Biology subjects

Inoue, S.

Publications and source records attributed to Inoue, S..

At least 19 recordsLinked to original sources

bioRxiv2026

NECTIN4 defines a subtype-dependent tumor state coupling enfortumab vedotin targetability to antitumor immunity in bladder cancer

NECTIN4 is the target of enfortumab vedotin (EV), but its biological role in bladder cancer (BC) and its relationship to EV plus PD-1 blockade remain unclear. Integrating clinical multi-omics, spatial and single-cell immune profiling, and experimental models, we identify NECTIN4 as a subtype-dependent determinant of tumor state and antitumor immunity. In basal BC cells, NECTIN4 suppressed inflammatory and mesenchymal programs. Consistently, NECTIN4-high basal/squamous muscle-invasive BCs showed favorable survival, higher checkpoint inhibitor response rates, and a coordinated CD8+ T cell-inflamed microenvironment with no comparable associations in luminal papillary tumors. In orthotopic models, murine Nectin4 promoted CD8+ T cell-dependent tumor control, whereas human NECTIN4 conferred EV sensitivity. EV also attenuated NECTIN4-dependent TIGIT binding. Conversely, NECTIN4-low models were preferentially sensitive to HDAC inhibition. NECTIN4 therefore couples antibody-drug conjugate targetability to a subtype-specific tumor-immune state permissive for checkpoint blockade, whereas the NECTIN4-low state defines a distinct inflammatory-mesenchymal state with an alternative therapeutic vulnerability.

cancer biology↗

bioRxiv2026

Characterization of central carbon metabolism of Acinetobacter sp. Tol 5 based on 13C-metabolic flux analysis

Acinetobacter sp. strain Tol 5 is an attractive microorganism because of its ability to utilize various aromatic compounds, including toluene. However, because Tol 5 is a non-model microorganism, the utilization of carbon sources in central carbon metabolism remains unclear. In this study, the activity of the central carbon metabolism was characterized using 13C-metabolic flux analysis (13C-MFA). Tol 5 was cultured in synthetic medium using ethanol or acetate as the sole carbon source. In addition, because toluene is degraded into pyruvate and acetyl-CoA via the toluene dioxygenase pathway, this situation was mimicked by culturing with ethanol and lactate as co-substrates. The 13C-MFA confirmed flux through the glyoxylate shunt, which bypasses part of the TCA cycle when either ethanol or acetate was used as the sole carbon source. In ethanol, the flux ratio of isocitrate dehydrogenase at the isocitrate branch point decreased markedly. Unlike acetate, the conversion of ethanol to acetyl-CoA is accompanied by NADH production. Therefore, the observed change in this flux ratio was influenced by the excess NADH generated. Under ethanol-lactate conditions, Tol 5 simultaneously utilized ethanol and lactate. The ethanol consumption rate decreased to less than half of that observed when ethanol was the sole carbon source. Most of the lactate uptake was converted to pyruvate and metabolized via the TCA cycle. The glyoxylate shunt flux was lower than that in ethanol. These metabolic characteristics provide valuable insights into the use of Tol 5 as a next-generation microbial cell factory.

microbiology↗

bioRxiv2026

Highly plastic macrophage niches orchestrate acquired quiescence and reactivation in breast-cancer bone metastasis

Recurrence and metastasis remain major causes of cancer mortality, sustained by therapy-resistant micrometastatic cells. Bone is a frequent site of breast-cancer relapse, yet the cues that reawaken disseminated cells remain poorly defined. We identify a previously unrecognized, highly plastic CXCL16 macrophage population that integrates tumor-associated macrophage programs found in distant metastatic sites such as lung and brain with non-tumor disease-associated traits in bone marrow. These CXCL16 macrophages establish a transient niche that restrains disseminated cancer-cell proliferation. Single-cell transcriptomics delineate functional remodeling of myeloid niches within the bone metastatic microenvironment: a CXCL16 macrophage niche that transiently constrains metastatic growth, and G-CSF macrophage and neutrophil niches that reignite tumor outgrowth. In primary tumors, cancer-associated fibroblasts (CAFs) aberrantly secrete G-CSF in response to cancer-cell signals, expanding G-CSF-receptor-positive subset of cancer cells with high metastatic potential. In advanced human bone metastases, CXCL16 macrophages localize to CAF-rich stroma but are excluded from cancer-cell clusters, indicating immune evasion. Together, these findings uncover CAF-bone-marrow cross-talk as a therapeutic target linking stromal inflammation, immune remodeling, and metastatic progression.

