Biology subjects

Ma, B.

Publications and source records attributed to Ma, B..

At least 19 recordsLinked to original sources

bioRxiv2026

Transient Partial Inhibition of DNA Polymerase α Promotes Chromatin Plasticity and Enhances Human Cell Reprogramming

Somatic cell reprogramming requires extensive remodeling of epigenomic and transcriptional state, yet the mechanisms that enhance chromatin plasticity during cell-fate conversion remain incompletely understood. Beyond its essential role in genome duplication, DNA replication provides a dynamic window for chromatin remodeling and may therefore influence cellular plasticity during reprogramming. Here, we show that low-dose, transient inhibition of the lagging-strand DNA polymerase (Pol ) complex markedly enhances reprogramming of human peripheral blood mononuclear cells into induced pluripotent stem cells (iPSCs). Fine-tuned inhibition of the catalytic subunit PolA1 increases cell survival under reprogramming-associated stress and enhances cellular resilience to diverse external stresses, preserves genomic stability, and accelerates the acquisition of pluripotent features. PolA1-inhibited cells undergo rapid increases in chromatin accessibility following reprogramming factor delivery and display accelerated progression toward the iPSC state. Consistent with enhanced developmental plasticity, colonies derived from PolA1 inhibitor-treatment display dome-shaped morphology, increased expression of several naive pluripotency markers such as SUSD2. Integrated imaging and single-nucleus multiomic analyses establish transient PolA1 inhibition as a potent regulator of early chromatin remodeling during reprogramming. Functionally, PolA1 inhibitor-derived iPSC lines maintain stable pluripotency and exhibit enhanced differentiation potential, including increased embryoid body budding and accelerated cardiomyocyte differentiation and maturation. Notably, transient and low-dose PolA1 inhibition alters replication-coupled histone incorporation without reducing DNA replication speed or affecting cell cycle progression. This two-step regime primes human PBMCs for sustained gains in reprogramming efficiency with pre- and post-treatment. Together, these findings identify replication-coupled chromatin regulation as an underappreciated mechanism regulating human cell fate plasticity and demonstrate that transient modulation of Pol can simultaneously enhance epigenomic plasticity and preserve genome stability during reprogramming.

molecular biology↗

bioRxiv2026

LysM: a bacterial self-encoded peptidoglycan hydrolase as an antimicrobial and anti-biofilm agent for Gram-negative nosocomial pathogens

Agents with both anti-biofilm property and broad-spectrum antimicrobial activity are urgently needed, especially for drug-resistant nosocomial pathogens. Here, we report a bacterial genome-encoded glycoside hydrolase, named LysM, with lytic activity against multiple Gram-negative nosocomial pathogens and little toxicity to human cells. 3D structural modelling and site-directed mutagenesis indicates that LysM is a peptidoglycan-specific glycoside hydrolase. In combination with EDTA, exogenous LysM induces explosive bacterial cell lysis, exhibits potent bactericidal activity, and eliminates 24 h-old biofilms formed by Gram-negative ESKAPE group pathogens with the highest activity on three nosocomial pathogens, namely Pseudomonas aeruginosa, Acinetobacter baumannii, and Enterobacter cloacae, all of which contain LysM homologs in their genome. More strikingly, LysM supplied in growth media without EDTA, also inhibits the growth of several nosocomial pathogens. Single-cell tracking analysis reveals that LysM completely prevent biofilm formation of P. aeruginosa by reducing bacterial surface-adherence and extending the doubling time. A LysM-based cocktail consisting of LysM, EDTA, and matrix exopolysaccharide-degrading glycosidase PslG, shows potent anti-biofilm activity against P. aeruginosa, including drug-resistant clinical isolates, both in vitro and in human airway organoids (HAOs) biofilm infection model. In addition, LysM enhances the efficacy of many antibiotics from all major classes, including colistin, tobramycin, ciprofloxacin, and tetracycline. Our findings not only provide a clinical candidate for the treatment of nosocomial infections but also open a new path for antimicrobial discovery and anti-biofilm strategies, as homologues of LysM are present in the genomes of many bacteria and phages.

