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Fujita, S.

Publications and source records attributed to Fujita, S..

At least 19 recordsLinked to original sources

UNKAI: A protein functional identity prediction model based on ESM-C latent representations and the attention mechanism

The rapid advancement of genome sequencing technologies has led to the accumulation of a vast number of protein sequences in public databases. However, a significant proportion of these proteins remain functionally uncharacterized. Concurrently, the expansion of protein sequence data has enabled the development of protein language models (pLMs). By distilling billions of years of evolutionary history into a latent representational space, these models have acquired an unprecedented capacity to predict both the tertiary structures and functions of proteins. In this study, we developed a deep learning-based method to predict whether two proteins catalyze the same enzymatic reaction. Our approach leverages latent representations generated by ESM Cambrian (ESM C), a state-of-the-art pLM, which are then processed through a neural network architecture integrating an attention mechanism. Our method outperformed existing approaches, including those based solely on full-length sequence similarity. Notably, it also surpassed our previous LightGBM-based model, which relied on structural similarity scores derived from AlphaFold-predicted models. Analysis of the attention weights reveals that our model autonomously highlights biologically significant sites, such as catalytic and binding residues. This demonstrates that integrating pLMs with attention mechanisms can enhance the accuracy and interpretability of protein function prediction while eliminating the need for manual feature engineering.

bioinformatics↗

Novel African Rhinolophus bat ACE2 sequences reveal the determinants of Afro-Eurasian sarbecovirus entry

Sarbecoviruses, including SARS-CoV and SARS-CoV-2, are frequently linked to Rhinolophus bats as their putative natural reservoirs. Angiotensin-converting enzyme 2 (ACE2), a host carboxypeptidase widely expressed in mammalian tissues, plays a critical role in sarbecovirus infection by serving as the cellular receptor for the viral spike (S) protein. Given recent human outbreaks and pandemics caused by members of sarbecoviruses, and the wide distribution of Rhinolophus bats, it is essential to maintain surveillance of these viruses while improving our understanding of their interactions with bat hosts, particularly the ACE2 receptor. However, while Rhinolophus bats from Asia have been relatively well studied, African Rhinolophus bats remain underrepresented and require further investigation. In this study, five Rhinolophus bat lung samples were obtained from Zambia, and ACE2 genes from these individuals were cloned and sequenced. We further evaluated the susceptibility of ACE2 variants to a panel of sarbecoviruses, revealing key residues that influence viral infectivity. ACE2 polymorphism was observed among Rhinolophus simulator individuals, revealing multiple ACE2 genotypes within the sampled population. However, R. simulator ACE2s did not permit infection by the clade 3 Afro-Eurasian sarbecoviruses tested in this study. Notably, RhGB01 and BM48-31 virus utilized only Rhinolophus blasii ACE2. Mutational analyses further suggested that ACE2 residues 31 and 41 play important roles in modulating spike-ACE2 interactions. This study reports 4 unique ACE2 sequences of R. simulator and R. blasii, and provides new insights into the molecular interactions between African Rhinolophus species ACE2s and the S protein of sarbecoviruses circulating in Africa and Europe. ImportanceAs putative natural reservoirs of sarbecoviruses, including SARS-CoV and SARS-CoV-2, Rhinolophus bats play a critical role in the emergence of zoonotic coronaviruses, making it essential to understand their interactions with these viruses for future pandemic preparedness. While Asian Rhinolophus bats have been relatively well studied, African species remain underrepresented, highlighting the need for further investigation. In this study, we cloned and sequenced ACE2 genes of five Rhinolophus bats collected in Zambia, Africa. We identified ACE2 polymorphism among Rhinolophus simulator individuals, although this variation was not associated with susceptibility to the clade 3 Afro-Eurasian sarbecoviruses examined. In addition, we identified key ACE2 residues that govern SARS-CoV-2 spike-ACE2 interactions and contribute to distinct infectivity patterns across species. These findings expand our understanding of the molecular determinants of sarbecovirus host range and support improved surveillance and risk assessment of emerging coronaviruses.

