Biology subjects

Ma, J.

Publications and source records attributed to Ma, J..

At least 73 records · Page 4Linked to original sources

bioRxiv2026

Evolutionary Divergence of Structure-Function Coupling between Human and Macaque: Spatial Patterns and Transcriptomic Basis

Structural-functional coupling (SFC) provides critical insights into how the brains structural architecture constrains functional organization for supporting higher-order cognition. Investigating evolutionary differences in SFC between humans and macaques may provide new insights into the neural basis of unique human cognitive abilities. In this study, we analyzed multimodal magnetic resonance imaging data from anesthetized and awake adult rhesus macaques and adult humans to examine cross-species divergences in SFC. Moreover, we integrated transcriptomic data to elucidate the molecular mechanisms underlying evolutionary differences in SFC patterns. We find that humans and macaques exhibit distinct SFC patterns: the human brain shows high SFC in the lateral and medial prefrontal cortex, whereas macaques show high SFC in the sensorimotor cortex. Notably, language-related regions in the human lateral temporal cortex exhibit relatively low SFC. Furthermore, the human whole-brain SFC pattern and the evolutionary differences in SFC between humans and macaques are negatively correlated with cortical evolutionary expansion. By integrating human and macaque transcriptomes, we reveal that the macaque SFC specifically associated genes are primarily involved in basic physiological functions, whereas the human SFC specifically associated genes exhibit evolutionary adaptations in synaptic function, neurotransmitter secretion, and other molecular processes. Moreover, the human-specific genes showing significant overlap with Human Accelerated Regions genes were mainly enriched in cell types of astrocyte and oligodendrocyte and in diseases of schizophrenia and Alzheimers diseases. Overall, these findings advance our understanding of the intricate relationships of SFC in human and macaque brains and provide novel insights in understanding evolutionary conservation and species specificity in cognitive function and gene regulation.

evolutionary biology↗

bioRxiv2026

Uncertainty-aware synthetic lethality prediction with pretrained foundation models

Synthetic lethality (SL) offers a promising paradigm for targeted cancer therapy, yet experimental identification of SL gene pairs remains costly, context-dependent, and biased toward well-studied genes. Existing computational approaches often rely on curated protein-protein interaction (PPI) networks and Gene Ontology (GO) annotations, which limit their ability to generalize to novel genes. Here we introduce CO_SCPLOWILANTROC_SCPLOWO_SCPCAP-C_SCPCAPO_SCPLOWSLC_SCPLOW, a two-stage, graph-free framework that leverages pretrained biological foundation models to predict SL pairs with calibrated uncertainty. In Stage 1, we apply a pretrained single-cell foundation model to bulk RNA-seq profiles of cancer cell lines to obtain context-aware embeddings and perform in silico gene knockouts to generate delta embeddings. These perturbation signals are further conditioned on a data-driven gene prior and supervised with CRISPR viability readouts to learn knockout-aware viability embeddings. In Stage 2, we derive pairwise features from these embeddings and train a lightweight classifier to distinguish SL from non-SL pairs. To enable reliable experimental prioritization, CO_SCPLOWILANTROC_SCPLOWO_SCPCAP-C_SCPCAPO_SCPLOWSLC_SCPLOW incorporates conformal prediction, producing calibrated and interpretable prediction sets that highlight high-confidence SL candidates. Across two evaluation settings, including zero-shot generalization to unseen gene pairs and to unseen genes, ablation analyses show that viability pretraining and the gene prior substantially improve performance while avoiding reliance on PPI and GO features. CO_SCPLOWILANTROC_SCPLOWO_SCPCAP-C_SCPCAPO_SCPLOWSLC_SCPLOW therefore transforms pretrained biological representations into practical, uncertainty-aware hypotheses that support robust and scalable discovery of therapeutic targets.

bioinformatics↗

bioRxiv2026

A regenerative stem cell-derived matrix accelerates functional dermal wound repair in a diabetic model