microbiology↗

bioRxiv2026

Proximity-induced protein deglycosylation by endogenous O-GlcNAcase

O-GlcNAcylation is an important post translational modification that regulates numerous cellular processes, yet tools enabling selective removal of O GlcNAc from individual proteins via endogenous O-GlcNAcase (OGA) in living cells remain limited. Here, we report De-O GlcNAcylation-targeting chimeras (DOGTACs), a chemically induced proximity strategy that selectively reduces O GlcNAc from target proteins by recruiting endogenous OGA. Initial designs incorporating potent competitive OGA inhibitors efficiently engaged OGA but failed to induce de-O-GlcNAcylation, revealing that catalytic competence is essential for productive proximity-driven editing. By attenuating inhibitor potency while retaining sufficient OGA engagement, we developed optimized DOGTACs that promote concentration- and time-dependent, target-specific de-O-GlcNAcylation in living cells without perturbing global O-GlcNAc levels. Furthermore, we successfully applied DOGTAC to additional target proteins across multiple cell lines. Collectively, this work established attenuated competitive inhibitors as effective recruitment modules for catalytic enzyme engagement and a novel framework, DOGTAC, for targeted de-O-GlcNAcylation via endogenous OGA recruitment in living cells.

cell biology↗

bioRxiv2026

LncRNA-mediated organization of oncogenic chromatin state is a targetable vulnerability in leukemia

Long non-coding RNAs (lncRNAs) are increasingly recognized as critical regulators of gene expression underlying various cellular functions, however, the functional and mechanistic contributions of most lncRNAs to tumorigenesis remain poorly defined, and targeting of lncRNAs is challenging with conventional therapeutic approaches. Here, we uncover human PAN3-AS1 and its murine ortholog Lnc35682, previously uncharacterized lncRNAs embedded within a conserved syntenic genomic locus, as highly expressed in acute myeloid leukemia (AML). Using genetic mouse models, human cell lines and primary patient samples, we show that PAN3-AS1 is essential for leukemia maintenance but dispensable for normal hematopoiesis. Mechanistically, we find that elevated PAN3-AS1 influences chromatin accessibility, thus promoting leukemia gene expression programs. This is mediated, at least in part, by PAN3-AS1s association with the nuclear lamina through a defined functional region that is required for its leukemogenic function. We further characterize a feed-forward regulatory circuit between PAN3-AS1 and its neighboring gene FLT3 that directly links the aberrant lncRNA functions to the FLT3-mutant AML subtype. To therapeutically exploit the regulatory node, we engineer a myeloid leukemia-preferentially targeted lipid nanoparticle (LNP) formulation and demonstrate effective delivery of siRNAs against endogenous targets into leukemia cells in experimental animals. LNP-siPAN3-AS1 alone or in combination with a clinically used FLT3 inhibitor, Gilteritinib, reduces leukemia burden and significantly delay leukemogenesis in vivo. Overall, our study uncovers a therapeutic vulnerable lncRNA-centric circuitry and provides compelling preclinical evidence for the development and application of a novel RNA targeting-LNP based therapy for treatment of myeloid leukemia. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/744058v1_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@10c9d42org.highwire.dtl.DTLVardef@12864e7org.highwire.dtl.DTLVardef@f87f6aorg.highwire.dtl.DTLVardef@750cbf_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2026

Analysis of the conformational dynamics of amylose oligomers using molecular dynamics simulations

Amylose contributes to starch crystallinity, but the stability of packed amylose double helices in water at elevated temperature remains insufficiently characterized. Here, we used molecular dynamics simulations to test whether chain length affects the short-timescale stability of A-type amylose oligomers in water. Six systems differing in chain length (6, 12, or 24 glucose units per chain) and oligomer size (isolated double strand or dodecamer of six double strands) were simulated, and five independent 1-s production runs were analyzed for each simulated condition. Oligomers with six glucose units showed structural collapse accompanied by increased water penetration. By contrast, dodecamers with 12 or 24 glucose units largely retained packed double-helical organization over the simulated timescale, although fraying was observed at their ends. These results indicate that chain length and lateral packing strongly affect the early structural response of amylose-like crystalline segments in hot water. The present simulations do not establish the ultimate fate of longer oligomers at longer timescales, but they identify a relative stability difference that is relevant to molecular interpretations of hydration-driven disordering in starch.