microbiology↗

Experimental SARS-CoV-2 infection using horseshoe bats

Horseshoe bats are known as the natural reservoir of sarbecoviruses. To understand how horseshoe bats coexist with sarbecoviruses in nature, experimental infection can provide direct evidence. However, in vivo infection studies using horseshoe bats have been lacking because of the difficulty of maintaining insectivorous bats in a laboratory setting. Here, we established a stable husbandry system for greater horseshoe bats (Rhinolophus ferrumequinum) and performed experimental infection with SARS-CoV-2. In contrast to Syrian hamsters which showed substantial viral replication, infected horseshoe bats exhibited low-level but persistent viral replication in the lung without overt disease. Histological analyses revealed that inflammatory lesions in the bat lungs were spatially restricted and temporally delayed compared with those in hamsters. Transcriptomic analyses further showed preferential activation of tissue repair pathways but limited inflammatory responses following infection. Notably, bats expressed several interferon-stimulated genes prior to infection. Our results suggest that a host strategy combining constitutive antiviral state, limited inflammation and enhanced tissue repair may result in controlled viral replication without overt disease, likely enabling horseshoe bats to coexist with sarbecoviruses.

microbiology↗

Division of labor between ERK and Notch signaling coordinates distinct stem cell populations during jellyfish tentacle regeneration

Regeneration restores lost tissues through diverse cellular strategies that vary across species and tissue contexts. In many systems, regeneration involves blastema formation; however, how blastema formation is coordinated in tissues employing multiple stem/progenitor populations remains unclear. Here, we investigate tentacle regeneration in the hydrozoan jellyfish Cladonema, in which two distinct proliferative populations contribute to regeneration: resident homeostatic stem cells (RHSCs), which supply differentiated cell types, and repair-specific proliferative cells (RSPCs), which are transiently induced upon injury to form the blastema. While extensive cell death occurs shortly after injury, it is largely dispensable for blastema formation. Using pharmacological inhibition and cell proliferation analyses, we identify ERK/MAPK signaling as a key regulator of blastema formation. ERK signaling is rapidly activated at the injury site and selectively promotes proliferation of RSPCs without affecting RHSC proliferation. In contrast, inhibition of Notch signaling disrupts nematocyte differentiation and induces hyperproliferation of RHSCs, while leaving RSPC proliferation unchanged, indicating that Notch signaling governs the balance between differentiation and self-renewal in RHSCs. Together, these findings reveal a division of labor between conserved signaling pathways and support a model in which regeneration is achieved through spatially and functionally compartmentalized control of distinct stem cell populations.

developmental biology↗

Non-invasive genomic sampling uncovers novel connectivities and origins of confiscated gorillas

BackgroundGorillas are a group of African great apes with two species and four subspecies that are currently critically endangered or endangered. Previous studies that analysed the genetics of wild gorillas from non-invasive samples, such as faeces or hair, analysed short mitochondrial or nuclear markers, which may not reflect the wider nuclear genome. Recent technical advances in target capture hybridisation, enrich the endogenous DNA content of non-invasive samples, allowing contiguous genomic regions to be sequenced. ResultsHere, we generated georeferenced genetic data from faecal and hair samples of 280 wild gorillas, sampled from three of the four gorilla subspecies, across large parts of their present-day distributions. With this expanded representation of gorilla genetic diversity in the wild, we detected three population clusters in western lowland gorillas, with the Sangha River and its affluents acting as significant barriers to gene flow. We reconstructed patterns of past population connectivity between western lowland gorillas in the north-eastern distribution range and Cross River gorillas, which may have been facilitated by a migration corridor also used by the Central and Nigeria-Cameroon chimpanzee subspecies. Finally, we predicted the geographic origins of wild-born gorillas, achieving a mean prediction error of 65 km, with a population-level resolution for mountain gorillas and some populations of western lowland gorillas. ConclusionOur work characterises fine-scale population structure in western lowland gorillas, which will be informative for future conservation strategies. This proof of concept in predicting geographic locations of wild gorillas, will be useful for future applications to geolocalise trafficked or rescued gorillas.

evolutionary biology↗

Genetic diversity in horseshoe bat ACE2 and sarbecovirus spike proteins mutually shape one another

