Biology subjects

Lao, Y.

Publications and source records attributed to Lao, Y..

11 recordsLinked to original sources

bioRxiv2026

PTMExplorer: A Multi-Dimensional Integrative Visualization Platform for Protein Post-Translational Modification Function and Structure

Deciphering the functions of post-translational modifications (PTMs) is a critical bridge connecting large-scale modification proteomics data to mechanistic studies. However, most existing tools for visualizing PTM omics data are limited to site catalogs or single-dimensional feature displays. They lack the capability to simultaneously map user-derived differential modification sites onto multi-dimensional contexts, including protein three-dimensional (3D) structure, evolutionary conservation, functional sites, and disease associations. This limitation makes it difficult for researchers to rapidly assess the biological importance of candidate sites from among a vast number of differentially modified sites. Here, we present PTMExplorer, an interactive platform for the multi-dimensional visualization of protein PTMs. PTMExplorer comprises three core modules: PTM Inspector, built upon ProtVista, provides a multi-track, sequence-feature integrated view incorporating intrinsically disordered region (IDR) prediction (via flDPnn), surface accessibility calculation (via FreeSASA), and UniProt functional annotations; PTM 3D Locator, leveraging the Nightingale/Mol* engine, anchors modification sites onto AlphaFold/Protein Data Bank (PDB) 3D structures through residue mapping via PDBe-SIFTS; and PTM Overview, utilizing the R circlize package, presents a panoramic polar circos plot illustrating modification distribution and inter-group differential regulation. Additionally, three major disease-associated modification databases (PTMD, qPTM, and PhosCancer) are integrated as PTM-Disease Nexus, enabling co-localization comparison between user-defined differential sites and reported disease-related sites. PTMExplorer currently supports eight model organisms, accepts user-uploaded differential analysis results, and provides multi-dimensional annotations and various visualization options (https://www.bioladder.cn/PTMExplorer/). Using a multi-omics dataset from hepatocellular carcinoma (18 patients, 9 modification types) as a case study, we demonstrate the practical utility of PTMExplorer in screening potential biomarkers, revealing multi-modification coordination mechanisms, and distinguishing between absolute and relative quantification patterns.

bioinformatics↗

bioRxiv2026

DPF3a couples H3K36me2-dependent chromatin remodeling to genome architecture during myogenesis

Chromatin remodelers are generally thought to regulate gene expression through nucleosome mobilization and modulation of local chromatin accessibility. Whether these complexes can instead control transcription through higher-order chromatin organization remains poorly understood. Here we identify the BAF subunit DPF3a as an essential regulator of skeletal muscle regeneration and myogenic differentiation. DPF3a functions through association with the H3K36me2/3 reader HRP2 and preferentially localizes to a distinctive chromatin state characterized by focal depressions within broad H3K36me2 domains. Biochemical reconstitution demonstrates that H3K36 methylation directly enhances remodeling activity of the DPF3a-containing cBAF complex. Unexpectedly, despite profound transcriptional defects, loss of DPF3a produces minimal changes in local chromatin accessibility. Instead, DPF3a is required for long-range chromatin looping associated with activation of myogenic genes. Together, our findings uncover a non-canonical mechanism whereby a chromatin remodeler regulates transcription primarily through three-dimensional genome organization rather than local accessibility control, and establish histone modification-guided chromatin remodeling as a key principle in gene regulation.