biophysics↗

bioRxiv2026

ATAD3A structurally links mtDNA replication and mitochondrial fission

Mitochondrial function depends on the maintenance of its genome, and disruptions in copy number and distribution are hallmarks of mitochondrial disorders. Mitochondrial DNA (mtDNA) replication is spatially and temporally linked to mitochondrial division (i.e., fission). However, the signal that coordinates these two events, which are physically separated by the barrier of two mitochondrial membranes, remains unknown. To gain insight into this coordination, we employed correlative cryo-electron tomography (cryo-ET) to analyze the microenvironment surrounding replicating nucleoids. Mitochondrial regions containing replicating mtDNA exhibit a unique membrane architecture defined by the presence of clustered, membrane-spanning tethers that traverse the inner membrane space. Using a combination of superresolution microscopy and genetically encoded cryo-ET tagging technology, we identify these tethers as the AAA+ ATPase ATAD3A. We further show that ATAD3A knockdown reduces recruitment of the mitochondrial fission machinery, whereas overexpression promotes its recruitment and subsequent fission. Our work suggests that ATAD3A forms nanoscale linkages that coordinate these two distinct processes, revealing a new structural paradigm for organellar communication across distinct membrane-defined environments. HighlightsO_LIReplicating mitochondrial DNA (mtDNA) nucleoids are surrounded by a distinct membrane microenvironment. C_LIO_LIATAD3A forms membrane-spanning tethers enriched at replicating mtDNA sites. C_LIO_LIATAD3A enrichment is necessary and sufficient to recruit mitochondrial fission machinery and induce fission at mtDNA replication sites. C_LIO_LIATAD3A structurally couples mtDNA replication state to mitochondrial fission across distinct organellar subcompartments. C_LI

Cell Biology↗

bioRxiv2026

UV inactivation of bacteria and viruses on surfaces: mechanistic insights and testing method comparisons

Germicidal UV devices offer a promising solution to mitigate surface-mediated pathogen transmission, providing effective disinfection without material corrosion. This study evaluated the surface inactivation kinetics of two bacteria and two bacteriophages using a low-pressure (LP) mercury UV lamp (254 nm) and a filtered krypton chloride (KrCl*) excimer lamp (222 nm). Three deposition methods (Spray, Spread, and Pipette) and two extraction methods (Swab and Elute) were compared. The UV dose response on surfaces followed a two-region non-linear model due to shielding from dried deposition constituents, primarily through UV absorption. KrCl* excimer exhibited similar bacterial inactivation but slightly lower viral inactivation than LP UV lamp (maximum inactivation [~] 1 log lower), but its safety profile makes it compelling in occupied spaces. Compared to aqueous conditions, bacteria were more UV sensitive on surfaces, whereas viruses were more resistant. The deposition methods affected the inactivation results, with the Spray method resulting in higher bacteria inactivation. While the extraction methods had limited effect on inactivation efficacy, the Swab method provided higher inactivation detection limits ([~] 2 log higher) and more consistent extraction efficiency. This study provides mechanistic insights into the effects of deposition conditions, UV wavelengths, and microbial characteristics on UV surface disinfection and contributes to standardization of testing methods. TOC Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC="FIGDIR/small/734141v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@12d6656org.highwire.dtl.DTLVardef@199a2f6org.highwire.dtl.DTLVardef@199f168org.highwire.dtl.DTLVardef@1b7f727_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2026

Molecular underpinning of Hydra viridissima-algal facultative symbiosis and vertical algal transmission