Despite the growing prevalence of non-healing diabetic wounds, no current treatment options overcome multifactorial deficits in repair. To this end, a mesenchymal stromal cell-derived regenerative extracellular matrix (rECM) was evaluated for the ability to accelerate cutaneous wound repair in leptin receptor-deficient (db/db) diabetic mice with paired full-thickness dorsal skin defects. A single dose of rECM significantly accelerated wound closure compared with vehicle controls. Also, rECM dose-dependently improved overall histological healing scores and modulated granulation tissue dynamics, with the highest dose promoting rapid resolution of granulation tissue relative to wound area. Spatial transcriptomics and immunofluorescence revealed that rECM drove robust formation of de novo peripheral nerve clusters characterized by the Schwann cell marker, p75. The rECM also enhanced vascular maturation in healed wounds, increasing average blood vessel size, smooth muscle actin-positive vessels, and vessel density within myofibroblast-rich regions. In a complementary 3D angiogenic sprouting model, rECM accelerated endothelial invasion and filopodia extension, and at higher concentrations induced contraction of collagen matrices consistent with accelerated resolution of granulation tissue. These data demonstrate that rECM accelerates closure of diabetic skin defects by coordinating faster granulation tissue remodeling with enhanced peripheral nerve formation and vascular maturation.

physiology↗

bioRxiv2026

Type 2 diabetes amplifies environmental Pb toxicity and intergenerational risk

Type 2 diabetes (T2D) and environmental lead (Pb) exposure intersect as prevalent global health challenges. Prior studies report elevated blood Pb in T2D, but the underlying mechanism and directionality remain unresolved. Here we show that T2D fundamentally alters Pb toxicokinetics, converting otherwise modest exposure into progressive organ injury and heritable risk. In a human cohort, elevated blood Pb in T2D specifically associated with impaired renal function rather than glycaemic control or adiposity. In db/db mice, diabetes increased oral Pb absorption through delayed gastrointestinal transit and epithelial barrier dysfunction, leading to enhanced systemic retention. With sustained exposure, renal Pb accumulation induced tubular stress that reduced effective clearance, creating a feed-forward loop in which diabetes promotes Pb retention and retained Pb accelerates kidney injury. Chronic Pb exposure exacerbated diabetic kidney injury without inducing hyperglycaemia, and short-course chelation attenuated renal stress markers. Beyond somatic toxicity, combined paternal T2D and Pb exposure programmed renal and neurobehavioral phenotypes across generations via sperm RNA. These findings redefine Pb from a passive environmental exposure to a context-dependent driver of disease, identify T2D as a vulnerability state that reshapes toxicant handling, and establish a mechanistic link between metabolic disease, environmental exposure and intergenerational risk. This work highlights the need for vulnerability-informed clinical management and environmental regulation in populations burdened by T2D.

genetics↗

bioRxiv2026

MG53 Protects Against Intestinal Inflammation by Inhibiting NLRP3 Inflammasome Activation

Background and ObjectivesInflammatory bowel disease (IBD) involves dysregulated immune responses and chronic intestinal inflammation. The nod-like receptor pyrin domain-containing 3 (NLRP3) inflammasome plays a critical role in IBD pathogenesis, but regulatory mechanisms remain not fully understood. Mitsugumin 53 (MG53, also known as TRIM72), originally identified as a critical membrane repair protein, has emerged as a novel regulator of inflammatory processes. To investigate the protective role of MG53 in colitis and elucidate its mechanisms in regulating NLRP3 inflammasome activation in IBD. MethodsWe used dextran sodium sulfate (DSS)-induced colitis models comparing MG53 knockout (MG53-/-) and wild-type (WT) mice, assessing disease severity, MG53 tissue uptake, and therapeutic effects of recombinant human MG53 (rhMG53). In vitro studies examined rhMG53s effects on NLRP3 inflammasome activation, caspase-1 cleavage, interleukin-1{beta} (IL-1{beta}) secretion, and MG53-NLRP3 interactions. ResultsMG53-/- mice showed more severe colitis with increased weight loss, higher disease activity scores, shortened colons, and greater inflammation. DSS treatment induced the accumulation of circulating MG53 in inflamed colonic tissue. rhMG53 administration ameliorated colitis severity in MG53-/- mice and dose-dependently suppressed NLRP3 inflammasome activation in vitro. MG53 interacted with NLRP3 and reduced apoptosis-associated speck-like protein containing a CARD (ASC) speck formation and NLRP3 oligomerization without affecting upstream signaling or NLRP3 stability. ConclusionMG53 is a physiological regulator of NLRP3 inflammasome activation that protects against colitis, suggesting therapeutic potential for IBD.