cancer biology↗

bioRxiv2026

Distribution-Constrained Optimization for Reliable ML-Guided 5'UTR Sequence Design

The 5' untranslated region (5' UTR) shapes translation initiation, so its design is central to mRNA therapeutics and to improving protein-production cell lines. Deep-learning models that predict translation efficiency, measured as mean ribosome load (MRL), from the 5' UTR sequence have been combined with genetic algorithms (GAs) for sequence optimization. However, optimizing against a model trained on offline data risks reward hacking that exploits the models estimation error outside the training distribution, yielding sequences that score highly in prediction yet fail to perform in the wet lab. Yet for 5' UTR design, few studies have systematically examined which region should be treated as untrustworthy (the definition of out-of-distribution, OOD) or which constraints keep the search away from it. We present a constrained optimization that keeps candidates within a trust region where the predictors validated accuracy holds; here "reliable" denotes keeping candidates within the training distribution over which prediction has been validated, not a guarantee of measured performance. As the OOD score, we compare the k-nearest-neighbor (KNN) distance in the predictors embedding space against a pseudo-perplexity (PPPL) from the encoder and LM head, and show that for nucleotide sequences--whose vocabulary is small--PPPL fails to separate in- vs out-of-distribution, whereas the KNN distance is an effective OOD score that can define a trust region even from unlabeled native UTR sequences. Using the KNN distance as a hard GA constraint keeps all candidates inside the trust region while maintaining predicted MRL: under unconstrained optimization most final-generation candidates (72-96% across seeds) left the trust region (self-KNN p95), whereas the hard constraint holds predicted MRL at the unconstrained level and yields about 4.3x more selectable low-risk candidates than post-hoc filtering of the unconstrained output. Comparing an output extrapolation guard, reference-sequence similarity and structural accessibility (RNAplfold), we find that the guard and the similarity constraint also suppress OOD as a side effect, whereas making accessibility a secondary objective broadens the search without suppressing OOD.

bioinformatics↗

bioRxiv2026

Large-scale dimensional behavioral profiling dissociates fear memory from locomotor confounds in mice: The necessity of baseline-normalized metrics

Fear conditioning is widely used to assess associative memory in mice, yet percent freezing conflates memory with baseline locomotor and anxiety-related traits. A systematic survey of recent studies (2020-2025) found that fewer than 1% statistically integrate locomotor activity into freezing analyses. Here, we address this gap using a large-scale dataset of >10,000 mice across >160 comparisons, including genetic mutations, pharmacological interventions and aging, tested in 15 standardized behavioral paradigms. Conventional freezing scores covaried strongly with general locomotor activity, obscuring memory-related phenotypes. Multiple factor analysis identified two principal behavioral dimensions, locomotor activity and learning/memory: conventional freezing aligned with the locomotor dimension, whereas freezing subtraction and the activity suppression ratio mapped onto the memory dimension and improved detection of synaptic plasticity phenotypes. These analyses show that baseline locomotor normalization is essential for interpreting fear conditioning as a memory assay and provide an open framework for selecting and reporting locomotor-normalized metrics.

neuroscience↗

bioRxiv2026

A Passive-Oxygenation Silicone Platform for Biomass Production: Maximizing Labor Productivity and Process Efficiency in Cellular Agriculture Development