The symbiosis between algae and animals represents a relatively recent evolutionary innovation, and exemplified by species in the Cnidaria phylum. Cnidaria can harbor algae within a modified cellular organelle called the symbiosome in a process called endosymbiosis. This animal-algal symbiosis can be facultative or obligate. Algae acquisition occurs either by horizontal transmission, where free-swimming planula gain algae through feeding, or by algae deposition into the developing oocyte in vertical transmission[1-5]. Most studies focus on anthozoans that perform obligate endosymbiosis and transmit algae horizontally. How facultative endosymbiosis in combination with vertical algal transmission impacts the evolutionary adaptation between host and symbiont remains unclear. By studying Hydra viridissima, which performs facultative endosymbiosis and transmits its Chlorella algae vertically, we define different cell types and identify the endoderm cell lineage that gives rise to three major cell types hosting algae. Compared to obligate endosymbiosis[6, 7], Hydra viridissima algal host cells exhibit distinct features, including algal uptake, elevated oxidative phosphorylation and redox activities, and express genes that can provide ammonium for their algal symbionts. We further show when and how the developing Hydra oocytes may take up algae and where oocytes may obtain lipids. Since Hydra is amenable to genetic manipulations, our findings should enable mechanistic studies of how facultative endosymbiosis and vertical transmission evolve and adapt in a changing climate.

molecular biology↗

bioRxiv2026

AlaRS-mediated lactylation shields PARP1 from ubiquitination to promote HNSCC progression

Abstract textMetabolic reprogramming in solid tumors causes massive lactate accumulation, yet how this drives oncogenesis remains incompletely understood. Here, we identify alanyl-tRNA synthetase (AlaRS) as a cellular lactyltransferase that promotes head and neck squamous cell carcinoma (HNSCC) progression. AlaRS directly catalyzes site-specific lactylation of the DNA repair protein PARP1 at lysines K249 and K667. Crucially, we uncover a competitive post-translational crosstalk wherein this lactylation directly antagonizes PARP1 ubiquitination. This "lactyl-shield" prevents proteasomal degradation, hyper-stabilizing the PARP1-Mortalin complex and sustaining tumor proliferation via p53/p21 dysregulation. To therapeutically exploit this mechanism, we identified chelerythrine (CHE) as a potent, selective inhibitor that directly binds the AlaRS catalytic center. CHE abrogates AlaRS lactyltransferase activity, destabilizes PARP1, and robustly suppresses HNSCC xenograft growth in vivo. These findings establish a novel metabolic-post-translational axis linking lactate accumulation to oncoprotein stabilization, providing a blueprint for targeting tRNA synthetase moonlighting functions in cancer.

molecular biology↗

bioRxiv2026

Rewiring c-Myc Transcriptional Activity with an O-GlcNAcylation Targeting Chimera (OGTAC)

c-Myc is a transcription factor that drives tumorigenesis in many cancers. It is notoriously difficult to directly target c-Myc, mainly due to its lack of well-defined druggable pockets. O-linked {beta}-N-acetylglucosamine modification (O-GlcNAcylation) is a post-translational modification (PTM) playing an important role in regulating c-Myc functions in cancer. However, previous studies have primarily relied on global perturbations to investigate c-Myc O-GlcNAcylation, making it difficult to determine its direct functional consequences due to concurrent cellular effects. Here, we report a bifunctional O-GlcNAcylation TArgeting Chimera (OGTAC) molecule, which can induce the proximity of c-Myc and O-GlcNAc transferase (OGT) in living cells, thereby enhancing the O-GlcNAcylation of c-Myc. The c-Myc-targeting OGTAC exhibits anti-proliferation effect against cancer cells. Mapping of c-Myc occupancy on genome indicates that OGTAC rewires c-Myc transcriptional activity and reprograms expression of the downstream oncogene MALAT1, in an O-GlcNAcylation-dependent manner. Overall, OGTAC presents a novel chemically induced proximity (CIP)-based tool to target and rewire c-Myc activity in cancer. Graphic abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=135 SRC="FIGDIR/small/722559v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@d1c640org.highwire.dtl.DTLVardef@2eb70corg.highwire.dtl.DTLVardef@f38970org.highwire.dtl.DTLVardef@c421c8_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2026

Distinct HPO axis responses and ovarian aging trajectories to chronic unpredictable mild stress in reproductively young versus middle-aged female mice