immunology↗

bioRxiv2026

MATR3 is essential for oocyte growth and maturation quality through a dual molecular mechanism

The molecular mechanisms governing mRNA accumulation during oocyte growth, essential for developmental competence, remain poorly understood. This study investigates the role of Matrin-3 (MATR3), a highly expressed RNA-binding protein in growing oocytes (GOs), using oocyte-specific knockout mouse models and human oocyte maturation arrest (OMA) samples. The results showed that MATR3 was more abundant in GOs than fully-grown oocytes (FGOs), highly expressed in the nucleus of non-surrounded nucleolus (NSN) oocytes, and exited the nucleus during the NSN-to-surrounded nucleolus (SN) transition. In OMA patients, MATR3 nuclear localization was missed, with smaller oocytes than FGOs. Further, Matr3 deletion in mouse GOs caused restricted oocyte growth, global transcription disorders, follicle development failure, blocked GO-granulosa cell communication (via reduced Gdf9 and Radixin expression), and infertility. Mechanistically, MATR3 regulated transcription by recruiting H3K9me2-demethylating lysine-specific demethylase 3B or binding target gene promoters, like Radixin. These findings reveal a critical role of MATR3 in orchestrating transcription and paracrine signaling during oogenesis and suggest its potential as a diagnostic and therapeutic target for OMA.

physiology↗

bioRxiv2026

Optimization of Agrobacterium-mediated transformation of commercial heirloom tomato cultivars to develop novel traits via CRISPR/Cas9 Genome Editing

Genetic improvement using new genome editing approaches rely on the efficient delivery of the CRISPR/Cas system in the vegetable crop tomato. Previous protocols for tomato transformation have primarily focused on a handful of cultivars (M82, Alisa Craig, Microtom, Sweet-100) with very little commercial relevance, and it is not clear if these protocols can be implemented directly in other commercially relevant varieties. Heirloom tomatoes are sought for their deep and diverse flavor but have not been subjected to systematic crop improvement via conventional breeding or biotechnology approaches such as transgenesis or genome editing. Therefore, we tested the transformation and regeneration capacity of six different heirloom cultivars known for their superior taste and market relevance in the US. Subsequently, we optimized rooting conditions and used the GRF4-GIF1 chimeric developmental regulator to successfully recover transgenic plants. Finally, we evaluated the efficiency of targeted genetic modification using the CRISPR/Cas9 genome editing system in several of these cultivars. We demonstrate that our optimizations led to successful transformation of several heirloom varieties, including the generation of edited plants for target genes modifying plant architecture and flowering time. Our results set the foundation for a biotechnology platform to deliver improved traits to local and regional heirloom varieties using genome editing.

plant biology↗

bioRxiv2026

Ophiuchus-Ab: A Versatile Generative Foundation Model for Advanced Antibody-Based Immunotherapy

Antibodies exhibit extraordinary specificity and diversity in antigen recognition and have become a central class of therapeutics across a wide range of diseases. Despite this clinical success, antibody design remains fundamentally challenging. Antibody function emerges from intricate and highly coupled interactions between heavy and light chains, which complicate sequence-function relationships and limit the rational design of developable antibodies. Here, we reveal that modeling antibody sequence space at the level of paired heavy and light chains is essential to faithfully capture inter-chain dependencies, enabling a deeper understanding of antibody function and facilitating antibody discovery. We present Ophiuchus-Ab, a generative foundation model pre-trained on largescale paired antibody repertoires within a diffusion language modeling framework, unifying antibody generation and representation learning in a single probabilistic formulation. This framework excels diverse antibody design tasks, including CDR infilling, antibody humanization, and light-chain pairing. Beyond generation, diffusion-based pre-training yields transferable representations that enable accurate prediction of antibody properties, including developability, binding affinity, and specificity, even in low-data regimes. Together, these results establish Ophiuchus-Ab as a versatile foundation model for modeling antibodies, providing a foundation for next-generation antibody-based immunotherapy.

