Biology subjects

Wang, Q.

Publications and source records attributed to Wang, Q..

At least 181 records · Page 10Linked to original sources

bioRxiv2025

Employing the intravesical delivery route to target on the kidney

Background and ObjectiveThere is an unmet demand for kidney-targeted non-invasive drug-delivery systems to enhance the therapeutic index and at the same time reduce extrarenal side effects. Here, we performed a proof-of-concept study to evaluate the validity and efficacy of intravesical delivery for kidney targeting in mice. MethodsProbes with a wide range of molecular sizes were tested for their efficiency to reach the kidney via intravesical delivery in female mice. The therapeutic efficacy and side effects of pazopanib in treating a renal adenocarcinoma were compared between delivery via intravesical route vs. intraperitoneal route. Intravesical delivery of empagliflozin was tested for its action on the proximal tubules. Finally, intrapelvic infusion was examined as another retrograde route for targeting the kidney. Key findings and limitationsIntravesical infusion was valid for a retrograde delivery of molecules up to 500 kDa to the kidney. Empagliflozin, an antagonist of sodium-glucose cotransporter 2 (SGLT2), could efficiently act on the proximal tubules via the intravesical route, and had an action on glucose excretion. In an orthotopic kidney carcinoma model, intravesical delivery of pazopanib was more efficacious in limiting tumor growth, accompanied with much milder adverse effects on extrarenal organs, compared to a systemic delivery when a same dose was administered. This was due to a higher intrarenal drug concentration achieved and a markedly lower level of drug leaked to the blood via intravesical delivery. Conclusions and Clinical ImplicationsThe intravesical delivery route has a great potential to benefit kidney therapeutics and research.

physiology↗

bioRxiv2025

Diminished and altered cellular senescence response in delayed wound healing of aging

The transient upregulation of cellular senescence within wound tissues has been demonstrated to be an important biological process facilitating efficient tissue repair. Dysregulation of this transient wound-induced senescence-like response can result in impaired healing outcomes. Given the established age-related decline in tissue regenerative capacity, we hypothesized that alterations in this senescence response contribute to the delayed healing of cutaneous wounds in aged individuals. Our investigation demonstrated a significant delay in the closure of full-thickness dorsal skin wounds in aged mice compared to their young counterparts. Analysis of the wound microenvironment revealed a transient upregulation of senescence-associated markers (p16, p21, senescence-associated {beta}-galactosidase) and senescence-associated secretory phenotype factors in the wound tissue of young mice, a response that was markedly attenuated in aged mice. Single-cell RNA sequencing analysis of all cells isolated from day 6 wounds identified a distinct population of p16+/p21+/Ki67- senescent fibroblasts in young mice, characterized by a transcriptional signature indicative of pro-healing extracellular matrix production, a finding corroborated in human wound tissue from young donors. Crucially, in aged wounds, we observed a lower quantity of these senescent cells, a deficit compounded by a qualitative, age-dependent shift in their function, moving away from beneficial extracellular matrix remodeling towards a more detrimental pro-inflammatory state, which ultimately can contribute to the delayed wound healing.

cell biology↗

bioRxiv2025

Global profiling of the proteome and acetylome in mice with abdominal aortic aneurysms

BackgroundAbdominal Aortic Aneurysm (AAA) is a life-threatening vascular condition characterized by a progressive dilation of the abdominal aorta and a high risk of rupture. Currently, no effective pharmacological therapies are available, and treatment relies mainly on surgical intervention. The lack of drug targets is largely due to limited understanding of the molecular mechanisms underlying AAA development. This study aims to identify potential therapeutic targets by performing comprehensive proteomic and acetylomic analyses in a mouse model of AAA. MethodsWe conducted global proteomic and acetylomic profiling on abdominal aortic tissues collected from an AAA mouse model. A total of 7,858 proteins and 1,790 acetylated proteins encompassing 4,581 acetylation sites were quantified. Bioinformatics analyses were performed to investigate the regulatory networks and biological pathways associated with these proteins and post-translational modifications. ResultsIntegrated analysis revealed that histone-related proteins were significantly enriched in pathways co-regulated by the proteome and acetylome. These pathways are influenced by histone acetyltransferases and deacetylases, implicating histone acetylation in AAA-related inflammatory and immune processes. Notably, the deacetylases Sirt2 and Sirt5 were identified as potential regulators of neutrophil extracellular trap (NET) formation through suppression of histone acetylation. ConclusionsThis study provides the first comprehensive proteomic and acetylomic dataset for abdominal aortic tissues in a mouse model of AAA, substantially expanding the mouse acetylation database. The results highlight the central role of histone modifications in immune dysregulation and inflammation, and propose Sirt2 and Sirt5 as candidate therapeutic targets that may inhibit NET formation and slow AAA progression. These findings suggest new avenues for drug development targeting the sirtuin family, although further experimental validation and cross-species studies are needed to confirm translational relevance. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=145 SRC="FIGDIR/small/657536v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@1c397a3org.highwire.dtl.DTLVardef@188180org.highwire.dtl.DTLVardef@de1a59org.highwire.dtl.DTLVardef@193e740_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2025