The commercial production of cell-based food is currently hindered by existing bioreactor technologies, which require substantial capital investment, specialized operating skills, and complex processing setups. To democratize cell-based food production, we developed the "oxy-thru cultivator"--a simple, autoclavable, closed-bag bioreactor fabricated from polydimethylsiloxane (PDMS). By leveraging the high oxygen-permeability of PDMS, this platform enables passive oxygenation across the entire vessel wall, eliminating the need for external aeration or mechanical sparging. During testing, the cultivator maintained a stable culture environment over 23 days, showing no cytotoxic leachables and retaining both structural integrity and sterility across 10 autoclave cycles. This robustness supported the continuous cultivation of DF-1 cells for 74 days. Using a standardized subculture scheme, we successfully harvested an estimated 2.60 g of cell-based biomass per cultivator over five passages. Notably, the platform achieved a 127% monthly labor productivity compared to conventional bioreactors and was easily operated by researchers without specialized training. Additionally, the system successfully supported the expansion of both mammalian and primary avian cell lines. With a minimal equipment footprint that reduces CapEx, and a reusable silicone vessel that lowers OpEx, the oxy-thru cultivator offers a highly practical, accessible pathway toward scaling up cellular agriculture. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC="FIGDIR/small/729703v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@644879org.highwire.dtl.DTLVardef@1d23680org.highwire.dtl.DTLVardef@1f83d9forg.highwire.dtl.DTLVardef@959bf7_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2026

Enzyme activity as an actionable axis for small-molecule precision oncology

Current precision oncology--molecular targeted therapies and immunotherapies--relies on genomic or expressed biomarkers, yet most cancer patients remain ineligible for these treatments. Here, we establish enzyme activity as an actionable and orthogonal axis for precision cancer medicine. Strategic activity-based screening of mouse organs and human clinical specimens with a panel of enzyme-reactive fluorescence probes identified {beta}-galactosidase 1 (GLB1) and {beta}-hexosaminidases (HEX) as broadly elevated tumor-selective biomarkers. Leveraging these activities, we developed 7-ethyl-10-hydroxycamptothecin (SN38)-based GLB1-and HEX-reactive prodrugs. These prodrugs exhibited dramatically reduced systemic toxicities and improved therapeutic windows, compared to a clinically used SN38-based prodrug, irinotecan (CPT-11). Both prodrugs demonstrated activity-dependent therapeutic efficacy, affording a dramatic reduction of tumor volumes across multiple in vivo models, including a subcutaneous patient-derived xenograft (PDX) of lung squamous cell carcinoma that lacked genetic alterations targeted by current precision medicine. Furthermore, this strategy is broadly applicable across various cytotoxic payloads, establishing a generalizable platform for small-molecule precision medicines. Our results define an enzyme-targeting paradigm for precision oncology, in which fluorescence probes serve as companion diagnostic tools to guide development and selection of appropriately targeted prodrugs, which are expected to provide safer and more efficacious treatment options for cancer patients with elevated enzyme activities.

cancer biology↗

bioRxiv2026

Gas-phase environment activates an alternative catabolic route in toluene-degrading Acinetobacter

Volatile aromatic compounds are important industrial feedstocks but also major environmental pollutants, highlighting the need for bioprocesses for their removal and valorization. Although gas-phase bioprocesses offer practical advantages for handling poorly water-soluble and highly volatile substrates, how gas-phase environments alter microbial metabolism remains poorly understood. Here, we investigated the effect of gas-phase conditions on toluene metabolism in the highly adhesive aromatic hydrocarbon-degrading bacterium Acinetobacter sp. Tol 5. A mutant lacking todC1, which encodes an essential component of the toluene dioxygenase, failed to grow on toluene in liquid culture but retained the ability to grow on solid media under a toluene atmosphere. Consistent with this phenotype, the mutant showed no detectable toluene degradation in the liquid phase, whereas it degraded toluene under gas-phase conditions after a prolonged lag phase. Gas chromatography-mass spectrometry (GC-MS) analysis revealed the accumulation of o-cresol and p-cresol specifically in the mutant under toluene vapor, indicating that toluene metabolism had shifted to an alternative route involving cresol intermediates. In addition, transcriptome analysis identified strong induction of the mph operon encoding phenol monooxygenase (PMO), suggesting that PMO is a likely candidate enzyme mediating TDO-independent toluene oxidation under gas-phase conditions. Together, these results demonstrate that the gas-phase environment can activate an alternative catabolic route in Tol 5 that is not active during conventional liquid cultivation. Our findings highlight the importance of direct metabolic analysis under gas-phase conditions for understanding and designing bioprocesses using highly volatile substrates.