bioinformatics↗

bioRxiv2026

Endothelial MRG15 Is a Mechanosensitive Suppressor of Atherosclerosis

Disturbed blood flow induces early endothelial inflammation in atherosclerosis, yet the precise mechanisms of endothelial sensing of disturbed flow remain incompletely understood. In this study, we integrated proteomic profiling of human endothelial cells (ECs) subjected to disturbed flow with coronary artery disease risk related genes from large-scale genome-wide association studies (GWAS). We identified mortality factor 4-like protein 1 (MORF4L1, also called MRG15) as a critical modulator in the pathogenesis of atherosclerosis induced by disturbed flow. We show that the expression of MRG15 is markedly reduced in ECs of human aortic atherosclerotic plaques, as well as in ECs exposed to disturbed flow in mouse carotid artery and in cultured human umbilical vein endothelial cells (HUVECs). Endothelial-specific deletion of Mrg15 significantly worsened, while its overexpression attenuated endothelial inflammation and atherosclerotic lesions in turbulent blood flow- or Western diet-induced mouse atherosclerosis model. Single-cell transcriptomics showed that Mrg15 deficiency increased endothelial inflammation and intercellular adhesion molecule 1 (Icam1) expression, and enhanced integrin-mediated adhesion pathways. Mechanistically, MRG15 facilitated the recruitment of enhancer of zeste homolog 2 (EZH2) to maintain repressive histone H3 lysine 27 trimethylation (H3K27me3) marks on the promoters of ICAM1 and integrin subunit alpha 5 (ITGA5). Disturbed blood flow rapidly led to an elevation of protein neddylation, which subsequently induced neddylation-dependent degradation of MRG15 within endothelial cells. This degradation of MRG15 alleviated the transcriptional repression of ICAM1 and ITGA5, thereby enhancing monocyte adhesion to ECs. These findings highlight endothelial MRG15 as a mechanosensitive suppressor of atherosclerosis induced by disturbed flow. Consequently, MRG15 emerges as a promising novel therapeutic target for atherosclerosis.

molecular biology↗

bioRxiv2026

Targeting autoimmunity in Rheumatoid Arthritis with nanoparticles displaying Liprin-1 peptide

Nanoparticle-based strategies offer a promising tool for inducing antigen-specific immune tolerance across various autoimmune conditions, by acting as master switch to turn-off the autoimmune response. Building on our previous work demonstrating the therapeutic potential of plant-made nanoparticles in rheumatoid arthritis, we present a systematic evaluation of key parameters--including dosing regimen, route of administration, and immunization schedule--to optimize both efficacy and safety. We developed virus-based nanomaterials expressing Tomato Bushy Stunt Virus (TBSV) nanoparticles engineered to display the immunodominant Liprin-1 peptide in the Nicotiana benthamiana plant platform. The mechanisms responsible for the observed protective effects against rheumatoid arthritis were also investigated. Our findings highlight the critical role of repeated intravenous administration and precise dosing in promoting regulatory T cell (Treg) induction and cytokine modulation. Furthermore, we dissected the individual contributions of the Liprin-1 peptide and the viral capsid scaffold in driving immune tolerance. These results support the potential of plant-derived nanoparticles as a versatile and effective platform for antigen-specific immunotherapy, with rheumatoid arthritis serving as a proof-of-concept model for broader applications in the field of autoimmunity. One Sentence SummaryPlant-made nanomaterials induce tolerance in arthritis models via repeated IV dosing, triggering an antigen-specific regulatory immune response.

pharmacology and toxicology↗

bioRxiv2025

Dose-dependent NFI regulation of progenitor lifespan and output underlying human neocortical malformation