Ultra-high field strength electroporation enables efficient DNA transformation and genome editing in nontuberculous mycobacteria

Efficient DNA delivery is essential for genetic manipulation of mycobacteria and for dissecting their physiology, pathogenesis, and drug resistance. Although electroporation enables transformation efficiencies exceeding 105 CFU per {micro}g DNA in Mycobacterium smegmatis and Mycobacterium tuberculosis, it remains highly inefficient in many non-tuberculous mycobacteria (NTM), including Mycobacterium abscessus. Here, we discovered that NTM such as M. abscessus exhibit exceptional tolerance to ultra-high electric field strengths, and that hypertonic preconditioning partially protects cells from electroporation-induced damage. Using ultra-high electric field strength (3kV/mm) electroporation, we achieved dramatic improvements in plasmid transformation efficiency--up to 106-fold in M. abscessus, 83-fold in Mycobacterium marinum, and 24-fold in Mycobacterium kansasii--compared to standard conditions (1.25 kV/ mm). Transformation efficiency was further influenced by the choice of selectable marker. Ultra-high field strength electroporation also markedly enhanced allelic exchange in M. abscessus expressing Che9c RecET recombinases, increasing recovery of gene deletion mutants by over 1,000-fold relative to conventional electroporation. In parallel, oligonucleotide-mediated recombineering for targeted point mutations produced nearly 10,000-fold more mutants under ultra-high field conditions. Together, these findings establish ultra-high field electroporation as a robust, broadly applicable platform for genetic engineering of NTMs. This method substantially enhances transformation efficiency and enables construction of advanced genetic tools--including expression libraries and CRISPRi knockdown libraries--in species that have historically resisted genetic manipulation. ImportanceInfections caused by non-tuberculous mycobacteria (NTM), including Mycobacterium abscessus, are increasing globally, yet genetic manipulation of these pathogens remains technically challenging due to inefficient DNA delivery and low gene editing success. The ultra-high electric field strength electroporation strategy described here overcomes these barriers, enabling dramatic improvements in both transformation and genome editing efficiency. This advance paves the way for high-throughput functional genomics in NTMs, including the construction of genome-wide knockout, CRISPRi knockdown, and expression libraries. Broad adoption of this approach will accelerate discovery of genetic determinants of virulence and drug resistance, facilitating the development of antimicrobials and vaccines.

microbiology↗

bioRxiv2025

Lost in communication: How Mueller glia cells fail to maintain retinal integrity in USH1C retinal organoids

Usher syndrome type 1, caused by pathogenic variants in the USH1C gene, leads to congenital deafness and progressive retinal degeneration resulting in vision loss. While auditory deficits can be compensated by cochlea implants and hearing aids, no treatment exists to prevent retinal degeneration. Here, we generated retinal organoids from induced pluripotent stem cells of two USH1C patients to elucidate the cellular and molecular mechanisms driving ocular pathogenesis. Single-cell RNA sequencing of both healthy and USH1C retinal organoids identified differential expression of genes related to phototransduction in photoreceptors, as well as alterations in cell adhesion and canonical Wnt signaling in Muller glia cells. Analysis of intercellular communication revealed an overall reduced signaling efficiency, particularly affecting Muller glia-mediated retinal adhesion processes. Morphological characterization of organoids confirmed transcriptome changes by showing degeneration of the outer limiting membrane and loss of adherens junction architecture. Moreover, photoreceptors revealed increased susceptibility to morphological and functional changes related to phototransduction. These results demonstrate that disruption of Muller glia signaling contributes to a loss of retinal integrity, providing novel insights into USH1C pathogenesis and offering targets for therapeutic interventions.