Psychosocial stressors are key contributors to ovarian functional decline. Chronic unpredictable mild stress (CUMS) is widely used to model stress-induced premature ovarian insufficiency (POI) in mice; however, current animal models do not adequately reflect middle-aged women, who represent a key population exposed to chronic psychosocial stress, nor do they capture the dynamic progression toward POI. Here, female C57BL/6 mice aged 2 or 6 months were subjected to CUMS for 8 or 12 weeks. Estrous cyclicity, endocrine profiles, ovarian histology, and transcriptomic changes in HPO axis-related tissues were systematically analyzed. After 8 weeks of exposure, 2-month-old mice exhibited impaired pituitary responsiveness to estradiol negative feedback, as evidenced by dysregulated FSH secretion, indicating reduced stress tolerance compared with 6-month-old mice. Following 12 weeks of CUMS exposure, both age groups showed significant reductions in ovarian size and follicle numbers across all developmental stages. These findings demonstrate that CUMS induces an age-dependent progression toward POI, with short-term exposure eliciting compensatory phases preceding overt ovarian insufficiency, accompanied by distinct endocrine and reproductive alterations and differential responsiveness of the HPO axis. Transcriptomic analyses revealed age-dependent stress responses: ovaries of 2-month-old mice displayed marked activation of inflammatory and immune-related pathways, whereas 6-month-old mice showed sustained upregulation of protein kinase-related signaling networks. Notably, the 6-month-old CUMS model more closely recapitulates stress-associated reproductive aging in adult women. In briefCUMS has been widely used to establish mouse models of psychosocial stress-induced POI. However, current animal models do not adequately reflect middle-aged women, who represent a key population exposed to chronic psychosocial stress, nor do they capture the dynamic progression toward premature ovarian insufficiency (POI). In this study, we demonstrate that different durations of CUMS exposure induce distinct stages of ovarian dysfunction in both young and middle-aged mice, with short-term exposure driving age-dependent compensatory phases and prolonged exposure leading to overt POI, both accompanied by divergent endocrine and reproductive alterations, alongside age-dependent changes in HPO axis responsiveness to CUMS. Notably, the 6-month-old CUMS model shows greater clinical relevance in recapitulating chronic psychosocial stress and stress-related reproductive aging in adult women.

physiology↗

bioRxiv2026

Revisiting phage-host inference from metagenome-assembled genomes reveals the need for scaffold-level validation

Accurate phage-host prediction underpins ecological interpretation and experimental validation of uncultivated phages, yet most current approaches rely heavily on metagenome-assembled genomes (MAGs). This reliance implicitly assumes that MAG-level quality metrics, such as completeness and contamination, ensure scaffold-level correctness. Here, we demonstrate that this assumption is frequently violated. Tetranucleotide frequency (TNF) analyses reveal that CRISPR-associated regions often diverge compositionally from their host genomes, increasing the risk of misbinning, while experimentally validated phage-host genome pairs show consistent TNF separation between phages and their hosts, challenging compositional binning assumptions. Through controlled simulations, we further show that the incorporation of substantial amounts of TNF-matched, non-host phage sequences into high-quality MAGs does not substantially alter standard completeness and contamination estimates, rendering such errors largely undetectable. Together, these results expose a systematic and underappreciated source of bias in MAG-based phage-host inference and highlight the need for explicit scaffold-level validation beyond conventional genome-quality metrics. We further propose a practical scaffold-level validation framework that integrates structural linkage and taxonomic consistency to improve the reliability of phage-host inference from MAGs.

microbiology↗

bioRxiv2026

A Divergent Class of Arylamine N-Acetyltransferases Catalyzes Convergent Amidative Condensation of Polyketides in Manumycins Biosynthesis