molecular biology↗

bioRxiv2026

Progressive Loosening of a Dual Autoinhibitory Interface Activates PP2A-B56δ

Protein phosphatase 2A containing the B56{delta} regulatory subunit (PP2A-B56{delta}) is a critical signaling enzyme whose dysregulation is associated with cancer, neurodegenerative disorders, and Jordan's syndrome, a severe intellectual disability disorder caused by mutations in B56{delta}. Unlike other PP2A holoenzymes, PP2A-B56{delta} is regulated through a unique dual autoinhibition mechanism in which the N- and C-arms occlude the catalytic site while a substrate-mimicking short linear motif (SLiM) blocks the substrate-binding pocket. Although disease-associated mutations have been shown to alter enzyme activity, the molecular mechanism underlying activation of PP2A-B56{delta} and the effects of pathogenic mutations remain poorly understood. Here, we combined cryo-electron microscopy (cryo-EM), enhanced-sampling molecular dynamics (MD) simulations, Markov state model (MSM) construction, and transition-state analysis using Transition State identification via Dispersion and vAriational principle Regularized neural networks (TS-DAR) to characterize the conformational landscape of the disease variant E198K. Our cryo-EM analysis identified two distinct structures of E198K: an inactive closed-form with the N/C-arms resolved and an active loose-form in which the N/C-arms become highly flexible and could not be fully resolved. These structures therefore established that activation is governed by conformational changes of the N/C-arms but did not reveal the underlying mechanism. Starting from the inactive closed-form, we generated over 1,600 trajectories with an average length of 1,260 ns combined for E198K and wild-type (WT) PP2A-B56{delta}. TS-DAR identified four metastable states and two major activation pathways connecting inactive and active conformations. We found that activation occurs through progressive loosening of the N/C-arm interface while maintaining the overall holoenzyme architecture, rather than a complete opening of the interface. This mechanism exposes both the catalytic site and substrate-binding pocket. Comparison of E198K and WT revealed that the disease-associated mutation shifts the conformational equilibrium toward active states while leaving the transition-state ensemble largely unchanged. Mechanistically, E198K disrupts a salt-bridge network and weakens interactions between the internal loop and the C-arm that normally stabilize active-site occlusion. The resulting increase in C-arm mobility promotes active-site exposure and explains the elevated catalytic activity of the mutant. Together, these findings establish a previously uncharacterized activation mechanism for PP2A-B56{delta} and provide an atomic-level explanation for how the pathogenic E198K mutation allosterically promotes holoenzyme activation.

biophysics↗

bioRxiv2026

CK2 variant function and disease modelling in Drosophila reveal allelic heterogeneity and Wnt/β-catenin-mediated phenotypes

Heterozygous pathogenic variants in CSNK2A1 or CSNK2B encoding the Casein Kinase 2 (CK2) protein complex, lead to pediatric neurodevelopmental disorders, Okur-Chung Neurodevelopmental Syndrome (OCNDS) and Poirier-Bienvenu Neurodevelopmental Syndrome (POBINDS). OCNDS and POBINDS are characterized by a range of symptoms, including developmental delay, intellectual disability, facial dysmorphism, and seizures. Despite over 250 reported cases of OCNDS and POBINDS, we do not fully understand how specific alterations in CK2 relate to the heterogeneity observed in patients. To investigate this, we used the fruit fly, Drosophila melanogaster, as a model system. To assess variant impact, we co-expressed human CSNK2A1 and CSNK2B reference or disease-causing variants in flies. In parallel, we determined the role of Drosophila CkII in the developing and mature nervous system, specifically in neurons and glia. We found that 12/13 variants tested act as full or partial loss-of-function with one CSNK2A1 variant showing gain-of-function. Phospho-proteomic studies in neurons revealed separate signatures for loss- and gain-of-function variants. We found that neuronal and glial CkII is critical for organismal development. Reduction of neuronal CkII in the adult nervous system causes motor and seizure-like phenotypes. Finally, given the known role of CK2 in potentiating Wnt/{beta}-catenin signalling, we show that Wnt agonists partially rescue phenotypes associated with adult-specific neuronal reduction of CkII. This work generates Drosophila models of CSNK2A1 and CSNK2B expression to functionally assess variant impact, as well as an adult-specific neuronal loss-of-function model for drug screening and mechanistic studies.

genetics↗

bioRxiv2026

Asymmetric Hydration and Protonation Switching of Dual Aspartates Drive Flagellar Rotation

The bacterial flagellar motor is an intricate nanomachine that transforms chemical energy from ion gradients into mechanical rotation, enabling bacterial movement. While stator unit architectures are conserved across species, the molecular link connecting ion translocation to rotational force generation remains elusive. In this study, we refined the cryo-EM structures of MotAB from Campylobacter jejuni (CjMotAB) and integrated a suite of approaches--including single-structure based pKa predictors and free energy perturbation (FEP) calculations, as well as standard and constant-pH molecular dynamics (CpHMD) simulations of various structural models representing the plugged, unplugged, and plug-removed states with different protonation states of D22--to dissect its rotational mechanism. Based on pKa calculations, the D22 residues in chains F and G of MotB were identified as proton carriers supporting the previous hypotheses. Importantly, we observed asymmetric hydration patterns of the two D22 residues in the MotB dimer, along with their hydrogen bonding interactions with MotA T189, which contribute to functional specialization. Our findings reveal that MotA rotation requires two essential prerequisites: plug removal and alternating D22 protonation switching, coupled with dynamic gauche-trans conformational changes in the sidechain of D22. This work clarifies how protonation dynamics and structural asymmetry synergistically regulate CjMotAB rotation, advancing our understanding of bacterial flagellar motor function and providing a foundational framework for investigating diverse ion-driven biological motors.