cell biology↗

bioRxiv2025

The SHR/Akr Y chromosome reveals repeated turnover of the rat pseudoautosomal region

Crossing-over between Chr X and Chr Y was first observed 90 years ago, in the brown rat, Rattus norvegicus. However, the sequence of the rat pseudoautosomal region (PAR) has remained a mystery. We produced a near-complete sequence of Chr Y from the SHR strain, along with nearly a megabase of sequence from both telomeres of Chr X. Both telomeric ends of Chr Y display extensive homology to Chr X, but no homology to the ancestral PAR of placental mammals. Using rat Y BACs probes for FISH on meiotic cells, we show that pairing almost always occurs between the tips of Yq and Xp, which are virtually identical in nucleotide sequence, but contain no protein-coding genes. Homology at the other ends of Chr X and Chr Y is likely the result of a recent transposition of five genes from Xq to Yp. These sequences are only 99.5% identical and pair infrequently, but show signs that it may have been pseudoautosomal in the ancestor of the rat genus. The rat Chr Y sequence presents opportunities for experimental studies of meiotic phenomena in a tractable model organism. The short PAR, with a correspondingly high recombination rate, represents a unique substrate for molecular studies of crossing-over. Likewise, the absence of extensively co-amplified testis gene families on the rat X and Y suggests they might serve as control for the intense competition between selfish elements that completely remodeled the mouse sex chromosomes.

genomics↗

bioRxiv2025

Single-cell Characterization of DNA Hydroxymethylation of the Mouse Brain During Aging

DNA methylation dynamics, including 5-hydroxymethylcytosine (5hmC) and 5-methylcytosine (5mC), critically regulate brain function, yet conventional methods cannot distinguish these modifications, obscuring their distinct roles in gene regulation and aging. We present Joint- Cabernet, a bisulfite-free single-cell platform enabling simultaneous profiling of 5hmC, 5mC, and transcriptomes. Applying Joint-Cabernet to 84,071 nuclei from adult and aged mouse brains, we resolved cell-type-specific DNA hydroxymethylation landscapes, revealing elevated 5hmCG and 5hmCH levels in transcriptionally active genes across neuronal subtypes and spatial gradients in cortical layers. During aging, 5hmCG accumulates globally but is selectively enriched at open chromatin loci, aligning with the upregulation of cell-type-specific genes in distinct brain cell types. This single-cell DNA methylation brain cell atlas provides a framework for studying methylation- driven mechanisms in brain aging and neurodegenerative diseases.

genetics↗

bioRxiv2025

Rational design of respiratory syncytial virus dimeric F-subunit vaccines in protein and mRNA forms

