Biology subjects

Zhang, W.

Publications and source records attributed to Zhang, W..

At least 325 records · Page 18Linked to original sources

bioRxiv2024

Molecular Insights into the Regulation of GNPTAB by TMEM251

In vertebrates, newly synthesized lysosomal enzymes traffick to lysosomes through the mannose-6-phosphate (M6P) pathway. The Golgi membrane protein TMEM251 was recently discovered to regulate lysosome biogenesis by controlling the level of GlcNAc-1-phosphotransferase (GNPT). However, its precise function remained unclear. In this study, we demonstrate that TMEM251 is a two-transmembrane protein indispensable for GNPT stability, cleavage by Site-1-Protease (S1P), and enzymatic activity. We reconcile conflicting models by showing that TMEM251 enhances GNPT cleavage and prevents its mislocalization to lysosomes for degradation. We further establish that TMEM251 achieves this by interacting with GOLPH3 and retromer complexes to anchor the TMEM251-GNPT complex at the Golgi. Alanine mutagenesis identified F4XXR7 motif in TMEM251s N-tail for GOLPH3 binding. Together, our findings uncover TMEM251s multi-faceted role in stabilizing GNPT, retaining it at the Golgi, and ensuring the fidelity of the M6P pathway, thereby providing insights into its molecular function.

cell biology↗

bioRxiv2024

E-box independent chromatin recruitment turns MYOD into a transcriptional repressor

MYOD is an E-box sequence-specific basic Helix-Loop-Helix (bHLH) transcriptional activator that, when expressed in non-muscle cells, induces nuclear reprogramming toward skeletal myogenesis by promoting chromatin accessibility at previously silent loci. Here, we report on the identification of a previously unrecognized property of MYOD as repressor of gene expression, via E-box-independent chromatin binding within accessible genomic elements, which invariably leads to reduced chromatin accessibility. MYOD-mediated repression requires the integrity of functional domains previously implicated in MYOD-mediated activation of gene expression. Repression of mitogen-and growth factor-responsive genes occurs through promoter binding and requires a highly conserved domain within the first helix. Repression of cell-of-origin/alternative lineage genes occurs via binding and decommissioning of distal regulatory elements, such as super-enhancers (SE), which requires the N-terminal activation domain as well as two chromatin-remodeling domains and leads to reduced strength of CTCF-mediated chromatin interactions. Surprisingly, MYOD-mediated chromatin compaction and repression of transcription do not associate with reduction of H3K27ac, the conventional histone mark of enhancer or promoter activation, but with reduced levels of the recently discovered histone H4 acetyl-methyl lysine modification (Kacme). These results extend MYOD biological properties beyond the current dogma that restricts MYOD function to a monotone transcriptional activator and reveal a previously unrecognized functional versatility arising from an alternative chromatin recruitment through E-box or non-E-box sequences. The E-box independent repression of gene expression by MYOD might provide a promiscuous mechanism to reduce chromatin accessibility and repress cell-of-origin/alternative lineage and growth factor/mitogen-responsive genes to safeguard the integrity of cell identity during muscle progenitor commitment toward the myogenic lineage.

molecular biology↗

bioRxiv2024

C2AM-Unet: Coordinate and Channel Attention Mixing Flexible Architecture for Retinal Vessel Segmentation

Accurate retinal Microvascular segmentation demands a balanced combination of anatomical fidelity and hemodynamic relevance. However, existing methods fall short in preserving critical structures such as capillary junctions and bifurcations, thus limiting clinical applications and causing fragmentation. To address these limitations, we propose DFMS-Net, a novel dualfield segmentation framework that synergistically integrates geometric-field modeling--achieved through the Spatial Pathway Extractor (SPE) and Transformer-based Topology Interaction (TTI) for preserving structural continuity--and functional-field optimization--realized by the Semantic Attention Amplification (SAA) module for enhancing semantic visibility--via a unified Dual-Field Hemo-dynamic Attention (DFHA) mechanism. This core module enables joint enhancement of vessel continuity, accurate resolution of complex branching patterns, and recovery of low-contrast capillaries, all under physiological guidance. By co-optimizing geometric and functional cues within a unified attention learning paradigm, DFMS-Net produces segmentations that are both morphologically accurate and hemodynamically plausible. Furthermore, we propose two specialized variants using a streamlined Double SPE Attention for vessel continuity refinement. To address the directionality-dependent nature of structural damage in retinal ischemia and glaucoma, we propose Variant-1--a streamlined architecture that emphasizes dual-stage directional refinement to enhance trajectory coherence and improve the detection of topological disruptions, while Variant-2 supports high-resolution analysis of capillary dropout in early diabetic retinopathy through detailed microvascular recovery. Extensive experiments on retinal (DRIVE, STARE) and coronary angiography (DCA1, CHUAC) datasets demonstrate that DFMS-Net achieves state-of-theart performance. Meanwhile, its strong generalization capability offers a promising foundation for diagnosing both retinal and cardiovascular diseases. The code will be available at https://github.com/699zjl/DFMS-Net-new.