biophysics↗

bioRxiv2025

Transcriptomic and protein analysis of human cortex reveals genes and pathways linked to NPTX2 disruption in Alzheimer's disease

BackgroundCognitive resilience to Alzheimers disease (AD) pathology is associated with preserved expression of NPTX2, an activity-regulated synaptic protein involved in circuit plasticity, excitation-inhibition balance, and complement-linked synapse regulation. However, the broader molecular programs coordinated with NPTX2 in resilient individuals remain unclear. MethodsWe analyzed postmortem middle temporal gyrus tissue using targeted PRM-MS proteomics in 135 individuals and bulk RNA-seq in an expanded 575-sample cohort. NPTX2-associated molecular coordination was assessed within cognitively normal low-pathology controls (CN-Lo), cognitively normal high-pathology controls (CN-Hi), mild cognitive impairment (MCI), and AD. Correlation-based approaches were applied using NPTX2 protein and NPTX2 mRNA expression as anchors to define resilience mechanisms in CN-Hi subjects. ResultsNPTX2 protein abundance was preserved across all controls regardless of age and pathology but reduced in MCI and AD. NPTX2 mRNA expression was also invariant across pathology within controls and reduced in MCI and AD but decreased markedly with age. Targeted proteomics identified NPTX2 relationships with synaptic and inhibitory-circuit proteins that were preserved across control groups, alongside CN-Hi-specific recruitment of trafficking, lysosomal, metabolic, and proteostasis-associated proteins. Transcriptome-wide correlations with NPTX2 revealed differences in gene co-expression between groups, identifying a prominent activity-dependent program including BDNF, VGF, SCG2, SST, SERTM1, DUSP4, and EGR4, that was preserved in both CN-Lo and CN-Hi subjects, while genes recruited to the NPTX2 network specifically in CN-Hi implicated immune, neuroprotective, translation, and proteostasis-related pathways. Coupling differential gene expression analysis with co-expression, we further identified five candidate resilience genes whose expression and NPTX correlation was preserved across controls, but lost in MCI and AD: SST, MAL2, TAC1, SERTM1, and RFK. Expression of genes in distinct NPTX2 co-expression classes can be freely explored in our bulk RNA-seq data and other public AD transcriptomic datasets at NeMO Analytics. ConclusionFindings suggest that cognitive resilience in the context of AD neuropathology engages a coordinated molecular state distinct from both persevered cognition without pathology and MCI/AD, which is organized around preserved and selectively remodeled NPTX2-associations. Rather than reflecting broad transcript abundance changes, resilience was characterized by maintained synaptic and inhibitory programs, and adaptive proteostasis and trafficking pathways that distinguish resilient high-pathology individuals from low-pathology controls or symptomatic AD.

bioinformatics↗

bioRxiv2025

Antimuscarinic drugs exert β-arrestin-biased agonism at the muscarinic acetylcholine type 1 receptor

