA-Based Designer APCs Elicit Robust CD8⁺ and CD4⁺ T Cell Responses

mRNA-based therapeutics have demonstrated notable success in SARS-CoV-2 vaccines and are emerging in cancer immunotherapy. However, conventional mRNA cancer vaccines are limited by the low immunogenicity of tumor-associated and neoantigens. We addressed this limitation by formulating a modular, liposome-based mRNA cocktail comprising three distinct mRNAs encoding tumor antigen, the co-stimulatory molecule CD80, and membrane-tethered IL-2. Administration of this mRNA mixture transforms somatic cells into designer antigen-presenting cells (APCs) in vivo, which simultaneously express the antigen, a co-stimulatory molecule, and a cytokine. These designer APCs more effectively activated tumor antigen-specific CD8 T cells than mRNA encoding the antigen alone and elicited robust anti-tumor immune responses. In addition, substituting IL-2 in the mRNA mixtures with membrane-tethered IL-12 led to the expansion and differentiation of endogenous antigen-specific Th1 helper T cells in vivo. Importantly, this platform activated NY-ESO-1-specific CD8 T cells both in human PBMCs in vitro and in HLA-A*02:01-transgenic mice, highlighting its translational potential. This modular mRNA strategy reprograms somatic cells in situ into designer APCs, providing a flexible and translatable platform for precision immunotherapy.

immunology↗

bioRxiv2025

Transgenerational polarity axis inheritance during Ceratopteris embryogenesis

For sexually reproducing organisms to pass on their genetic information, progeny must successfully establish. Various life history strategies have evolved, using either dispersal of large numbers of progeny or intensive nurturing of a few. Most plants use the former strategy, but ferns generate a single embryo in the exact same location as the mother, and it is unknown how progeny success is promoted, or how embryogenesis is adapted. By studying Ceratopteris richardii embryogenesis, we find that maternal tissues guide orientation of the early embryo body axis, thus aligning its root pole towards the homologous maternal rhizoids. We find that axis polarity inheritance is mediated by maternal tissue mechanical patterns, and thus identify a robust mechanism for progeny establishment as a nurturing strategy in plants.

plant biology↗

bioRxiv2025

Title: KymoTip: High-throughput Characterization of Tip-growth Dynamics in Plant Cells

Live imaging data analysis often requires an objective, local, and accurate way of quantification of cell dynamics. In the research field of polarized tip-growth, the cell fluctuations and/or fluctuations in tip position and growth direction hampers automated analyses of huge amounts of imaging sequences. The fluctuated nature in data makes it unclear how cell shape and growth are linked to intracellular events that could be the actual driving force of cell growth. To overcome these difficulties, we developed a powerful and user-friendly tool called KymoTip with an available format. In this software, novel functions such as coordinate normalization, tip-bottom detection, and signal kymograph were implemented. We confirmed that not only plasma membrane-labeled fluorescent images, but also images such as bright-field and cortical microtubule markers --so long as the cell contours can be identified-- are amenable to KymoTip. Furthermore, by combining markers for cell contours with those that visualize intracellular structures, it becomes possible to quantitatively analyze various intracellular events, such as nuclear migration and calcium wave, in conjunction with cellular growth dynamics. Since KymoTip can be handled by non-specialist, it is expected to promote understanding of what happens at the sub- and cellular level with high throughput outcomes. Significance statementFaced with fluctuations in cell coordinates and cell tip positions, position correction of live imaging data and accurate detection of tip position are key challenges in plant developmental biology. We solved them with a powerful and user-friendly tool, KymoTip, that can realize cell position correction, cell tip detection with cell centerline, and quantification of intracellular events.

bioinformatics↗

bioRxiv2025

A simple and versatile plasma membrane staining method for visualizing living cell morphology in reproductive tissues across diverse plant species

Plant reproduction involves dynamic spatiotemporal changes that occur deep within maternal tissues. In ovules of Arabidopsis thaliana (A. thaliana), one of the two synergid cells degenerates at fertilization, while the fertilized egg cell (zygote) undergoes directional elongation followed by asymmetric division to initiate embryonic patterning. However, morphological analysis of these events has been hampered by the limitations of conventional cell wall staining, which fails to label cells lacking complete walls, and by the requirement for transgenic fluorescent reporters to visualize cell outlines. Here, we report that the membrane-specific fluorescent dye FM4-64 readily permeates ovules, allowing clear visualization of reproductive cell morphology both before and after fertilization. This staining method supports high-resolution time-lapse imaging and quantitative analysis of early embryogenesis in living tissues. Importantly, it is applicable not only to the angiosperm A. thaliana but also to the liverwort Marchantia polymorpha (M. polymorpha) and the fern Ceratopteris richardii (C. richardii), enabling the visualization of live reproductive cell structures within maternal tissues and revealing fertilization-associated morphological changes. This simple and robust method thus provides a valuable tool for spatiotemporal and quantitative analyses of reproductive processes across a broad range of plant species, without the need to generate transgenic lines.

plant biology↗

bioRxiv2025

Distinct localization patterns of actin microfilaments during early cell plate formation in plants through deep learning-based image restoration