bioinformatics↗

bioRxiv2024

Dynamic interplay between RNA N6-methyladenosine modification and porcine reproductive and respiratory syndrome virus infection

N6-methyladenosine (m6A) has attracted significant attention for its role in various biological processes, including RNA stability, translation, and the immune response. Understanding the role of m6A in viral infections is crucial to deep the complex interaction between virus and host cells. Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) is a significant pathogen affecting swine health worldwide. Here, we firstly identified the m6A peaks in the PRRSV genome by m6A RNA immunoprecipitation sequencing (m6A-seq). Seven m6A-enriched regions within the PRRSV genome were detected, with one located in the N protein-coding region and the others distributed across nonstructural protein-coding regions. Notably, the Nsp2-coding region contained the highest m6A peak, spanning approximately 178 nucleotides. Functional analyses demonstrated a positive correlation between m6A modification levels and PRRSV replication in porcine alveolar macrophages (PAMs), as modulating the expression of m6A methyltransferases and demethylases significantly influenced viral replication. Moreover, treatment with the universal methylation inhibitor 3-deazaadenosine (3-DAA) effectively suppressed PRRSV replication, suggesting its potential as a novel anti-PRRSV therapeutic. To further elucidate the role of m6A in PRRSV infection, we analyzed the m6A landscape in PAMs infected with pandemic and highly pathogenic PRRSV strains. Among the 4,677 transcripts exhibiting altered m6A modification levels, the MAPK14 gene as well as other genes in p38/MAPK signaling pathway potentially emerged as the preliminary targets of m6A-mediated epigenetic regulation during PRRSV infection. These findings provide new insights into the epigenetic mechanisms underlying PRRSV infection and may facilitate the development of targeted anti-PRRSV strategies and therapeutics. IMPORTANCEThe involvement of m6A in viral genome has broader implications for our understanding of virus evolution and pathogenicity. The ability of viruses to adapt to their hosts and exploit host cellular mechanisms is a critical factor in their success as pathogens. As m6A is a modification that can be manipulated by both host and virus, studying its role in viral infections may reveal new insights into viral evolution. Understanding how different viruses utilize m6A could inform strategies for developing antiviral therapies that target these interactions. Such therapies could disrupt the ability of viruses to exploit m6A modifications, potentially reducing their virulence and enhancing host defenses.

microbiology↗

bioRxiv2024

Molecular epidemiology of carbapenem-resistant hypervirulent Klebsiella pneumoniae: Risk factors and resistance mechanism of ceftazidime/avibactam in China