Polyketides are prized for their structural complexity and therapeutic potential, yet the incorporation of amide bonds into their frameworks typically relies on linear, nonribosomal peptide synthetase (NRPS)-dependent assembly. The direct, convergent coupling of distinct polyketide chains via amide bond formation--an ideal strategy for combinatorial biosynthesis--has remained largely elusive. Here, we report the discovery of a novel family of arylamine N-acetyltransferases (NATs) from manumycin-type biosynthetic pathways that catalyze an unprecedented intermolecular amidative chain transfer/condensation between an acyl carrier protein (ACP)-tethered polyketide donor and a free polyketide acceptor. Biochemical, structural, and molecular dynamics studies reveal that the representative enzyme, ColC2, possesses a distinctive substrate-binding pocket that diverges from canonical arylamine NATs, conferring exceptional promiscuity toward diverse acyl donors and acceptors. We demonstrate the utility of this biocatalyst by coupling arylamines with either synthetic acyl-thioesters or polyketide synthase (PKS) machinery to generate a library of non-natural polyketide amides and manumycin derivatives. These findings establish a new paradigm for amide bond formation in polyketide biosynthesis and position arylamine NATs as powerful tools for the development of novel therapeutics through combinatorial synthesis.

synthetic biology↗

bioRxiv2026

Nucleotide-dependent Structural Selection Governs c-Src Phosphorylation of Oncogenic KRas4B-G12D

c-Src, the first identified oncogene, and KRas, one of the most frequently mutated proteins in human cancers, can activate each other to promote tumorigenesis. Although normal KRas proteins cycle between the inactive GDP bound and the active GTP bound forms, oncogenic mutants are predominantly GTP bound. Importantly, c-Src preferentially recognizes and phosphorylates the GTP-bound form of KRas but the molecular mechanism underlying this specific recognition remains unknown. Here, we employ molecular simulation tools to identify the mechanism underlying the c-Src recognition of the GTP-loaded state of the G12D mutant of KRas4B (the most prevalent in humans). Combining extensive all-atom Molecular Dynamics simulations and Markov State Models analysis, we found that the most populated states of GTP-bound KRas maintain more open and dynamic switch regions, facilitating easier access of c-Src to the phosphorylation sites of KRas (Tyr32 and Tyr64). These states are sparsely populated for GDP loaded KRas. Docking calculations refined by molecular dynamics identify two c-Src specific regions (residues 340-359 and 453-473) able to stabilize phosphorylation-competent KRas conformations. Therefore, c-Src engages highly populated macrostates of GTP-loaded KRas, while interactions with the GDP form are limited to rare conformations. These KRas conformations selectively recognized by c-Src constitute privileged targets for the rational design of peptide-based or small-molecule inhibitors that specifically target active KRas4B-G12D while sparing the inactive GDP-bound form.

biophysics↗

bioRxiv2026

Engineering armoured in vivo CAR T cells through targeted delivery and transient mRNA gating

In vivo generation of chimeric antigen receptor (CAR) T cells offers an off-the-shelf, scalable alternative to ex-vivo therapies, but is constrained by inefficiency, toxicity and durability. We report the first armoured in-vivo CAR T platform with tunable, transient activity, that enhances safety and efficacy. Our platform utilises CD8 targeted lipid nanoparticles (LNP) to transiently deliver CAR and armouring membrane-bound IL12 mRNAs. Payloads utilize a T-cell-restricted (T-trex) strategy to constrain CAR and IL-12 expression to T cells, enabling potent antigen-dependent activity with spatio-temporal control. Selective targeting of T-cells with a T-trex CAR showed enhanced expression. The addition of T-trex IL12 mRNA provides a tunable, localised amplification of CAR function, enhancing antigen-dependent cytotoxicity and cytokine production with robust tumour control. These findings establish a novel dose-efficient armoured in-vivo CAR T approach and general framework for programmable immune cell engineering.

immunology↗

bioRxiv2026

Development of a BM7G((TKO/hCD46/hCD55/hTHBD/hEPCR) Donor Pig with Endogenous Promoter-Driven Transgenes for Xenotransplantation