Angiotensin-converting enzyme 2 (ACE2) serves as the entry receptor for a wide diversity of sarbecoviruses naturally harboured by horseshoe bats (genus Rhinolophus). Despite the extensive circulation of these viruses in many horseshoe bat species, the potential interactions between virus and receptor evolution remain poorly understood. We sampled individuals of the intermediate horseshoe bat (Rhinolophus affinis) across Vietnam and identified 15 genotypes of ACE2 proteins, 10 of which are previously unreported. Phylogenetic analysis and infectivity assays with a panel of 36 sarbecovirus spike proteins indicated that the R. affinis ACE2 phylogeny has geographic structuring and genotypes originating from different geographic regions exhibit distinct infectivity phenotypes. We detected site-specific positive selection on ACE2 site 24 with the associated substitutions largely affecting the receptors sarbecovirus infectivity profile. Together, our findings suggest that the R. affinis within-species ACE2 diversity has likely been shaped through selection by past sarbecovirus infection. Similarly on the virus end, we use mutagenesis assays and structural analysis through cryo-EM, to delineate the proximal evolution of the Ra22QT77 spike defined by specialization to the ACE2 genotypes of horseshoe bats found in and near southern Vietnam, where the virus was sampled. Our findings contribute to a better understanding of host-sarbecovirus co-evolution dynamics and provide valuable insights into the receptor usage determinants of these viruses. HighlightsO_LIWe identify a total of 15 ACE2 genotypes in R. affinis bats from Vietnam. C_LIO_LIACE2 intraspecific polymorphism is geographically separated and associated with distinct sarbecovirus infectivity. C_LIO_LISite 24 of R. affinis ACE2 experiences positive selection and controls susceptibility to sarbecoviruses. C_LIO_LIBat ACE2s and sarbecovirus spikes are bidirectionally shaped by each others evolution. C_LI

microbiology↗

Escape from Cell Uptake: Drug-Free Cancer Therapeutics Regulated Hydrophobicity and Negative Charge

Self-aggregation and inducing cell membrane disruption in response to tumor microenvironment-stimuli is expected to be a promising approach for cancer treatment, but is limited by its insufficient stimuli-responsive cytotoxicity due to a lack of in-depth understanding of molecular characteristics, resulting in low selectivity of cell death induction. In this study, we focused on engineering polymer aggregation in detail to further improve tumor microenvironment-responsive cytotoxicity. PVA-U with grafting degrees (G.D.) of 3% (PVA-U3), 15% (PVA-U15), and 25% (PVA-U25) were synthesized and their aggregation properties cytotoxicity was evaluated. The difference in half maximal inhibitory concentration (IC50) values between pH 7.4 and pH 6.5 for PVA-U15 was 4.3-fold, which was greater than that of PVA-U25 at 2.8-fold, suggesting that tumor microenvironment-responsive cytotoxicity could be regulated by controlling G.D. of UDCA. Interestingly, PVA-U15 formed aggregates in the pericellular environment and adsorbed on the cell, effectively inducing cell death whereas PVA-U3 and PVA-U25 showed internalization in the cell. These results indicated that the balance of the surface charge and hydrophobicity could contribute to the adsorption on the cell membrane. These findings are expected to contribute to the development of membrane disruption strategies to control the aggregation properties and cell membrane interaction. Table of ContentsCancer therapeutics strategy targeting tumor microenvironment and cell adsorption of PVA-U with varying granting degree (G.D.). PVA-U is expected to aggregate and disrupt cell membrane in response to tumor microenvironment. PVA-U could regulate cell membrane adsorption by its negative charge and hydrophobicity due to controlling G.D. of UDCA, resulting in optimized tumor microenvironment selectivity. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=181 SRC="FIGDIR/small/671160v1_ufig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@e8d4acorg.highwire.dtl.DTLVardef@1e27a1borg.highwire.dtl.DTLVardef@1beb5d8org.highwire.dtl.DTLVardef@1948d51_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

Novel humanized anti-Nav1.7 antibodies with long-lasting, side-effect-free analgesic effects

Neuropathic pain remains difficult to treat effectively because of the limitations of current pain medications. The Nav1.7 sodium channel plays a crucial role in pain sensation. The development of selective inhibitors has been challenging because of the high similarity among Nav channel subtypes. To address this issue, we developed monoclonal antibodies that selectively target Nav1.7 and humanized them for clinical use for the first time. When administered systemically to neuropathic pain model rats, a potent analgesic effect was observed that lasted for at least 96 hours. Electrophysiological studies revealed that the antibody reduced mechanically-evoked and spontaneous neuronal activity in the model. Importantly, the antibodies did not impact physiological pain or motor function. Collectively, our findings suggest that these novel Nav1.7-targeting antibodies are potentially effective analgesic drugs for chronic pain, including neuropathic pain. Our novel humanized anti-Nav1.7 antibodies have the potential to be used for the development of new analgesic, and one such antibody, S-151128 is currently in clinical trial.