microbiology↗

bioRxiv2026

Integrated omics analysis reveals reorganization of nitrogen and lipids metabolism in a toluene-degrading bacterium

Gas-phase bioprocesses that immobilize microbial cells on solid carriers enable the efficient conversion of poorly water-soluble gaseous substrates, thereby offering significant potential to advance bioremediation and bioproduction. However, microorganisms in the gas phase are exposed to various environmental stresses, mainly due to the absence of bulk water. While survival strategies of microorganisms in gaseous environments have been studied in environmental microbiology, the metabolic adaptations that sustain bacterial cell activity remain poorly understood. In this study, we elucidated the comprehensive metabolic alterations of a highly adhesive bacterium Acinetobacter sp. Tol 5 degrading toluene under gas- and aqueous-phase conditions. An integrated approach combining metabolomics, lipidomics, and transcriptomics revealed significant differences in metabolic profiles between cells under these conditions. Under the gas-phase condition, the degradation of amino acids and nucleic acids was significantly promoted, and the intracellular glutamate pool was maintained at high levels. Notably, citrulline was found to accumulate specifically under the gas-phase condition, representing a stress response similar to that reported in Cucurbitaceae plants during drought. Furthermore, lipidomics revealed the lipid composition of Tol 5 and demonstrated a shift in response to environmental conditions. Specifically, the degradation of intracellular storage lipids was promoted under gas-phase conditions, suggesting a crucial link to bacterial survival in water-limited environments. These findings provide critical insights into the adaptation strategies of bacteria adapting to gaseous environments, offering fundamental information for the rational design of robust gas-phase bioprocesses and a deeper understanding of environmental microbiology.

microbiology↗

bioRxiv2026

Integration of UAS-based spatial surveys and bio-logging tracking enhances precision in population size estimation

Accurately estimating wildlife population sizes, essential for ecological theory and conservation management, yet remains challenging. Although unmanned aerial systems (UASs) combined with machine learning, have revolutionized population estimation, they face limitations in addressing the hierarchical population processes from individual behavior to colony-and population-level dynamics. To overcome this limitation, we developed a data integration framework that jointly analyzes multiple datasets, representing different scales of the same underlying process, were jointly analyzed. Using a seabird colony as a model system, we integrated UAS-based count data in the colony with bio-logging-based tracking data to estimate population size by quantifying both the number of individuals present and the proportion absent from the surveyed area. These complementary datasets were linked using state-space models allowing accurate population estimates with explicit uncertainty quantification. Furthermore, we evaluated the robustness of the estimations with respect to sample size. Sub-sampling simulations revealed that estimation uncertainty was more sensitive to sample size in bio-logging-based tracking data than in UAS-based count data. This finding highlights the importance of understanding dataset-specific properties when designing effective investigations. Overall, our resource-efficient framework is broadly applicable across species and populations and demonstrates how integrating complementary observation methods can improve population estimates and inform conservation practice.

ecology↗

bioRxiv2026

An integrated scalable process for adherent cultivated meat production: From proliferative cell selection to safety-verified product development

Cultivated meat can contribute to the global food supply; therefore, establishing efficient production processes is an urgent task to meet the growing demand for sustainable protein. Although cell culture technology has historically focused on suspension cells in the pharmaceutical and, subsequently, cultivated meat industries, the development of efficient processes for most adhesion-dependent cells has lagged, with limited examples reported to date. Therefore, this study used primary duck liver-derived adherent cells to develop and evaluate an integrated three-step meat production process involving pre-culture to select highly proliferative cells, packed-bed bioreactor expansion for mass production, and final processing, including packaging and heating. The established process allowed the efficient growth of selected cell populations while maintaining their proliferative characteristics. Moreover, the developed product met the microbiological and heavy metal safety criteria. Comprehensive compositional analysis (nutritional, amino acid, and fatty acid profiles) revealed that the product exhibited a protein profile distinct from that of conventional duck liver paste, along with a unique lower-fat signature. As productivity is dependent on cell doubling time, culture duration and monthly production were estimated for cells from various animal species. Overall, this study established a practical integrated production process for cultivated meat using adherent cells, providing a technological foundation for cellular agriculture applicable to diverse cell types and useful for future food supply diversification. GraphicalAbstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/698307v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@18b1148org.highwire.dtl.DTLVardef@11bbcecorg.highwire.dtl.DTLVardef@102d99org.highwire.dtl.DTLVardef@21af6f_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2025