Previous studies indicate that both pirenzepine (PZ), a selective orthosteric muscarinic acetylcholine type 1 receptor (M1R) antagonist, and muscarinic toxin 7 (MT7), a negative M1R allosteric modulator (NAM), act via M1R to promote neuritogenesis in cultured adult rodent primary dorsal root ganglia (DRG) sensory neurons, in part, through {beta}-arrestin-dependent activation of extracellular signal-regulated protein kinase 1/2 (ERK1/2). Furthermore, these antagonists reverse nerve degeneration in a variety of rodent models of peripheral neuropathy through multiple complementary pathways. To understand the therapeutic effects and mechanism of M1R antagonist-induced ERK1/2 phosphorylation, we tested the hypothesis that PZ and MT7 possess {beta}-arrestin-biased agonism at M1R to drive activation of ERK and enhance neurite outgrowth. Treatment for up to 30 min with PZ and MT7 dose-dependently recruited {beta}-arrestin2 to M1R (analyzed using nano-BRET) and increased ERK phosphorylation in both HEK293 cells and DRG neurons. DRG neurons of different sub-types express M1R, and ERK activation by MT7 was only observed in M1R-positive neurons. These novel pharmacological effects occurred in the absence of activation of G protein signaling or receptor internalization. PZ phosphorylated M1R at six specific serine/threonine residues (T230, S251, T254, S321, T354, S356) of intracellular loop 3 (ICL3) and deletion mutation of these sites suppressed PZ and MT7 induction of {beta}-arrestin binding to M1R and inhibited ERK activation. With regard to PZ signaling, alanine substitution at S251 and T254 was sufficient to impede {beta}-arrestin binding and ERK activation. {beta}-arrestin-biased activity of PZ and MT7 involved the mobilization of casein kinase 2 (CK2) and this occurred in the absence of Gq or G protein receptor kinase (GRK) activity. Pharmacological or siRNA-based inhibition of CK2 blocked PZ-induction of {beta}-arrestin association, ERK activation and neurite outgrowth in DRG neurons. In conclusion, PZ/MT7 activated M1R toward the {beta}-arrestin signaling pathway in both HEK293 cells and DRG neurons to augment ERK activation and neurite outgrowth via engagement of CK2. One-sentence summaryAntimuscarinic drugs act as {beta}-arrestin-biased agonists via casein kinase 2 activation to promote ERK1/2 phosphorylation and neurite outgrowth O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/649213v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@1bb8da7org.highwire.dtl.DTLVardef@510218org.highwire.dtl.DTLVardef@608d69org.highwire.dtl.DTLVardef@e40d9e_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstract.C_FLOATNO Schematic presentation of the effect of muscarinic ligands at M1R associated signaling pathway. (A) Muscarine/carbachol acts as a balanced ligand by engaging both Gq and -arrestin signaling pathways and treatment with pirenzepine/MT7 blocks these effects. (B) Pirenzepine/MT7 acts as a -arrestin biased ligand by 1) phosphorylating of ICL3 region of M1R via CK2 (but not GRKs), 2) no activation of G protein signaling, 3) recruitment of -arrestin 2 and 4) ERK1/2 activation leading to neurite outgrowth in DRG sensory neurons. This figure was generated by BioRender under license number EK285MUOPQ. C_FIG

pharmacology and toxicology↗

bioRxiv2024

SysQuan: repurposing SILAC mice for the affordable absolute quantitation of the human proteome

Relative quantitation, used by most MS-based proteomics laboratories to determine protein fold-changes, requires samples being processed and analyzed together for best comparability through minimizing batch differences. This limits the adoption of MS-based proteomics in population-wide studies, and the detection of subtle but relevant changes in heterogeneous samples. Absolute quantitation circumvents these limitations and enables comparison of results across laboratories, studies, and longitudinally. However, high costs of the essential stable isotope labeled (SIL) standards prevents widespread access and limits the number of quantifiable proteins. Our new approach, called "SysQuan", repurposes SILAC mouse tissues/biofluids as system-wide internal standards for matched human samples to enable absolute quantitation of, theoretically, two-thirds of the human proteome using 157,086 shared tryptic peptides. We demonstrate that SysQuan enables quantification of 70% and 31% of the liver and plasma proteomes, respectively. We demonstrate for 14 metabolic proteins that abundant SIL mouse tissues enable cost-effective reverse absolute quantitation in, theoretically, 1000s of human samples. Moreover, 10,000s of light/heavy doublets in untargeted SysQuan datasets enable unique post-acquisition absolute quantitation. SysQuan empowers researchers to replace relative quantitation with affordable absolute quantitation at scale, making data comparable across laboratories, diseases and tissues, enabling completely novel study designs and increasing reusability of data in repositories. TOC Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/622109v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@c7a58org.highwire.dtl.DTLVardef@1344320org.highwire.dtl.DTLVardef@2321e3org.highwire.dtl.DTLVardef@85bf2d_HPS_FORMAT_FIGEXP M_FIG C_FIG

systems biology↗

bioRxiv2024

Identification of novel myokines and putative protein targets that mediate functional adaptations in response to chronic contractile activity induced skeletal muscle-extracellular vesicle treatment