BackgroundRespiratory syncytial virus (RSV) poses a significant public health threat, particularly to children and the elderly. Two protein-based vaccines and one mRNA vaccine have been approved, all targeting the prefusion conformation of the fusion (F) glycoprotein. However, it has been reported that the F protein transitions to the post-fusion state during storage, resulting in a reduction of the vaccines immunogenicity. MethodsIn this study, we engineered novel pre-F-based antigens to preserve pre-F-specific immunodominant epitopes while eliminating sub-potent ones. Based on this, we constructed a series of single-chain dimers and selected the one with the highest expression yield and melting temperature (Tm). Next, we created a heterodimer, scDimer AB. Structural and protein characterization analyses were conducted to verify our design. All monomeric and scDimer antigens were used to immunize rodent models. Additionally, we prepared the antigens in mRNA form and immunized BALB/c mice. Finally, we combined both antigen forms, administering intramuscular mRNA priming followed by intranasal protein delivery in mice. In all immunization strategies, viral challenges were performed in animals to evaluate the immunologic protective effects. FindingsThrough rational design, we developed a monomeric and two single-chain dimeric (scDimer) proteins with the expected characteristics, including complete II, V, and O epitopes and a partial III epitope. The scDimers elicited stronger binding and neutralizing antibody responses in rodent models compared to the monomer, and they also boosted T cell responses when combined with appropriate adjuvants. After three doses of scDimer immunization, live RSV was barely detectable in the tissues of infected animals. The copies of RNA encoding N-gene were significantly reduced in the immunized groups compared to the PBS-injected control groups. We also engineered mRNA versions of the antigens and verified their protective efficacy in mice. Notably, there were no significant differences between intranasal boost and intramuscular two doses after RSV challenged, suggesting that intranasal boost provided equivalent protection to intramuscular vaccination and could reduce the risk of vaccine-enhanced disease (VED) potentially. InterpretationThe scDimer-based RSV vaccines effectively protected rodents from RSV infections, highlighting their clinical potential. Our antigen design removed certain suboptimal epitope regions, enhancing the efficiency of antigen presentation and increasing the proportion of the most potent pre-F-specific neutralizing antibodies. This approach provides a novel perspective for future vaccine design. FundingNational Key R&D Program of China, National Science Foundation of China, Young Scientists in Basic Research, Chinese Academy of Sciences, and Special Program of China National Tobacco Corporation. RESEARCH IN CONTEXTO_ST_ABSEvidence before this studyC_ST_ABSThe licensed respiratory syncytial virus (RSV) vaccines were based on RSV pre-F structure. We conducted a PubMed search for articles published in English from databases inception to March 31, 2025, using the search terms "RSV" (interchanged with "respiratory syncytial virus"), "pre-F" (interchanged with "prefusion F") and "vaccine". Our search revealed that most of the articles focus on combining pre-F with RSV G protein or other proteins to form viral-like particles (VLPs) or nanoparticles. The second-largest category of articles examined the antibodies level elicited by the pre-F antigens in animal model or some special cohorts. Other studies addressed topics such as optimization of injection dosage and immunization strategies, methodologies for detecting pre-F-specific antibodies, and the development of mRNA-LNP vaccines. Notably, only 4 articles described novel designs of RSV pre-F antigens, all of which were trimeric and based exclusively on the RSV A subtype F protein sequence. Added value of this studyTo our knowledge, our design is the first single-chain dimer version of RSV pre-F antigen. The engineered antigens demonstrated high yield, thermostability, and immunogenicity comparable to that of DS-Cav1. Additionally, we developed the first chimeric RSV-antigen covering both subtype A and B, thereby broadening the potential protective spectrum. We further validated the immunogenicity of our designs in mRNA form and as a protein-mRNA combination. This research emphasizes the potential of constructing and combining minimized, stable antigen modules, offering a novel approach for future vaccine development. Implications of all the available evidenceIn this study, we present a novel approach for constructing and combining minimized, stable antigen modules for future vaccine design. The strong immunogenicity of our antigens underscores their potential for clinical application. Furthermore, given the structural similarities between the RSV antigen and those of other Pneumoviruses, such as human metapneumovirus (hMPV), our design strategy could be extended to the development of multivalent vaccines.

immunology↗

bioRxiv2025

A CRISPR/Cas9-based system using dual-sgRNAs for efficient gene deletion in Mycobacterium abscessus

The increasing global prevalence of Mycobacterium abscessus infections presents a significant clinical challenge due to the pathogens intrinsic resistance to multiple antibiotics and poor treatment outcomes. Despite the necessity of genetic tools for studying its physiology, pathogenesis, and drug resistance, efficient methods for large-fragment deletions remain underdeveloped. Here, we report a CRISPR/Cas9-based dual-sgRNA system employing Streptococcus thermophilus CRISPR1-Cas9 (Sth1Cas9), enabling efficient large-fragment knockout in M. abscessus with deletion efficiencies exceeding 90% at certain loci and spanning up to 16.7 kb. Furthermore, we systematically optimized the modular arrangement of genetic components in Cas9/dual-sgRNA expression plasmids and refined their construction workflow, achieving a significant reduction in cassette loss rates while enabling single-step plasmid assembly. Notably, deletion efficiency was position-dependent rather than correlated with target size, suggesting an influence of chromatin structure on editing outcomes. As the first CRISPR/Cas9-based platform capable of kilobase-scale deletions in M. abscessus, this system advances functional genomics studies and facilitates targeted investigations into virulence and antibiotic resistance mechanisms.

microbiology↗

bioRxiv2025

Scaled multidimensional assays of variant effect identify sequence-function relationships in hypertrophic cardiomyopathy