Nuclear Factor I (NFI) misexpressions in humans are associated with severe brain malformations, yet the underlying mechanisms remain poorly understood. Here we show that NFIs regulate the broad lineage progression and lifespan of radial glial progenitors (RGPs), and consequently neocortical development in a dose-dependent manner. Human cerebral organoids carrying patient-mimicking NFI mutations exhibit expression level-dependent bidirectional impairments in RGP temporal development, coinciding with patient phenotypes. In mouse models, selective removal of NFIs leads to a dramatic protraction of RGP lineage progression and lifespan, excessive progeny output, and neocortical overgrowth and abnormal folding, whereas overexpression of NFIs accelerates RGP lineage progression, resulting in developmental stage-dependent precocious productions of diverse neural progenies. Moreover, NFIs exhibit a positive auto-regulation and progressive increase in expression, and regulate distinct temporal-specific targets underlying RGP lineage progression. These results suggest that NFIs act as evolutionarily conserved key global temporal regulators of RGP lineage progression and neocortical development. HIGHLIGHTSO_LINFIs affect human RGP temporal development coinciding with patient phenotypes. C_LIO_LINFI removal protracts RGP lineage progression and lifespan with excessive progeny output. C_LIO_LINFI overexpression accelerates RGP lineage progression with precocious progeny output. C_LIO_LINFIs regulate distinct temporal-specific targets underlying RGP lineage progression. C_LI

neuroscience↗

bioRxiv2025

Bivalent CD47 Immunotoxin for Targeted Therapy of Lung Cancer

Lung cancer is the leading cause of cancer-related deaths in the world, with [~]2.5 million people diagnosed and [~]1.5 million deaths each year. While the last two decades have yielded substantial progress with systemic targeted and immune therapies improving treatment responses in subgroups of patients with advanced and refractory lung cancer, there is still a dire unmet clinical need to develop more effective therapies with durable responses. CD47 receptors are overexpressed on the surface of a variety of malignant tumor cells including lung cancer. Although there has been increasing interest in developing targeting antibodies against CD47 for immunotherapy, they failed clinical trials due to their dose-limiting toxicities, primarily hematopoietic toxicity. Using a unique diphtheria toxin resistant Pichia pastoris yeast expression system, we have developed a diphtheria toxin-based bivalent CD47 immunotoxin (bi-CD47-IT) for targeted therapy of CD47+ non-small cell lung cancer (NSCLC). Bi-CD47-IT demonstrated compelling preclinical efficacy in multiple NSCLC cell line-derived xenograft (CDX) and patient-derived xenograft (PDX) mouse models, including subcutaneous, orthotopic, metastatic and humanized models. This study demonstrates the remarkable preclinical activity of bi-CD47-IT against various lung cancer models, making bi-CD47-IT a novel and promising therapeutic approach for NSCLC.

cancer biology↗

bioRxiv2025

HLA micropolymorphisms confine neoantigen conformational adaptability and guide T cell receptor selectivity

T cell receptor (TCR) restriction by highly polymorphic major histocompatibility complex (MHC) proteins is a foundation of cellular immunity. Although the effects of MHC polymorphisms on peptide binding and selection are well established, how micropolymorphisms within MHC supertypes impact immune recognition is poorly understood. Here, we identified a novel mechanism through which the micropolymorphisms in two closely related HLA-A3 superfamily members determine TCR specificity. We previously showed that TCRs specific for a public neoantigen from the PIK3CA oncogene presented by HLA-A*03:01 were unable to recognize the same epitope in the context of HLA-A*03:02. We found here that the micropolymorphisms distinguishing A*03:02 from A*03:01 exert their effect not by altering peptide binding or static structures, but by changing the conformational ensemble of the neoantigen in the groove, preventing it from adopting a conformation compatible with TCR binding. The effect is rooted in how the two polymorphic sites interact with other co-varying, evolutionarily coupled polymorphisms, reflecting a cross-groove network of interactions that controls the conformational adaptability of the peptide/HLA complex. We suggest polymorphism-dependent conformational adaptability reflects an evolved feature of class I MHC proteins that amplifies the impact of peptides in the groove, further diversifying epitopes and contributing to how TCRs and other immunoreceptors differentiate between antigens. Beyond this mechanistic insight, our findings emphasize the need for high-resolution HLA typing in efforts across immunology, including antigen-specific immunotherapy.

immunology↗

bioRxiv2025

Diverse infections transcriptionally reprogram the intestinal epithelium and epithelial-immune cell interactions