The high morbidity and mortality rates associated with carbapenem-resistant hypervirulent Klebsiella pneumoniae have become a global public health problem. We analyzed the whole genome sequence and epidemiological data of 81 carbapenem-resistant Klebsiella pneumoniae strains and conducted sensitivity experiments to explore the optimal concentration of carbapenem-resistant Klebsiella pneumoniae. We verified the resistance and virulence genes of carbapenem-resistant Klebsiella pneumoniae using polymerase chain reaction and compared virulence between carbapenem-resistant hypervirulent Klebsiella pneumoniae a ndcarbapenem-resistant-non-hypervirulent Klebsiella pneumoniae through Galleria mellonella assays.The prevalent types of multilocus sequence typing in our carbapenem-resistant Klebsiella pneumoniae strains were ST11 (69.14%, 56/81). ST11-K64-O1/O2v1 has become an epidemic type in our hospital, with drug-resistance genes dominated by blaKPC-2 (90.12%, 73/81). Ceftazidime/avibactam has better antibacterial effects on carbapenem-resistant Klebsiella pneumoniae, and its resistance is mainly mediated by a resistance plasmid containing the blaKPC-2 gene. 52 carbapenem-resistant hypervirulent Klebsiella pneumoniae strains carry the blaKPC-2, with a high detection rate of siderophore. Further research has shown that the genetic environment of the blaKPC-2 contains TnpR_Tn3 and ISKpn27 upstream and ISKpn6 insertion sequences downstream. Galleria mellonella revealed that the survival rate of carbapenem-resistant hypervirulent Klebsiella pneumoniae was lower than that of carbapenem-resistant-non-hypervirulent Klebsiella pneumoniae, and the survival rate of carbapenem-resistant hypervirulent Klebsiella pneumoniae-K64 was lower than that of carbapenem-resistant hypervirulent Klebsiella pneumoniae-K47.The spread of carbapenem-resistant hypervirulent Klebsiella pneumoniae strains is alarming, necessitating molecular monitoring of hypervirulent strains and strengthening the nosocomial infection prevention and control measures. Antibiotics should be used reasonably based on patients infection status and local epidemiological data. ImportanceCarbapenem-resistant Klebsiella pneumonia is a global public health concern. We conducted a molecular epidemiological analysis of 81 carbapenem-resistant hypervirulent Klebsiella pneumoniae strains from our hospital and explored the resistance mechanism of ceftazidime/avibactam. We identified the blaKPC-2 gene to be primarily responsible for carbapenem resistance in our hospital, and ST11-K64 was the dominant clone. A high-risk clone, ST307-K102, was detected, necessitating strengthened surveillance to prevent its spread. This studys findings contribute to the optimization of antimicrobial management and indicate that genomic research can assist with the tracking of hospital infection outbreaks and the improvement of infection-control measures.

microbiology↗

bioRxiv2024

Deep Learning Unravels Potential Antibiotic-Resistance Drugs against Klebsiella Pneumoniae

BackgroundKlebsiella pneumoniae (KP) is a critical pathogen recognized by the World Health Organization due to its high mortality rates and resistance to antibiotics. Developing effective treatments for KP is essential to mitigate the growing threat of multidrug-resistant infections. This study leverages deep learning techniques to identify and validate potential drug candidates against KP, accelerating the drug discovery process and reducing associated risks. ResultsA comprehensive dataset was constructed, comprising 3,475 drugs selected from the DrugBank database, filtered for their potential efficacy and safety. Using convolutional neural networks (CNNs) for drug-drug interaction analysis, a prediction accuracy of 72% was achieved. Evolutionary Scale Modeling (ESM) was employed to calculate molecular similarities between drugs and KP strains, identifying five promising candidates with structural similarities exceeding 85% to known KP targets. A drug repurposing knowledge graph (DRKG) further validated these findings, with the PairRE model outperforming other approaches in ranking potential drug candidates. ConclusionsThis study presents a scalable and efficient framework for identifying and validating drug candidates to combat antibiotic-resistant KP. By integrating deep learning models, molecular similarity analysis, and knowledge graph techniques, this approach shortens the drug discovery timeline from years to months. The identified candidates hold significant promise for preclinical and clinical testing, offering a pathway to effective therapies against KP and other multidrug-resistant pathogens.

bioinformatics↗

bioRxiv2024

Intranasal dantrolene nanoparticles inhibit inflammatory pyroptosis in 5XFAD mice brains