Desiccation-tolerant Acinetobacter as a robust chassis for gas-phase bioprocesses

BackgroundGas-phase bioprocesses that immobilize microbial cells on solid carriers enable efficient conversion of volatile or poorly water-soluble substrates, but they also expose cells to fluctuating humidity and transient desiccation that can compromise viability and catalytic performance. In Gram-negative bacteria, especially in pathogens, desiccation is known to impose multifactorial stress, including loss of cellular water and energetics, damage to DNA and proteins, oxidative stress, and disruption of the cell envelope. However, for chassis strains used in or being considered for gas-phase bioprocesses, their desiccation tolerance and responses to desiccation stress remain incompletely defined. ResultsHere, we evaluated the viability, energy status, and gas-phase toluene degradation of Acinetobacter sp. Tol 5 as a chassis for gas-phase processes after controlled desiccation (8%, 52%, and >95% RH), in comparison with Acinetobacter baylyi ADP1, Pseudomonas putida, and Escherichia coli. To minimize adhesion-related bias in post-desiccation measurements, we used an ataA-deficient Tol 5 mutant (Tol 5 {Delta}ataA) for the assays. Acinetobacter strains maintained high viability during desiccation for 16 days, whereas P. putida and E. coli showed significant loss of viability at 8% RH and 52% RH. Intracellular ATP measurements further indicated that desiccation reduced intracellular ATP in all strains, but E. coli rapidly exhausted ATP at 8% and 52% RH, whereas Acinetobacter strains and P. putida retained intracellular ATP. In a gas-phase toluene degradation assay, immobilized Tol 5 retained higher toluene-degrading activity after desiccation than P. putida mt-2. Transcriptome profiling revealed a multilayered Tol 5 response involving DNA and RNA maintenance, cell envelope trafficking and remodeling, redox-responsive functions, and broad repression of growth-associated metabolism. ConclusionsOur results highlight Acinetobacter, particularly Tol 5, as a promising chassis strain for gas-phase bioprocesses and suggest potential mechanistic targets for stabilizing bacterial cells under low-water-activity conditions.

microbiology↗

bioRxiv2025

Chromosome-scale genome assembly and annotation of the two-spotted cricket Gryllus bimaculatus (Orthoptera: Gryllidae)

The two-spotted cricket, Gryllus bimaculatus, is a key hemimetabolous model organism for developmental biology, neuroscience, and regeneration. The existing reference genome is, however, highly fragmented into 47,877 scaffolds, hampering chromosome-scale analyses for these fields. Here, we report a high-quality, chromosome-scale genome assembly for the white-eyed mutant strain of this cricket, generated using a combination of Nanopore and PacBio HiFi long reads, integrated with Hi-C data. The final 1.62 Gbp assembly achieves a scaffold N50 of 107.4 Mbp, a significant improvement in contiguity over the previous 6.3 Mbp N50. We anchored 94.45% of the assembly into 15 pseudomolecules, consistent with the known karyotype (n = 15). The genome completeness (BUSCO v6.0.0 insecta_odb12) reached 98.1%. We also updated the annotation, identifying 14,964 protein-coding genes. This gene set shows markedly improved completeness (BUSCO v6.0.0 insecta_odb12: 95.7%) compared with the previous annotation (81.2%) and successfully recovers all nine essential neuropeptide genes previously reported as missing from the draft assembly. This chromosome-scale genomic resource provides an essential foundation for comparative and functional genomics in G. bimaculatus.

genomics↗

bioRxiv2025

Metabolic pathway analysis of an Acinetobacter strain capable of assimilating diverse hydrocarbons