The distal small intestine plays vital roles in host physiology by regulating nutrient and fluid homeostasis. Despite being impacted in Crohns disease and a major target for a range of infections, we know relatively little about the complexity of cellular responses and cell-cell communication in the ileum during infection. Single cell and spatial transcriptomics have emerged as powerful technologies to study tissue heterogeneity in the gut, but these tools have focused on the large intestine, in part due to the accessibility of this tissue for biopsies and its importance in cancer. Here we present GutPath, an atlas of over 500,000 single cells with RNA and protein expression profiles for 91 cell states in the ileum across diverse infectious archetypes. We show that GutPath accurately captures established immune responses to infection while revealing pathogen-specific responses in enterocytes. To highlight the discovery potential of this atlas, we identify a novel enterocyte cell state present during Yersinia pseudotuberculosis infection that is spatially linked to bacterial load and tissue pathology. GutPath establishes a much-needed resource for the immunology community that will accelerate the study of the transcriptional diversity of cellular landscapes in the small intestine.

immunology↗

bioRxiv2025

Fourier Helicity Spectra as Quantifiers of Multiscale Chirality

Quantifying the chirality of three-dimensional (3D) objects is crucial for understanding interactions of nanoscale structures, biological molecules, and hierarchical materials. Although chirality is simultaneously present at multiple scales for each case, existing chirality measures rely only on singular scale-specific quantifiers, and some of them yield incorrect chirality assignments for biomolecules. Here we introduce the Fourier Helicity Spectra, a mathematical framework to decompose geometry of chemical and biological structures encompassing the scale dependence of mirror asymmetry. Fourier decomposition of 3D objects into helical harmonics enables one to compute a handedness-resolved characteristics of complex shapes, while reducing computational cost. We show its applicability for a diverse range of chemical systems, from biomacromolecular electron densities to tomographic reconstructions of inorganic nanostructures. Furthermore, inverse transform of isolated spectral components into real space enables visualization of helicity-matched interfaces, identifying how chirality governs biomolecular interactions. We anticipate that this methodology can be generalized with alternative basis sets to encompass a broad range of chiral symmetries governing biological signaling, polarization optics, drug design, and advanced synthesis.

bioinformatics↗

bioRxiv2025

Targeting Putrescine Homeostasis with Lactiplantibacillus plantarum PS128 to Mitigate Autism-like Behaviors in Fmr1 KO Mice

BackgroundAutism spectrum disorder (ASD) is biologically heterogeneous and has limited mechanism-informed interventions targeting core behavioral symptoms. Emerging evidence suggests that gut microbial metabolism shapes neurobehavioral outcomes, yet the specific metabolic pathways linking gut ecology to brain function remain incompletely understood. One candidate pathway is polyamine metabolism, which links microbial amino acid metabolism to host regulation and represents a plausible but underexplored contributor to ASD-related phenotypes. Notably, the psychobiotic Lactiplantibacillus plantarum PS128 has been reported to improve social behaviors in individuals with ASD, although the molecular basis for these effects is unclear. In this study, we used Fragile X mental retardation 1 knockout (Fmr1 KO) mice, a well-established ASD model, to investigate microbiota-dependent metabolic mechanisms underlying autism-like behaviors. ResultsWe found that autism-like behaviors in Fmr1 KO mice were associated with gut dysbiosis, impaired intestinal barrier integrity, disruption of the arginine-ornithine-polyamine pathway, and elevated putrescine in the prefrontal cortex (PFC). Supplementation with PS128 remodeled the gut microbiota, reduced inflammation- and disease-associated taxa, improved intestinal structure and permeability, and restored polyamine homeostasis. Targeted metabolomics revealed an increased PFC-to-serum putrescine ratio in KO mice, which was normalized following PS128 intervention. This correction was accompanied by reduced expression of polyamine transporters in the PFC, including ATP13A family members. Causal experiments supported a functional role for putrescine, as peripheral elevation of putrescine induced autism-like behaviors in wild-type mice, whereas pharmacological inhibition of putrescine synthesis ameliorated behavioral deficits in Fmr1 KO mice. ConclusionsThese findings identify putrescine metabolic dysregulation as a key contributor to autism-like phenotypes and support the existence of an ASD subtype defined by disruption of the arginine-ornithine-polyamine axis. Integrated multi-omics and causal perturbation analyses support a model in which microbiota-targeted intervention rebalances systemic polyamine regulation along the gut-brain axis, thereby improving ASD-relevant behaviors. Our work provides mechanistic evidence linking microbial metabolism to neurochemical homeostasis and highlights the translational potential of metabolic stratification in ASD. Graphical AbstractWe identify dysregulated microbiota-driven polyamine homeostasis as a causal contributor to autism-like behaviors in Fmr1 knockout mice. A psychobiotic intervention restores gut-brain metabolic balance and rescues social and repetitive behavioral deficits through coordinated microbial and host transport regulation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/694366v2_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@e1e780org.highwire.dtl.DTLVardef@125a3baorg.highwire.dtl.DTLVardef@201228org.highwire.dtl.DTLVardef@1880650_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Mining of natural diversity enables efficient and expressible peptide asparaginyl ligases