pharmacology and toxicology↗

Self-Assembling Synthetic Peptides Remodel Mitochondrial Properties via Nanopore Formation

Pore-forming peptides (PFPs) are powerful tools for engineering biological membranes in vitro and in living cells. They can be derived from natural sources or designed de novo, with synthetic peptides offering distinct advantages in rational design unconstrained by natural sequences. While native PFPs have been targeted to specific organelles to modulate their functions, the potential of synthetic PFPs for organelle engineering remains largely unexplored. Here, we show that the 28-amino acid synthetic peptides, SVG28 and its variant SVG28D2, can be selectively targeted to mitochondria for functional alteration. More specifically, molecular dynamics simulations first confirmed the structural stability of the SVG28 nanopore even when fused to the TOM20 transmembrane domain, a mitochondrial targeting sequence. In isolated lipid bilayers, TOM20-SVG28 formed pores with a 0.3-nS, conductance, or 1.0-nm pore. The expression efficiency was further enhanced by two amino acid substitutions (SVG28D2). In cells, both TOM20-SVG28 and TOM20-SVG28D2 localized to the mitochondria, induced morphological alterations, and reduced both membrane potential and ATP production. These observations in silico, in vitro and in cellulo collectively indicate that the synthetic SVG28 peptides self-assemble into pores in the outer mitochondrial membrane, likely permitting proton permeation. Importantly, the matrix protein Su9 remained confined within mitochondria, suggesting preserved inner membrane integrity, and thus specific effect on the target. This work establishes a framework for the design and application of genetically-encoded synthetic PFPs in living cells, opening new avenues for cellular engineering.

synthetic biology↗

VEGF-C-mediated Cardiac Lymphangiogenesis Promotes Inflammation Resolution in Autoimmune Acute Myocarditis in Mice

BACKGROUNDAcute myocarditis is an immune-mediated inflammatory disease characterized by myocardial inflammation and edema. Cardiac lymphatic vessels are essential for fluid clearance and immune regulation; however, their role in modulating inflammation in autoimmune myocarditis remains largely unclear. METHODSWe investigated the impact of cardiac lymphangiogenesis using a murine model of experimental autoimmune myocarditis induced by cardiac myosin peptide immunization. Human autopsy samples were analyzed for lymphatic expansion. Mice were treated with VEGF-C C156S, a VEGFR3-specific agonist, beginning one week after immunization. Cardiac lymphangiogenesis, myocardial edema, inflammatory infiltration, fibrosis, and cardiac function were evaluated by immunohistochemistry, echocardiography, gene expression analysis, and water content measurements. RESULTSThe VEGF-C treatment accelerated cardiac lymphangiogenesis, enhanced lymphatic drainage, reduced myocardial edema, and attenuated inflammatory cell infiltration and fibrosis. Echocardiography showed the preservation of left ventricular function. VEGF-C selectively decreased the accumulation of iNOS inflammatory macrophages without broadly suppressing T cells or reparative macrophages. Bulk RNA sequencing confirmed the down-regulation of inflammatory gene signatures associated with macrophage activation. CONCLUSIONSThe early stimulation of cardiac lymphangiogenesis by VEGF-C promotes the resolution of inflammation, reduces myocardial injury, and preserves cardiac function in autoimmune myocarditis. Targeting the cardiac lymphatic system may represent a promising therapeutic strategy for acute myocarditis.

pathology↗

Observation of Field Cancerization in Human Breast Tissue with Raman Hyperspectral Imaging and Artificial Neural Networks