BackgroundThis study investigates the effects of intranasal dantrolene nanoparticles on inflammation and programmed cell death by pyroptosis in 5XFAD Alzheimers Disease (AD) mice. Methods5XFAD and wild type (WT) B6SJLF1/J mice were treated with intranasal dantrolene nanoparticles (5 mg/kg), daily, Monday to Friday, for 12 weeks continuously, starting at 9 months of age. Blood and brain were harvested at 13 months of age, one month after completion of 12 weeks intranasal dantrolene nanoparticle treatment. Blood biomarkers function of liver (Alanine transaminase, ALT), kidney (Creatinine), and thyroid (TSH: Thyroid-stimulating hormone) were measured using ELISA. The changes of whole brain tissue proteins on Ca2+ release channels on membrane of endoplasmic reticulum (type 2 ryanodine and type 1 InsP3 receptors, RyR-2 and InsP3R-1), lipid peroxidation byproduct malondialdehyde (MDA)-modified proteins, 4-HNE, pyroptosis regulatory proteins (NLR family pyrin domain containing 3 (NLRP3), cleaved caspase-1, full length or N-terminal of Gasdermin D (GSDMD), cytotoxic (IL-1, IL-18, IL-6, TNF-a) and cytoprotective (IL-10) cytokines, astrogliosis (GFAP), microgliosis (IBA-1) and synapse proteins (PSD-95, Synapsin-1) were determined using immunoblotting. Body weights were monitored regularly. ResultsIntranasal dantrolene nanoparticles significantly inhibited the increase of RyR-2 and InsP3R-1 proteins, MDA-modified proteins, 4-NHE, pyroptosis regulatory proteins (NLRP3, cleaved caspase-1, N-terminal GSDMD), cytotoxic cytokine (IL-1{beta}, IL-18, IL-6, TNF-), biomarkers for astrogliosis (GFAP) and microgliosis (IBA-1), and the decrease of cytoprotective cytokine (IL-10) and synaptic proteins (PSD-95, synpasin-1). Intranasal dantrolene nanoparticles for 12 weeks did not affect blood biomarkers for function of liver, kidney, and thyroid, not did it change body weight significantly. ConclusionIntranasal dantrolene nanoparticles significantly inhibit the increase of RyR-2 and InsP3R-1 Ca2+ channel receptor proteins, ameliorate activation of the pyroptosis pathway and pathological inflammation, and the associated loss of synapse proteins. Intranasal dantrolene nanoparticles for three months did not affect liver, kidney and thyroid functions or cause other side effects.

neuroscience↗

bioRxiv2024

Shroom3-Rock interaction and profibrotic function: Resolving mechanism of an intronic CKD risk allele.

Common intronic enhancer SNPs in Shroom3 associate with CKD in GWAS, although there is paucity of detailed mechanism. Previously, we reported a role for Shroom3 in mediating crosstalk between TGF{beta}1- & Wnt/Ctnnb1 pathways promoting renal fibrosis (TIF). However, beneficial roles for Shroom3 in proteinuria have also been reported suggesting pleiotropic effects. Here we focused on identifying the specific profibrotic Shroom3 motif. Given known therapeutic roles for Rho-kinase inhibitors in experimental CKD, and the established interaction between Shroom3 and Rock via its ASD2 domain, we hypothesized that Shroom3-mediated ROCK activation played a crucial role in its profibrotic function in high expressors. To test this hypothesis, we developed transgenic mice and cell lines that inducibly overexpressed wild-type- (WT-Sh3) or ASD2-domain deletion- Shroom3 (ASD2{Delta}-Sh3). Prior scRNAseq data showed that during TIF, Shroom3 and Rock co-expression occurred in injured tubular cells and fibroblasts, highlighting cell-types where this mechanism could be involved. Using HEK293T cells, we first confirmed absent ROCK binding and inhibited TGF{beta}1-signaling with ASD2{Delta}-Sh3-overexpression vs WT-Sh3. In mIMCD cells, ASD2{Delta}-Sh3 overexpression, reduced Rock activation (phospho-MYPT1), pro-fibrotic and pro-inflammatory transcripts vs WT-Sh3. Fibroblast proliferation (3T3) was also reduced with ASD2{Delta}-Sh3. In vivo, we studied ureteric obstruction (UUO) and Aristolochic nephropathy (AAN) as TIF models. In AAN, inducible global-, or Pan-tubular specific-, WTSh3-overexpression showed increased azotemia, and TIF vs ASD2{Delta}-Sh3 mice. WT-Sh3 mice consistently showed significant enrichment of Rho-GTPase, TGF{beta}1- and Wnt/CtnnB1- signaling in kidney transcriptome, paralleling Shroom3-coexpressed genes in tubulo-interstitial transcriptomes from human CKD. In UUO, again WT-Sh3 mice recapitulated increased fibrosis vs ASD2{Delta}-Sh3. Importantly, ASD2{Delta}-Sh3 did not develop albuminuria vs WT-Sh3, while mutating a disparate Fyn-binding Shroom3 motif induced albuminuria in mice, suggesting motif-specific roles for Shroom3 in the kidney. Hence, our data show a critical role for the Rock-binding, ASD2-domain in mediating TIF in milieu of Shroom3 excess, with relevance to human CKD.