Peptide asparaginyl ligases (PALs) are powerful tools for protein engineering but are limited by natural rarity and poor expression. We mined 23 cyclotide-rich Viola species, uncovering 29 new PALs that expanded the known repertoire to 47. A dual-objective screen identified VdiPAL1 as the best-performed natural PAL, with twice efficiency of wt-VyPAL2 and 12 mg L-1 soluble expression in E. coli. A broad P2 specificity including Trp/Ile/Leu/Phe/Tyr/Met was discovered across diverse PALs, which enables sequential click-compatible liposome dual-functionalization. 1.8L[A] crystal structure of VdiPAL1 reveals a pre-organized near-attack conformation (NAC), supported by constant-pH MD simulations linking pH-dependent reactivity to NAC geometry. Our homology- and structure-based design yielded VyOpt1, a quintuple mutant of VyPAL2 with over 24-fold improved expression via enhanced cap-domain foldability in a single design-test cycle. This work expands the PAL family and demonstrates a transferable cap-domain-based engineering strategy, highlighting natural diversity as a powerful driver of enzyme discovery and optimization. TOC summaryMining of cyclotide-rich Viola genus expanded the total number of natural peptide asparaginyl ligases (PALs) to 47, including the discovery of highly expressible VdiPAL1. Its high-resolution structure provided new insights for mechanism, and its sequence guided us to a generalized PAL engineering strategy, leading to 24-fold increment in VyPAL2 expression.

biochemistry↗

bioRxiv2025

RBM15 promotes lung adenocarcinoma progression and palbociclib sensitivity via m6A-mediated STIL activation and downstream cyclin D1/CDK4 signaling

RNA-binding motif protein 15 (RBM15), an N6-methyladenosine (m6A) methyltransferase, plays a crucial role in the progression of lung adenocarcinoma (LUAD); however, the underlying mechanisms remain insufficiently defined. In this study, we investigated the contribution of RBM15-mediated m6A modification to LUAD pathogenesis. Our data showed that RBM15 was highly expressed in LUAD tissues and cell lines. Notably, its expression levels were positively correlated with tumor TNM staging and negatively correlated with differentiation status. The overexpression of RBM15 enhanced cell proliferation, migration, invasion, and cell cycle progression, while simultaneously inhibiting apoptosis. Conversely, RBM15 knockdown reversed these phenotypes and induced G2/M-phase arrest. In vivo experiments further confirmed that RBM15 knockdown led to the suppression of tumor growth. RNA sequencing, MeRIP sequencing, and bioinformatics analyses identified STIL as a gene directly regulated by RBM15 through m6A modification, a finding subsequently validated by luciferase reporter assays. Western blot and rescue experiments demonstrated that RBM15 and STIL cooperatively activate the cyclin D1/CDK4 signaling pathway, thereby promoting LUAD progression. However, this effect was markedly attenuated by CDK4/6 inhibitor application. Collectively, these findings indicated that the RBM15-STIL-cyclin D1/CDK4 axis represents a promising therapeutic target for LUAD.

cancer biology↗