Tissue-conserving partial mastectomy surgeries are often used to treat breast cancer, in which malignancies and surrounding margin tissues are removed. Morphologies of margin tissues are often assessed intraoperatively by a histopathologist, requiring excision and time-consuming preparation for microscopic examination. Changes in morphology often appear only after the mutation of cells involved in tumor propagation, suggesting that margins and perhaps conserved tissue may already contain cancer-converted cells prior to morphological changes. In a process known as field cancerization (FC), benign tissue surrounding a malignancy is affected by continued exposure to carcinogens, triggering cancer-promoting alterations like conversions of normal adipocytes and fibroblasts to cancer-associated adipocytes and fibroblasts. We report successful FC detection in human breast tissue samples excised during breast-conserving surgeries in human patients using Raman hyperspectral imaging and artificial neural networks (ANNs). Spectra in the images are segmented based on tissue modalities such as adipose, connective, and glandular tissues using intensities at Raman shifts of known molecules and label propagation, a method of transductive learning. ANNs that discriminate cancer from noncancer spectra return high quality (>95% accuracy) and reproducible distinction of spectra originating from cancerous tissue at high signal-to-error ratios (S/Es). Good distinction is returned in all modalities, with adipose-rich modalities returning the most reliable classifications, suggesting reliable discrimination of normal and cancer-associated adipocytes. Spectral S/Es are increased using larger spectral measurement areas, but this causes spectral mixing across different tissue modalities and significant biological variation, failing to deliver a corresponding increase in classification quality.

cancer biology↗

Molecular basis of sarbecovirus evolution and receptor tropism in natural hosts, potential intermediate hosts, and humans

The spike protein of many sarbecoviruses binds to the angiotensin-converting enzyme 2 (ACE2) receptor and facilitates viral entry. The diversification of the sarbecovirus spike gene and the mammalian ACE2 gene suggests that sarbecoviruses and their hosts have co-evolved, and the genetic diversity in these genes affects the host tropism of sarbecoviruses. Better comprehending the evolutionary potential of sarbecoviruses can lead to preparedness for the next pandemic. However, the host tropism of sarbecoviruses is not fully understood. Here, we performed round-robin pseudovirus infection assays using 53 sarbecoviruses and ACE2s from 17 mammals to elucidate the ACE2 tropism of sarbecoviruses in natural hosts, potential intermediate hosts and humans. We determined the factors responsible for the ACE2 tropism of sarbecoviruses through structural, phylogenetic analyses, and infection experiments, revealing which substitutions can expand the host range of sarbecoviruses. These results highlight the mechanisms modulating host tropism throughout sarbecovirus evolution.

microbiology↗

Cross-Species Biomechanical Determinants of Shape Diversity

How complex molecular mechanisms translate into diverse multicellular shapes remains unclear. By leveraging the bi-layered architecture of six cnidarian species that diverged 500 million years ago, we show that modularity in supracellular mechanics governs larval shape diversity. Using active surface theory, quantitative imaging, and an inducible genetic system, we identify species-specific variations in three biomechanical modules. Basally aligned stress fibers drive axial elongation, while oral geometry and aboral rigidity define shape polarity. Remarkably, manipulating these modules transforms one species shape into another, demonstrating the causal relationship between module variation and shape diversity. Our analysis also uncovers instances of mechanical redundancies, where distinct module combinations generate similar shapes. These findings provide a general framework for how molecular complexity funnels into mesoscale mechanical determinants shaping morphological diversity.

developmental biology↗

Enhanced prediction of protein functional identity through the integration of sequence and structural features

Although over 300 million protein sequences are registered in a reference sequence database, only 0.2% have experimentally determined functions. This suggests that many valuable proteins, potentially catalyzing novel enzymatic reactions, remain undiscovered among the vast number of function-unknown proteins. In this study, we developed a method to predict whether two proteins catalyze the same enzymatic reaction by analyzing sequence and structural similarities, utilizing structural models predicted by AlphaFold2. We performed pocket detection and domain decomposition for each structural model. The similarity between protein pairs was assessed using features such as full-length sequence similarity, domain structural similarity, and pocket similarity. We developed several models using conventional machine learning algorithms and found that the LightGBM-based model outperformed the models. Our method also surpassed existing approaches, including those based solely on full-length sequence similarity and state-of-the-art deep learning models. Feature importance analysis revealed that domain sequence identity, calculated through structural alignment, had the greatest influence on the prediction. Therefore, our findings demonstrate that integrating sequence and structural information improves the accuracy of protein function prediction.

bioinformatics↗

Direct non-transcriptional link between brassinosteroid perception and cortical microtubule reorientation drives hypocotyl growth