systems biology↗

bioRxiv2024

Spatially Resolved in vivo CRISPR Screen Sequencing via Perturb-DBiT

Perturb-seq enabled the profiling of transcriptional effects of genetic perturbations in single cells but lacks the ability to examine the impact on tissue environments. We present Perturb-DBiT for simultaneous co- sequencing of spatial transcriptome and guide RNAs (gRNAs) on the same tissue section for in vivo CRISPR screen with genome-scale gRNA libraries, offering a comprehensive understanding of how genetic modifications affect cellular behavior and tissue architecture. This platform supports a variety of delivery vectors, gRNA library sizes, and tissue preparations, along with two distinct gRNA capture methods, making it adaptable to a wide range of experimental setups. In applying Perturb-DBiT, we conducted un-biased knockouts of tens of genes or at genome-wide scale across three cancer models. We mapped all gRNAs in individual colonies and corresponding transcriptomes in a human cancer metastatic colonization model, revealing clonal dynamics and cooperation. We also examined the effect of genetic perturbation on the tumor immune microenvironment in an immune-competent syngeneic model, uncovering differential and synergistic perturbations in promoting immune infiltration or suppression in tumors. Perturb-DBiT allows for simultaneously evaluating the impact of each knockout on tumor initiation, development, metastasis, histopathology, and immune landscape. Ultimately, it not only broadens the scope of genetic inquiry, but also lays the groundwork for developing targeted therapeutic strategies.

genomics↗

bioRxiv2024

SARM1 Activation Promotes Axonal Degeneration Via a Two-Step Liquid-to-Solid Phase Transition

SARM1 protein plays a central role in axonal degeneration, a key process in many neurodegenerative diseases and nerve injuries. It mediates this by depleting axonal NAD+ through its NADase activity, catalyzed by the TIR domain. Normally, this activity is kept in an inactive state and becomes activated in response to various neuronal damage signals. However, the molecular mechanism behind SARM1 activation, particularly how activation is restricted to damaged axons, remains to be fully elucidated. In this study, using a class of pyridine-containing compounds that induce SARM1-dependent cell death and axonal degeneration, we reveal a two-step process of SARM1 activation. The first step involves sub-lethal "priming" activation of SARM1 mediated by the NAD+ precursor NMN, leading to the formation of covalent conjugates between the hydrolyzed product of NAD+, adenosine diphosphate ribose (ADPR), and the compounds catalyzed by the intrinsic base exchange activity of SARM1. In the second step, these ADPR-conjugates act as molecular glues, promoting the formation of self-proliferating superhelical filaments. On these filaments, SARM1s TIR domains adopt an active NADase configuration. As the superhelical filaments rapidly reach their limit of solubility, they precipitate out of the liquid phase as condensates with stable, fully activated NAD hydrolysis activity. Interestingly, we found that a series of reported SARM1 inhibitors currently under clinical development, which target the TIR enzymatic domain, can paradoxically activate SARM1s NADase activity via this mechanism. These findings provide new insights into how SARM1 activation is spatially restricted to damaged axons and offer important implications for the development of therapeutics targeting SARM1.

molecular biology↗

bioRxiv2024

PanTax: Strain-level taxonomic classification of metagenomic data using pangenome graphs

Microbes are omnipresent, thriving in a range of habitats from oceans to soils and even within our gastrointestinal tracts. They play a vital role in maintaining ecological equilibrium and promoting the health of their hosts. Consequently, understanding the strain diversity within microbial communities is crucial, as variations between strains can lead to distinct phenotypic expressions or diverse biological functions. However, current methods for taxonomic classification from metagenomic sequencing data have several limitations, including their reliance solely on species resolution, support for either short or long reads, or their confinement to a given single species. Most notably, the majority of existing taxonomic classifiers rely solely on a single linear representative genome as a reference, which fails to capture the strain diversity, thereby introducing single-reference biases. Here, we present PanTax, a pangenome graph-based taxonomic classification method that overcomes the shortcomings of single-reference genome-based approaches, because pangenome graphs possess the capability to depict the genetic variability present across multiple evolutionarily or environmentally related genomes. PanTax provides a comprehensive solution to taxonomic classification for strain resolution, compatibility with both short and long reads, and compatibility with single or multiple species. Extensive benchmarking results demonstrate that PanTax drastically outperforms state-of-the-art approaches, primarily evidenced by its significantly higher precision or recall (at both species and strain levels), while maintaining comparable or better performance in other aspects across various datasets. PanTax is a user-friendly open-source tool that is publicly accessible at https://github.com/LuoGroup2023/PanTax.