Xenotransplantation offers a potential solution to the organ shortage crisis. Multi-gene modification of pigs, such as knockout of three carbohydrate antigen-related genes and expression of immunoprotective proteins, can significantly improve xenograft survival. However, existing strategies face challenges: transposon-based transgenesis may lead to unstable expression, while exogenous promoters used in site-specific integration are susceptible to epigenetic silencing, hindering long-term stable expression. Therefore, developing a donor pig model capable of sustained multi-gene expression is critical. To address this, CRISPR-Cas9 was used to knockout three major glycan antigen genes to eliminate hyperacute rejection. Subsequently, four human protective genes were site-specifically integrated into the porcine Rosa26 safe-harbor locus, with expression driven by the endogenous Rosa26 promoter and the THBD core promoter for long-term stable and tissue-specific expression, and the selection marker was removed using Cre/loxP. Results showed complete absence of the three glycan antigens in BM7G pigs, while the four protective proteins were stably expressed in vascular endothelial cells and major organs. Among them, hCD55 and hCD46 were widely expressed, while hTHBD and hEPCR showed vascular-specific expression. In-vitro assays confirmed that BM7G porcine endothelial cells significantly reduced human antibody binding, effectively inhibited complement-dependent cytotoxicity, and decreased thrombin-antithrombin complex formation. In conclusion, by combining xenoantigen knockout with endogenous promoter-driven expression of multiple human protective genes, a seven-gene modified pig model with low immunogenicity and synergistic protective function was successfully constructed, providing an important donor resource for xenotransplantation preclinical research.

immunology↗

bioRxiv2026

Porcine extended pluripotent stem cell-derived somite-like mesoderm cells with Dox-driven PAX7 are robust seed cell resource for facilitating production of cultured meat

Cultured meat technology, with its significant advantages of shortening meat production cycles, reducing natural resource consumption, minimizing the risk of zoonotic disease transmission, and enabling precise control over nutritional composition and texture, offers a novel alternative source for human meat consumption. One of the major challenges to produce cultured meat in large scale is how to establish high.quality seed cells, which should have long term proliferative capacities and are able to differentiate into muscles efficienuy with simple procedures. Here, we first established an engineered porcine expanded potential stem cells (Tet-On-PAX7 EPSCs) containing Tet-On regulated PAX7 gene. Then the Tet-On-PAX7 EPSCs were induced to somite-liKe mesodermal cells. These somite-liKe mesodermal cells can be expanded over 1025-fold even after 40 passages in-vitro culture while retaining strong myogenic potential. The somite-like mesodermal cells treated with DOX for one day would differentiate into muscle stem cells (Muses), and the later were able to differentiate into muscles with an efficiency of up to 90% within just 7 days in 11-FSDeDa without Dox. Moreover, when somite-liKe mesodermal cells were seeded on patterned scaffolds, microcarrier scaffolds, or cultured in anchorage-independent suspension, they maintained high efficiency in muscle differentiation, confirming their potential to be used as seed cells for scaled cultured meat production.

developmental biology↗

bioRxiv2026

The binding of OTULIN restrains LUBAC activity to prevent TNF-driven immunopathology

Met1-linked ubiquitin chains (Met1-Ub), synthesised by the linear ubiquitin chain assembly complex (LUBAC) and disassembled by the deubiquitinase OTULIN, critically regulate inflammatory signalling. Although OTULINs activity is essential to prevent TNF-driven autoinflammatory pathology and embryonic lethality, the regulatory significance of its direct interaction with LUBAC remains unclear. Here, we reveal that mice harbouring a point mutation (OTULINY56A) in the OTULIN PUB-interacting motif, which disrupts OTULIN-LUBAC interaction, are viable without spontaneous immunopathology. However, OtulinY56A/Y56A mice exhibited hypersensitivity to TNF-induced toxicity, which was not prevented by inhibiting RIPK1 kinase-mediated cell death. Mechanistically, disruption of the OTULIN-LUBAC interaction led to Met1-linked autoubiquitination, which enhanced LUBACs activity and increased Met1-Ub accumulation at the TNF receptor signalling complex. This stabilised the signalling complex even after dissociation from TNF, increased NF-{kappa}B signalling and, contrary to loss of OTULIN or its activity, protected cells from TNF-induced apoptosis. During systemic Listeria monocytogenes infection, the increased response to TNF in OtulinY56A/Y56A mice exaggerated pathology without affecting bacterial burden. Collectively, we identify the physical association of OTULIN to LUBAC as a critical brake that restricts LUBACs function and Met1-Ub-dependent inflammatory signalling, thereby preserving tissue integrity and promoting disease tolerance during acute immune activation.

molecular biology↗