Sustainable bioproduction requires developing robust microbial chassis with broad metabolic versatility and suitability for industrial applications. Acinetobacter sp. Tol 5 is a highly adhesive bacterium capable of utilizing various hydrocarbons, making it a promising chassis candidate for immobilized whole-cell catalysis. In this study, we characterized the carbon metabolism of Tol 5 by reconstructing metabolic pathway maps from its genomic data and analyzing the transcriptomes of cells grown on ethanol, hexadecane, toluene, and phenol. Genomic analysis revealed that Tol 5 has limited capacity for sugar utilization but possesses a wide range of metabolic pathways for alkane and aromatic compounds, including five distinct aromatic degradation routes that expand the known metabolic diversity of the genus Acinetobacter. Transcriptome analysis identified the specific pathway genes induced in response to each carbon source and revealed substrate-dependent cross-regulation between aromatic degradation pathways. Gene disruption experiments further demonstrated that toluene dioxygenase facilitates rapid entry into exponential growth on phenol but reduces carbon assimilation efficiency, while phenol monooxygenase serves as the primary and indispensable route for phenol assimilation, revealing a different physiological role for toluene dioxygenase in Tol 5 compared with Pseudomonas putida strains. These findings provide a comprehensive view of the carbon metabolism of Tol 5 and highlight its potential as a microbial chassis for bioprocesses utilizing non-sugar feedstocks, while revealing new aspects of metabolic versatility in the genus Acinetobacter.

microbiology↗

bioRxiv2025

Identification and functional characterization of toluene degradation genes in Acinetobacter sp. Tol 5

Microbial degradation of aromatic compounds provides sustainable solutions for environmental remediation and bioconversion. Acinetobacter sp. Tol 5 is notable for its strong adhesiveness and potential as a biocatalyst for toluene degradation; however, its toluene metabolic pathway has not been fully elucidated. In this study, genomic analysis identified a cluster of genes in Tol 5 highly similar to the well-known tod operon of Pseudomonas putida, encoding enzymes responsible for toluene metabolism. Phylogenetic analyses indicated that these tod genes, unusual among Acinetobacter species, were likely acquired through horizontal gene transfer. Transcriptomic analyses revealed that todF and todC1 are co-transcribed, while the adjacent fadL2 gene, encoding a putative outer membrane transporter corresponding to P. putida todX, is independently transcribed. Functional characterization using gene-knockout mutants revealed that TodC1, the large subunit of dioxygenase, is essential for growth on toluene, whereas FadL2 is not essential. Growth experiments further showed that the todC1 knockout mutant could metabolize benzoate, but not toluene or benzene, confirming that the TOD pathway is the primary route for toluene and benzene degradation in Tol 5. The identification of the functional TOD pathway, which is unique within Acinetobacter, provides genetic and biochemical insights for the development of Tol 5 as an efficient immobilized biocatalyst for the bioremediation and bioconversion of aromatic compounds.

microbiology↗

bioRxiv2025

Mitorubin, berberrubine-based compounds that improve mitochondrial function, exhibit cardioprotective effects against age-related cardiac dysfunction

Mitochondria play a central role in cellular energy metabolism and homeostasis, and their dysfunction is closely linked to the progression of age-related diseases. The mitochondrial ubiquitin ligase MITOL (also known as MARCHF5) is a key regulator of mitochondrial dynamics and function, and reduced MITOL expression in the mouse heart has been implicated in mitochondrial dysfunction and cardiac aging. In this study, we identified berberrubine as a compound that promotes MITOL expression and activates mitochondria. We further assembled a group of berberrubine-based compounds, including its quinoid form and a newly developed water-soluble derivative, and collectively named them "Mitorubin" as mitochondria-activating compounds with therapeutic potential. While conventional berberrubine has poor water solubility, the addition of acetic acid significantly improved its solubility, enabling formulation as a solution. Mitorubin enhanced MITOL expression in cultured cells, increased mitochondrial DNA content and expression of mitochondrial proteins, and promoted mitochondrial respiration. In a model of age-related cardiac dysfunction, oral administration of Mitorubin restored mitochondrial function, improved cardiac performance, suppressed myocardial hypertrophy, and alleviated pulmonary congestion. Moreover, Mitorubin did not shorten lifespan in aged mice and significantly extended lifespan in high-fat diet-fed mice, suggesting both safety and efficacy under chronic administration. These findings suggest that Mitorubin is a promising mitochondrial activator and may represent a novel therapeutic strategy for age-related diseases.