BackgroundAn estimated 1 in 500 people live with hypertrophic cardiomyopathy (HCM), a disease for which genetic diagnosis can identify family members at risk, and increasingly guide therapy. Mutations in the myosin binding protein C3 (MYBPC3) gene account for a significant proportion of HCM cases. However, many of these variants are classified as variants of uncertain significance (VUS), complicating clinical decision-making. Scalable methods for variant interpretation in disease-specific cell types are crucial for understanding variant impact and uncovering disease mechanisms. MethodsWe developed a scaled multidimensional mapping strategy to evaluate the functional impact of variants across a critical domain of MYBPC3. We incorporate saturation base editing at the native MYBPC3 locus, a long-read RNA sequencing-enabled assay of variant splice effects, and measurements of HCM-relevant phenotypes, including MYBPC3 abundance, hypertrophic signaling, and ubiquitin-proteasome function in human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). ResultsOur multidimensional mapping strategy enabled high-resolution functional analysis of MYBPC3 variants in iPSC-CMs. Targeted transient base editing generated a comprehensive variant library at the native locus, capturing diverse variant effects on cellular HCM-relevant phenotypes. Our massively parallel splicing assay identified novel splice-disrupting variants. Integration of functional assays revealed that decreased MYBPC3 abundance is a key driver of HCM-related phenotypes. In parallel, downregulation of protein degradation was observed as a compensatory response to MYBPC3 loss of function, and novel disease mechanisms were identified for missense variants near a critical binding domain, underscoring their contribution to pathogenesis. Bayesian estimates of variant effects enable the reclassification of clinical variants. ConclusionsThis work provides a platform for extending genome engineering in iPSCs to multiplexed assays of variant effects across diverse disease-relevant cellular phenotypes, enhancing the understanding of variant pathogenicity and uncovering novel biological mechanisms that could inform therapeutic strategies.

genetics↗

bioRxiv2025

Tau Oligomerization Drives Neurodegeneration via Nuclear Membrane Invagination and Lamin B Receptor Binding in Alzheimer's disease

The microtubule-associated protein tau aggregates into oligomeric complexes that highly correlate with Alzheimers disease (AD) progression. Increasing evidence suggests that nuclear membrane disruption occurs in AD and related tauopathies, but whether this is a cause or consequence of neurodegeneration remains unclear. Using the optogenetically inducible 4R1N Tau::mCherry::Cry2Olig (optoTau) system in iPSC-derived neurons, we demonstrate that tau oligomerization triggers nuclear rupture and nuclear membrane invagination. Pathological tau accumulates at sites of invagination, inducing structural abnormalities in the nuclear envelope and piercing into the nuclear space. These findings were confirmed in the humanized P301S tau (PS19) transgenic mouse model, where nuclear envelope disruption appeared as an early-onset event preceding neurodegeneration. Further validation in post-mortem AD brain tissues revealed nuclear lamina disruption correlating with pathological tau emergence in early-stage patients. Notably, electron microscopy shows that tau-induced nuclear invagination triggers global chromatin reorganization, potentially driving aberrant gene expression and protein translation associated with AD. These findings suggest that nuclear membrane disruption is an early and possibly causative event in tau-mediated neurodegeneration, establishing a mechanistic link between tau oligomerization and nuclear stress. Further investigation into nuclear destabilization could inform clinical strategies for mitigating AD pathogenesis.

neuroscience↗

bioRxiv2025

Inflamed Microglia like Macrophages in the Central Nervous System of Prodromal Parkinson's Disease

We hypothesized that prodromal Parkinsons Disease (PD), characterized by REM Sleep Behavior Disorder and hyposmia, is an inflammatory disorder initiated in the gut that evolves into a neurodegenerative process manifest as classical PD. To test this hypothesis, we performed single-cell RNAseq analysis of cerebrospinal fluid (CSF) and blood from 118 individuals, comparing healthy subjects with prodromal PD and manifest PD to patients with the central nervous system (CNS) autoimmune disease, multiple sclerosis (MS). Surprisingly, we identified increased numbers of immune cells in the CSF of patients with prodromal PD. Single-cell RNA sequencing revealed increases in CSF-specific microglia-like macrophages with enhanced JAK-STAT and TNF signaling signatures in prodromal PD. CSF macrophages exhibited similar transcriptional profiles to dural macrophages and gut muscularis macrophages from -synuclein-expressing PD model mice. Finally, shared memory T cell clones were found in humans between gut and CSF T cells. These findings uncover a myeloid-mediated TNF inflammatory process in the CNS of patients with prodromal PD, with an immunological linkage between the gut and the CNS.

immunology↗

bioRxiv2025

Deep Blood Proteomics Identifies over 12,000 Proteins Providing Valuable Information about the State of the Human Body