bioinformatics↗

bioRxiv2024

Exploring methanogenic archaea and their thermal responses in the glacier-fed stream sediments of Rongbuk River Basin, Mt. Everest

Glacier-fed streams (GFS) are emergent sources of greenhouse gas methane, and methanogenic archaea in sediments contribute largely to stream methane emissions. However, little is known about the methanogenic communities in GFS sediments and their key environmental driving factors. This study analyzed stream sediments from the Rongbuk River basin on Mount Everest for methanogenic communities and their temperature responses through anaerobic microcosm incubations at 5{degrees}C and 15{degrees}C. Diverse methanogens were identified, including acetoclastic, hydrogenotrophic, and methylotrophic types. Substantial methane and CO2 production were detected across altitudes and increased significantly at 15{degrees}C, with both methane and CO2 production rate negatively correlated with altitude. While temperature sensitivity of CO2 production but not CH4 showed a negative altitude correlation. Methanogens increased substantially over long-term incubation, dominating the archaeal community. At 15{degrees}C, the relative abundance of several methanogenic groups was strongly correlated with altitude, with positive correlations observed for Methanomassiliicoccaceae and Methanoregulaceae, and negative correlations for Methanocellaceae and Methanotrichaceae, respectively. Besides altitude, factors like phosphorus, C-to-N ratio, and pH also affected methanogenic structure, methane and CO2 production, and temperature sensitivities. This study offers new insights into methanogens and methane production in GFS sediments, improving our understanding of GFS carbon cycling and its potential responses to climate change.

ecology↗

bioRxiv2024

A Full Window Data Independent Acquisition Method for DeeperTop-down Proteomics

Top-down proteomics (TDP) is emerging as a vital tool for the comprehensive characterization of proteoforms. However, as its core technology, top-down mass spectrometry (TDMS) still faces significant analytical challenges. While data-independent acquisition (DIA) has revolutionized bottom-up proteomics and metabolomics, they are rarely employed in TDP. The unique feature of protein ions in an electrospray mass spectrum, as well as the data complexity require the development of new DIA strategies. This study introduces a machine learning assisted Full Window DIA (FW-DIA) method that eliminates precursor ion isolation, making it compatible with a wide range of commercial mass spectrometers. Moreover, FW-DIA leverages all precursor protein ions to generate high-quality tandem mass spectra, enhancing signal intensities by [~]50-fold and protein sequence coverage by threefold in a modular protein analysis. The method was successfully applied to the analysis of a five-protein mixture under native conditions and Escherichia coli ribosomal proteoform characterization.

bioinformatics↗

bioRxiv2024

Deep Learning with Evolutionary Scale Modeling to discover Antibiotic-Resistance Drug against Klebsiella Pneumoniae

The development of effective antibiotics is crucial in combating antibiotic-resistant pathogens such as Klebsiella pneumoniae (KP). This study utilizes deep learning, specifically Evolutionary Scale Modeling (ESM), to identify potential drug candidates. By analyzing molecular similarities between known drugs and KP strains, we aim to accelerate the discovery of new treatments and reduce the risks associated with clinical trials. This approach may shed new lights in the field of drug repositioning.

bioinformatics↗

bioRxiv2024

A regulator controls both DNA damage response and anti-phage defense networks in Moraxellaceae

DNA damage chemicals, including many antibiotics, often induce prophage induction and phage outbreaks in microbial communities, a major threat to bacterial survival. Moraxellaceae contains clinically highly relevant strains with outstanding antibiotic and radio resistance, yet their cellular-level regulation in DNA damage response and anti-phage defense is largely unknown. Here, we identified a WYL family protein that had replaced the ubiquitous SOS system in evolution and directly regulated a transcriptional network of both DNA repair genes and anti-phage defense genes under DNA damage stress. This study depicts a mechanism that how bacteria maintain immunity to phages without compromising on antibiotic resistance, and sheds light on controlling the resistance of Moraxellaceae strains in clinical practice.