Brassinosteroids (BRs) are steroid-type phytohormones that are essential for plant growth, development and adaptation to environmental stresses. BRs promote cell wall remodelling through extensive reprogramming of gene expression, thus impacting on cell elongation and hypocotyl growth. The existence of non-genomic responses to BRs is however still unclear. Here we identified and characterized a new family of BRI1-interacting proteins identified by yeast two hybrid named MBAPs. We confirmed using several complementary approaches that MBAPs are genuine BRI1 partners in plant cells. We demonstrated that MBAPs are localized to cortical microtubules (CMTs) underlying the plasma membrane using a short conserved helix and where they contact with and are phosphorylated by BRI1. Combinations of mbap loss-of-function mutants showed hypersensitivity to BRs and establish MBAPs as negative regulators of BR responses. Surprisingly, mbap mutants showed unaffected typical downstream BR signaling readouts and BR target gene expression. Rather, mbap mutants displayed disordered CMT network and enhanced CMT reorganization upon BR perception. Altogether, our work shed light on the direct non-transcriptional connection between BR perception at the cell surface and CMT organization in the control of hypocotyl elongation.

plant biology↗

Model selection reveals selective regulation of blood amino acid and lipid metabolism by insulin in humans

Insulin plays a crucial role in regulating the metabolism of blood glucose, amino acids, and lipids in humans. However, the mechanisms by which insulin selectively controls these different metabolites are not yet fully understood. To address this question, we used mathematical modeling to identify the selective regulatory mechanisms of insulin on blood amino acids and lipids. Our study revealed that insulin negatively regulates the influx and positively regulates the efflux of lipids, which is consistent with previous findings. In contrast, we did not observe the previously reported negative regulation of BCAA influx by insulin; instead, we found that insulin positively regulates BCAA efflux. We also observed that the earlier peak time of lipids compared to BCAAs is dependent on insulins negative regulation of their influx. Overall, our findings shed new light on how insulin selectively regulates the levels of different metabolites in human blood, providing valuable insights into the pathogenesis of metabolic disorders and potential therapeutic interventions.

systems biology↗

Cold-induced muscle atrophy in zebrafish: insights from swimming activity and gene expression analysis

The investigation into the effects of cold acclimation on fish skeletal muscle function and its potential implications for muscle atrophy is of great interest to us. This study examines how rearing zebrafish at low temperatures affects their locomotor activity and the expression of genes associated with muscle atrophy. Zebrafish were exposed to temperatures ranging from 10 {degrees}C to 25 {degrees}C, and their swimming distance was measured. The expression levels of important muscle atrophy genes, Atrogin-1 and MuRF1, were also evaluated. Our findings show that swimming activity significantly decreases when the water temperature ranges from 10 {degrees}C to 15 {degrees}C, indicating a decrease in voluntary movement. Additionally, gene expression analysis shows a significant increase in the expression of Atrogin-1 and MuRF1 at 10 {degrees}C. This up-regulation could lead to muscle atrophy caused by decreased activity in cold temperatures. To investigate the effects of exercise on reducing muscle atrophy, we subjected zebrafish to forced swimming at a temperature of 8 {degrees}C for ten days. This treatment significantly reduced the expression of Atrogin-1 and MuRF1, emphasizing the importance of muscle stimulation in preventing muscle atrophy in zebrafish. These findings suggest that zebrafish can serve as a valuable model organism for studying muscle atrophy and can be utilized in drug screening for muscle atrophy-related disorders. Cold-reared zebrafish provide a practical and ethical approach to inducing disuse muscle atrophy, providing valuable insights into potential therapeutic strategies for addressing skeletal muscle atrophy.

physiology↗

Comparisons of two receptor pathways in a single cell-type reveal features of signalling specificity.

To respond appropriately to diverse stimuli, cells harbour numerous receptor pathways yet share downstream signalling components. Mitogen-Activated Protein Kinase (MPK) cascades, present in all eukaryotes, act as central hubs shared amongst diverse signalling pathways. How signalling specificity is maintained within a single plant cell-type is not well understood. We engineered a genetic background for direct comparisons of a developmental and an immunity pathway in the Arabidopsis root endodermis. We show the two pathways maintain distinct outputs despite similar MPK phosphorylation patterns. Systematic activation of all individual MPK Kinases (MKKs) in the endodermis revealed that different MKK groups can drive distinct outputs. We propose that specificity is achieved through a balance of combinatorial activation and cross-pathway inhibition within the MPK cascade. Our findings demonstrate the ability of MPK cascades to generate diverse outputs in one plant cell-type, providing new insights into the mechanisms contributing to signalling specificity.

plant biology↗