Blood is a valuable resource for clinical research, offering insight into physiological and pathological states. However, the specific proteins detectable in blood and the optimal proteomic methods for their detection have not been rigorously investigated and documented. To address this, we conducted various blood proteomic strategies, including directly blood proteomic analysis, high-abundance protein depletion, low-abundance protein enrichment, and extracellular vesicle enrichment using data-independent acquisition or targeted proteomics. These approaches identified 11,679 protein groups in plasma from healthy individuals. In 136 pancreatic ductal adenocarcinoma whole blood samples, 6,956 protein groups were found, including 678 not seen in healthy samples, expanding the total to 12,357 blood proteins. This represents the most comprehensive blood proteome to date. To support broader access and analysis, we developed the Human Blood Proteome (HuBP) database, detailing protein detectability, abundance, and reproducibility across workflows, sample types, and disease contexts.

systems biology↗

bioRxiv2025

Complete360: A Multi-Omics Liquid Biopsy Platform with Attomole Sensitivity and High Reproducibility

Advancements in liquid biopsy platforms can uncovered disease-associated molecular changes that may indicate disease. Despite recent progress in the detection of a number of analytes, the current liquid biopsy platforms struggle with specificity, sensitivity, and reproducibility, limiting their clinical utility and high-throughput applicability. Here, we describe an ultra-sensitive mass spectrometry-based liquid biopsy platform, Complete360, with an optimized library of detection parameters, meticulously refined assays to enable the simultaneous detection and quantification of a broad range of body-fluid proteins, metabolites, and lipids. The system addresses sample-specific and target-specific noises, as well as the micro-environments of proteomics during co-elution, achieving unprecedented levels of reproducibility, specificity, and sensitivity. This platform has facilitated the detection and quantification of numerous multi-omics signatures in human plasma samples, uncovering novel disease mechanisms never identified by previous approaches. Our study establishes a platform for rapid discovery and validation of molecular changes in disease panels, demonstrating its potential utilities in applications meeting clinical regulation guidelines.

systems biology↗

bioRxiv2025

The contribution of the locus coeruleus-norepinephrine system to the coupling between pupil-linked arousal and cortical state

Understanding how pupil-linked arousal couples with cortical state is crucial for uncovering the neural mechanisms underlying brain state-dependent cognitive and sensory processing. Pupil size fluctuations reflect rapid changes of the pupil-linked arousal system, indexing brain states as well as the activity of neuromodulatory systems, including the locus coeruleus-norepinephrine (LC-NE) system. We investigated the relationship among phasic pupil dilation, cortical state, and neuromodulation by combining optogenetic LC stimulation with EEG recordings and pupillometry in awake mice. A comparison between EEG signals during spontaneous phasic pupil dilation and those during phasic pupil dilation evoked by LC stimulation revealed distinct EEG power spectrums, with LC activation driving strong modulation in the alpha and beta bands. Using machine learning techniques, we trained a convolutional neural network classifier to distinguish between types of pupil dilation based on the power dynamics of individual EEG frequency bands. The results confirmed that EEG features in the alpha, beta, and gamma bands differ markedly between spontaneous phasic arousal and LC stimulation-evoked arousal. Moreover, pharmacological manipulations to either block or {beta} adrenergic receptors or agonize -2 adrenergic receptors were employed to explore how adrenergic receptors could influence the coupling between phasic pupil dilation and cortical state. With each manipulation uniquely modulating EEG power and pupil size, our results highlight the differentiated role of adrenergic receptors in the maintenance of coupling between pupil-linked arousal and cortical state. This study provides new insights into the complex relationship between pupil-linked arousal and cortical arousal state, underscoring the significant role of the LC-NE system in influencing these arousal states. Significance StatementThis study reveals the role of the locus coeruleus in pupil-linked arousal coupling with cortical arousal state by uncovering the different relationships between LC stimulation-evoked versus spontaneous phasic pupil dilations and cortical EEG. By integrating machine learning and noradrenergic pharmacological manipulation, our findings highlight the distinct cortical state associated with spontaneous phasic arousal and LC stimulation-evoked arousal, as well as the crucial role of different subtypes of adrenergic receptors in mediating the coupling between pupil size and cortical state.