microbiology↗

bioRxiv2024

AI Assisted Native Proteomics: Delineating Ribosomal Protein Conformations Pre- and Post-Assembly

The simultaneous identification of proteins as well as their conformations in a biological system would greatly enhance our understanding of cellular mechanisms and disease. As an emerging technique, native proteomics analyzes proteins in their native states, facilitating the acquisition of protein stoichiometry, post-translational modifications (PTMs), and interactions with ligands. However, revealing protein conformations at the proteome scale remains a significant challenge. In this study, we propose an AI-assisted native proteomics method that integrates a protein structure prediction (PSP) module with top-down proteomics (TDP) and native mass spectrometry (nMS) to acquire both proteome identity and conformations. First, protein sequences and PTMs are obtained using the TDP method, while protein solvent-accessible surface area (SASA) is measured in parallel by nMS. These data are then input into the PSP module to acquire the most probable conformation of the protein under nMS experimental conditions. We validate the feasibility and accuracy of this method through the analysis of both a globular protein and an intrinsically disordered protein. Additionally, we apply this approach to delineate the conformations of ribosomal proteins pre- and post-assembly. Interactions of ribosomal proteins with drug molecules are also explored. By enabling proteome identification and conformation characterization from relatively small amounts of endogenous proteins, this method would bridge the gap between structural biology and conventional proteomics technologies.

molecular biology↗

bioRxiv2024

Reducing functionally defective HSCs alleviates aging-related phenotypes in old mice

Aging is a process accompanied by functional decline in tissues and organs with great social and medical consequences. Developing effective anti-aging strategies is of great significance. In this study, we demonstrated that transplantation of young hematopoietic stem cells (HSCs) into old mice can mitigate aging phenotypes, underscoring the crucial role of HSCs in the aging process. Through comprehensive molecular and functional analyses, we identified a subset of HSCs in aged mice that exhibit "younger" molecular profiles and functions, marked by low levels of CD150 expression. Mechanistically, CD150low HSCs from old mice can effectively differentiate into downstream lineage cells but not their CD150high counterparts. Notably, transplantation of old CD150low HSCs attenuates aging phenotypes and prolongs lifespan of elderly mice compared to those transplanted with unselected or CD150high HSCs. Importantly, reducing the dysfunctional CD150high HSCs can alleviate aging phenotypes in old recipient mice. Thus, our study demonstrates the presence of "younger" HSCs in old mice, and aging-associated functional decline can be mitigated by reducing dysfunctional HSCs.

cell biology↗

bioRxiv2024

Optimizing Angiopep-2 Density on Polymeric Nanoparticles for Enhanced Blood-Brain Barrier Penetration and Glioblastoma Targeting: Insights from In Vitro and In Vivo Experiments

The blood-brain barrier (BBB) poses a formidable challenge to efficient drug delivery into the brain. One promising approach involves leveraging receptor-mediated transcytosis facilitated by Angiopep-2 peptide (Ang-2)-conjugated nanoparticles. However, the precise impact of Ang-2 density on BBB penetration remains poorly understood. In this study, we developed a versatile polymeric nanoparticle system with tuneable Ang-2 surface density and systemically examined its influence on BBB penetration through various in vitro assays and an in vivo study. Our findings revealed a nuanced relationship between Ang-2 surface density and BBB penetration across the different experimental setups. In 2D cell culture, we observed a positive correlation between Ang-2 surface density and cellular association in hCMEC/D3 cells, characterized by a distinctive inflection point. Conversely, in the Transwell model, higher Ang-2 density correlated negatively with BBB penetration, whereas the BBB-GBM-on-a-chip showed the opposite trend. These disparities may arise from differences in avidity under static versus dynamic conditions, potentially modulating nanoparticle interactions due to fluidic forces. In vivo studies revealed that higher Ang-2 densities facilitated nanoparticle transport across the BBB, consistent with the findings of the BBB-GBM-on-a-chip model. Furthermore, loading doxorubicin into nanoparticles with optimal Ang-2 density resulted in controlled pH-responsive release and enhanced anticancer effect against U87 GBM cells in both 2D cell cultures and a 3D BBB-GBM-on-a-chip model. These results underscore the critical importance of optimizing Ang-2 surface density for efficient BBB penetration and emphasize the utility of dynamic models in preclinical in vitro assessment of novel nanoparticle formulations for targeted delivery to the brain.

bioengineering↗