We have previously shown that skeletal muscle-derived extracellular vesicles (EVs) released post-chronic contractile activity (CCA) increased mitochondrial biogenesis in murine myoblasts, and decreased cell viability and induced apoptosis and senescence in non-small cell lung cancer cells. While the underlying mechanisms are unknown, the effects perpetuated were dependent on membrane-bound proteins. Here, we performed an extensive LC-MS/MS proteomic analysis on EVs from control and CCA myotubes. A total of 2900 proteins were identified in CON-EVs and CCA-EVs, including EV-associated proteins such as TSG101, tetraspanins (CD9, CD81, and CD63), flotillin-1, and annexins. Of these, 856 proteins are novel and not listed in EV databases (ExoCarta and Vesiclepedia), indicating that myotube-EVs harbor proteins not yet identified in EVs of different origin. Additionally, we identified 2062 unique proteins that have not yet been previously reported in myotube-EVs to date. Remarkably, of the 2900 total proteins identified, we observed 46 upregulated, and 25 downregulated differentially expressed proteins (DEPs) in CCA-EVs vs. control-EVs. Most of upregulated DEPs include EV-associated proteins. Comparing the 71 DEPs with proteins expressed in skeletal muscle indicated 61 of these as potential myokines. We identified actin cytoskeleton signaling, integrin signaling and muscle contraction as the most enriched pathways among the DEPs using different databases/software including FunRich, KEGG, STRING and Ingenuity Pathway Analysis. Using a relevance score that prioritized membrane-bound proteins with known function in mitochondrial biogenesis and inhibition of cancer growth, we identified top-scoring highly enriched DEPs of interest: IGF1R, ATP7A, PFN1, GJA1, PRKCA and ITGA6. We confirmed upregulation of these targets in EVs using immunoblotting. Among these top-scoring DEPs, PFN1, and ITGA6 are associated with EVs, with expression upregulated following acute exercise. In summary, we report the first comprehensive analysis of skeletal muscle-EV proteome following CCA, with identification of putative protein targets and signaling pathways that may execute the pro-metabolic and anti-tumorigenic effects of CCA-EVs.

cell biology↗

bioRxiv2023

Activation of Shh/Smo is sufficient to maintain oligodendrocyte precursor cells in an undifferentiated state but is not necessary for differentiation

Myelination is the terminal step in a complex and precisely timed program that orchestrates the proliferation, migration and differentiation of oligodendroglial cells. It is thought that Sonic Hedgehog (Shh) acting on Smoothened (Smo) participates in regulating this process, but that these effects are highly context dependent. Here, we investigate oligodendroglial development and remyelination from three specific transgenic lines: NG2-CreERT2 (control), Smofl/fl/NG2-CreERT2 (loss of function) and SmoM2/NG2-CreERT2 (gain of function), as well as pharmacological manipulation that enhance or inhibit the Smo pathway (SAG or cyclopamine treatment respectively). To explore the effects of Shh/Smo on differentiation and myelination in vivo, we developed a highly quantifiable model by transplanting OPCs in the retina. We find that myelination is greatly enhanced upon cyclopamine treatment and hypothesize that Shh/Smo could promote OPC proliferation to subsequently inhibit differentiation. Consistent with this hypothesis, we find that the genetic activation of Smo significantly increased numbers of OPCs and decreased oligodendrocyte differentiation when we examined the corpus callosum during development and after cuprizone demyelination and remyelination. However, upon loss of function with the conditional ablation of Smo, myelination in the same scenarios are unchanged. Taken together, our present findings suggest that the Shh pathway is sufficient to maintain OPCs in an undifferentiated state, but is not necessary for myelination and remyelination.

neuroscience↗

bioRxiv2023

Supporting central nervous system neuroprotection and remyelination by specific TLR4 antagonism

ApTOLL is an aptamer specifically designed to antagonize Toll-Like Receptor 4 (TLR4), a relevant actor for innate immunity involved in inflammatory responses in multiple sclerosis (MS) and other diseases. MS is a primary demyelinating, chronic, inmune and neurodegenerative disease of the central nervous system that normally debuts in young adults. The currently available therapeutic arsenal to treat MS is composed of immunomodulators but, to date, there are no (re)myelinating drugs available in clinics. Our present study shows cells expressing TLR4 in demyelinating lesions of MS patients (postmortem samples from cerebral cortex) and, as a derivative, we studied the effect of TLR4 inhibition with ApTOLL in animal models of MS (experimental autoimmune encephalomyelitis -EAE- and the cuprizone). The treatment with ApTOLL positively impacted the clinical symptomatology, and this was associated with better preservation plus restoration of myelin and oligodendrocytes in the demyelinated lesions of these animals, which suggests not only an immunomodulatory but also a remyelinating effect of the treatment with ApTOLL. This latter was corroborated on purified cultures of rodent and adult human oligodendrocyte precursor cells (OPCs), confirming the expression of TLR4 in this cell type. Altogether, the molecular nature of ApTOLL and its mechanism/s of action strongly supports this compound as a novel candidate to treat MS and other demyelinating scenarios.

neuroscience↗