neuroscience↗

bioRxiv2025

Environmental stress sensitivity determines bacterial Mn(II) oxidation

Bacterial Mn(II) oxidation is a widespread and ecologically significant process, with Mn oxides playing essential roles in biogeochemical cycling and environmental remediation. However, the environmental cues that initiate this microbial activity remain enigmatic. In this study, a spontaneous mutant was isolated from the previously characterized Arthrobacter strain that exhibited autonomous Mn(II)-oxidizing capacity, in contrast to the parental strain, which required cocultivation with other bacteria to initiate Mn(II) oxidation1. Comparative transcriptomic analysis suggested that a gene system encoding an environmental stress-sensing receptor controlled the transcription of boxA, a Mn(II)-oxidizing gene. These findings indicate that two conditions are required for bacterial Mn(II) oxidation: the presence of a Mn(II)-oxidizing gene and an environmentally responsive regulatory system capable of activating its expression. Based on this knowledge, eight additional bacterial strains were isolated that triggered Arthrobacter Mn(II) oxidation. Transcriptomic profiling revealed a robust positive correlation between boxA expression and genes encoding two-component signal transduction systems. Given their widespread distribution (approximately 23% of environmental bacterial genomes harbor Mn(II)-oxidizing gene homologs2) and constant exposure to diverse biotic and abiotic stressors, these bacteria are likely to undergo frequent Mn(II) oxidation triggered by microbial interactions or chemical signals. This study provides critical mechanistic insight into the environmental regulation of bacterial Mn(II) oxidation, addressing a longstanding gap in understanding its natural occurrence.

microbiology↗

bioRxiv2025

Brain-wide cell-type-specific noradrenergic modulation of the transcriptome

Neuromodulatory systems such as the locus coeruleus-norepinephrine (LC-NE) system exert a widespread influence on brain function, yet the transcriptional consequences of such neuromodulatory perturbations remain largely unknown across the many unique cell types in the brain. In this study, we establish a generalizable framework to map brain-wide, cell-type-specific gene expression changes in mice following in vivo chemogenetic activation or inhibition of LC neurons. Single-nucleus RNA sequencing revealed that LC perturbation induces widespread but highly cell type- and region-specific transcriptional program changes, shaped by the distribution of adrenergic receptor subtypes. These findings support a model in which a shared global signal of neuromodulatory tone can produce discrete, context-dependent cellular outcomes through distinct molecular gating mechanisms of cell-type-specific adrenergic receptor subtype combinations. By establishing gene expression as a quantifiable metric of neuromodulatory control, this study lays the foundation for transcriptionally informed interventions capable of modulating brain functions with cellular precision.

neuroscience↗

bioRxiv2025

Integrated Classification of Cortical Cells and Quantitative Projectomic Mapping Unveil Organizational Principles of Brain-Wide Connectomes at Single Cell Level

Molecularly defined cortical cell types have recently been linked to whole neuronal morphology (WNM), particularly those characterized by whole-brain-wide projections, such as intratelencephalic (IT), extratelencephalic (ET), and corticothalamic (CT) neurons. In contrast, classical morphological classifications (e.g., tufted TPC, small tufted SPC, and stellate SSC) are based primarily on local dendrosomatic and axonal structures, especially apical dendrites. This study bridges these perspectives by establishing a new neuronal taxonomy, analyzing the connectomes of defined cortical cell types, and comparing them with those obtained from bulk anterograde injections. Neurons were sparsely labeled via tamoxifen-inducible Cre lines with GFP reporters, and 1,419 WNM cells were comprehensively reconstructed with Vaa3D-TeraVR from [~]15 areas across six functional regions of molecularly labeled brains imaged with 2p-fMOST. These cells were newly classified by integrating current molecular-WNM and classical morphological perspectives, with sample size augmented by 1,455 publicly available WNM cells reconstructed from the Mouse-Light project and CEBSIT. This effort defined ten combined molecular-WNM-classical morphological cell types: L5ET_TPC, L6CT_NPC, L6b_HPC, and seven IT types--L2/3IT_TPC, L4IT_SSC, L4IT_UPC, L4IT_TPC, L5IT_SPC, L6IT_IPC, and L6IT_car3PC. Clustering, quantitative analyses and random Forest classifier objectively validated these types and revealed their distinct connectomes, along with convergent, topographic, and hierarchical organizations across their projection brain regions. At the single-cell level, multiple organizational principles governing cortico-cortical (C-C) and cortico-subcortical (C-subC) connectomes emerged with unprecedented detail, offering a precise GPS-like tool for in vivo recordings and robust datasets for neuronal network modeling. Comparisons with bulk anterograde injection data underscored the limitations of traditional methods in identifying projection targets. Overall, our approach provides significant insights into cortical circuitry and elucidates the complex interplay between neuronal molecular identity, whole morphology, and classical morphological classification.

neuroscience↗