biochemistry↗

bioRxiv2025

Altered laminin signaling destinates melanocyte reprogramming in vitiligo

Vitiligo is an acquired depigmenting skin disorder characterized by progressive melanocyte loss, but the cellular mechanisms driving this process remain unclear. Here, we identify melanocyte dedifferentiation as a central and reversible pathogenic mechanism in vitiligo. In healthy human skin, melanocytes reside within a basement membrane niche defined by dystroglycan-laminin-211 adhesion. In contrast, vitiligo lesions exhibit aberrant extracellular matrix remodeling, leading to an adhesion switch to integrin 3{beta}1-laminin-332 interactions. This shift promotes melanocyte dedifferentiation via Rho-F-actin-dependent activation of Hippo and MAPK pathways, resulting in c-Jun-mediated transcriptional changes. Dedifferentiated melanocytes lose their pigment-producing identity and acquire neural crest-like features with multilineage potential. Importantly, this process is reversible. Pharmacological inhibition of involved pathways restores melanocyte differentiation and induces repigmentation in both vitiligo mouse models and ex vivo patient skin. Notably, JAK inhibitors also promote redifferentiation independently of immune modulation. These findings uncover melanocyte dedifferentiation as a fundamental driver of vitiligo and a tractable therapeutic target, offering new opportunities for therapeutic intervention.

cell biology↗

bioRxiv2024

A new target of multiple lysine methylation in bacteria

The methylation of {varepsilon}-amino groups in protein lysine residues is known to be an important posttranslational modification in eukaryotes. This modification plays a pivotal role in the regulation of diverse biological processes, including epigenetics, transcriptional control, and cellular signaling. Although less studied in prokaryotes, recent research has begun to reveal the potential role of methylation in modulating bacterial immune evasion and adherence to host cells. In this study, we analyzed the cell surface proteins of the toluene-degrading bacterium Acinetobacter sp. Tol 5 by label-free liquid chromatography-mass spectrometry (LC-MS) and found that the lysine residues of its trimeric autotransporter adhesin (TAA), AtaA, are methylated. Over 130 lysine residues of AtaA, consisting of 3,630 amino acids and containing 232 lysine residues, were methylated. We identified the outer membrane protein lysine methyltransferase (OM PKMT) of Tol 5, KmtA, which specifically methylates the lysine residues of AtaA. In the KmtA-deficient mutant, most lysine methylations on AtaA were absent, indicating that KmtA is responsible for the methylation of multiple lysine residues throughout AtaA. Bioinformatic analysis revealed that the OM PKMT genes were widely distributed among gram-negative bacteria, including pathogens with TAAs that promote infectivity, such as Burkholderia mallei and Haemophilus influenzae. Although KmtA has sequence similarities to the OM PKMTs of Rickettsia involved in infectivity, KmtA-like PKMTs formed a distinct cluster from those of the Rickettsia type according to the clustering analysis, suggesting that they are new types of PKMTs. Furthermore, the deletion of Tol 5 KmtA led to an increase in AtaA on the cell surface and enhanced bacterial adhesion, resulting in slower growth. SignificanceMethylation of lysine residues is a posttranslational modification that plays diverse physiological roles in eukaryotes. In prokaryotes however, lysine methylation has been studied only in a limited number of pathogenic bacteria. In this study, we found novel lysine methylation across multiple residues of an outer membrane protein and its methyltransferase, KmtA, in a bacterium from activated sludge. KmtA, along with rickettsial outer membrane protein lysine methyltransferases, which are known to be involved in bacterial pathogenicity, exists in many species of gram-negative bacteria. This finding suggests that methylations are ubiquitous in prokaryotes and are involved in a variety of functions, offering potential strategies for controlling bacterial infections and enhancing the functions of beneficial bacteria for biotechnological applications.

microbiology↗