Phragmoplasts are plant-specific intracellular structures composed of microtubules, actin microfilaments (AFs), membranes, and associated proteins. Importantly, they are involved in the formation and expansion of cell plates that partition daughter cells during cell division. While previous studies have revealed the important role of cytoskeletal dynamics in the proper functioning of the phragmoplast, the localization and role of AFs in the initial phase of cell plate formation remain controversial. Here, we used deep learning-based image restoration to achieve high-resolution 4D imaging with minimal laser-induced damage, enabling us to investigate the dynamics of AFs during the initial phase of cell plate formation in transgenic tobacco BY-2 cells labeled with Lifeact-RFP or RFP-ABD2 (actin binding domain 2). This computational approach overcame the limitation of conventional imaging, namely laser-induced photobleaching and phototoxicity. The restored images indicated that RFP-ABD2 labeled AFs were predominantly localized near the daughter nucleus, whereas Lifeact-RFP labeled AFs were found not only near the daughter nucleus but also around the initial cell plate. These findings, validated by imaging with a long exposure time, highlight distinct localization patterns between the two AF probes and suggest that Lifeact-RFP labeled AFs play a role in initiating cell plate formation.

plant biology↗

bioRxiv2024

Agent-Based Simulation of Cortical Microtubule Band Movement in the Arabidopsis Zygote

Arabidopsis thaliana zygotes form a circumferential cortical microtubule (CMT) band that moves upward along the longitudinal axis of the cylindrical cell; however, how and why this CMT band moves remain unknown. Based on several recently proposed descriptive feedback mechanisms of CMT reorientation, we hypothesized that a directional cue (DC) changes CMT orientation. We constructed an agent-based simulation model with a DC to observe what happens during CMT formation using different parameters. We determined that the output CMT band can become larger than the input DC zone and identified two types of self-organization processes depending on the parameters used. Furthermore, using well-defined parameters that form a static CMT band, we artificially changed the position of the input DC zone, revealing that CMT band movement requires a DC to move at an appropriate speed.

plant biology↗

bioRxiv2024

Conjugation of dn-OPDA with amino acids inhibits its hormonal bioactivity in Marchantia polymorpha

Jasmonates are important phytohormones activating plant tolerance to biotic and abiotic stress, as well as different development processes. A conserved signalling pathway activated by distinct hormones in different plant species mediates these responses: dinor-12-oxo-phytodienoic acid (dn-OPDA) isomers in bryophytes and lycophytes, and JA-Ile in most vascular plants. The final responses depend, in many cases, on the accumulation of specialized metabolites. To identify novel compounds regulated by the dn-OPDA pathway in Marchantia, untargeted metabolomic analyses were carried out in response to dn-OPDA-regulated stress. A novel group of molecules were identified as dn-OPDA-amino acid conjugates (dn-OPDA-aas), and their accumulation after wounding and herbivory confirmed by targeted metabolic profiling in Marchantia and all species in which we previously found dn-iso-OPDA. Mutants in GRETCHEN-HAGEN 3A (MpGH3A) failed to accumulate dn-OPDA-aa conjugates, and showed a constitutive activation of the OPDA pathway and increased resistance to herbivory. Our results show that dn-iso-OPDA bioactivity is reduced by conjugation with amino acids. Therefore, a dichotomous role of jasmonate conjugation in land plants is highlighted: jasmonic acid (JA) conjugation with isoleucine (Ile) produce the bioactive JA-Ile in tracheophytes, whereas conjugation of dn-iso-OPDA with different amino acids disactivate the hormone in bryophytes and lycophytes.

plant biology↗

bioRxiv2024

Temporal changes in surface tension guide the accurate asymmetric division of Arabidopsis zygotes

In most plants, the zygote divides asymmetrically to define the body axis. In Arabidopsis thaliana, the zygote undergoes polar elongation maintaining a transverse band of cortical microtubules (MTs), and divides asymmetrically forming another MT band, preprophase band (PPB). How the MT band is maintained at the actively growing cell tip and whether it contributes to PPB formation remain elusive. By combining live-cell imaging and mechanical simulation, we show that zygote elongation induces a temporal change (large material derivative) in surface tension at the growing tip to maintain the MT band, which in turn supports polar elongation. The MT band then guides PPB to determine the cell division site. Therefore, autonomous mechanical feedback between cell elongation and MT organization ensures the zygote division asymmetry.

plant biology↗

bioRxiv2024

A viscoelastic-plastic deformation model of hemisphere-like tip growth in Arabidopsis zygotes

Plant zygote cells exhibit tip growth, producing a hemisphere-like tip. To understand how this hemisphere-like tip shape is formed, we revisited a viscoelastic-plastic deformation model that enabled us to simultaneously evaluate the shape, stress, and strain of Arabidopsis (Arabidopsis thaliana) zygote cells undergoing tip growth. Altering the spatial distribution of cell wall extensibility revealed that cosine-type distribution and growth in a normal direction to the surface creates a stable hemisphere-like tip shape. Assuming these as constraints for cell elongation, we determined the best-fitting parameters for turgor pressure and wall extensibility to computationally reconstruct an elongating zygote that retained its hemisphere-like shape using only cell contour data, leading to formulation of non-dimensional growth parameters. Our computational results demonstrate the different morphologies in elongating zygotes through effective non-dimensional parameters.

plant biology↗