Biology subjects

Le, Q.

Publications and source records attributed to Le, Q..

7 recordsLinked to original sources

bioRxiv2025

Superresolution imaging of live samples by centroid reassignment microscopy

Superresolution imaging has become one of the most important recent advances in microscopy development. However, most superresolution methods are ill-adapted for live-sample imaging because they are unacceptably slow, susceptible to artifacts, or require the use of specialized fluorophores and labeling protocols. We introduce a superresolution method called centroid reassignment microscopy (CRM) that overcomes these limitations. CRM is a simple variation on confocal microscopy wherein the single-element detector and small pinhole are replaced by a centroid detector and larger pinhole. Superresolution is obtained by reassigning the detected fluorescence to its centroid location of as a function of the scanning excitation focus location. Our method bears resemblance to the method of image scanning microscopy, which involves the use of an array detector, with the advantage that CRM provides improved resolution for the same number of detected photons while being simpler to implement and less sensitive to noise. CRM is light-efficient, fast (single-frame), robust to defocus aberrations, and requires no changes whatsoever in standard fluorescence imaging protocols, making it uniquely attractive for superresolution imaging of live, dynamic samples.

bioengineering↗

bioRxiv2025

Human meibomian gland organoids to study epithelial homeostasis and dysfunction

Meibomian glands (MGs) are holocrine glands that secrete lipids to maintain the homeostasis of ocular surface, and their dysfunction leads to dry eye disease. Herein, we established long-term 3D organoid culture for murine and human MGs, which retained the cell lineages and lipid-producing ability. The organoids mimicked the drug treatment responses and generated functional MGs after orthotopic transplantation. Inspired by organoid cultures, we found FGF10 eye drops could rescue all-trans retinoic acid-induced MG dysfunction in mice. Besides, nicotinamide uniquely hampered the human MG organoid expansion by inhibiting FGF10 signaling. Single-cell atlas and lipidome not only aligned the delineated cell types and featured lipids between human MGs and organoids, but highlighting MAPK signaling inhibition enhanced acinar cell differentiation and functional maturation of MG organoids. In summary, this study established an organoid platform to explore epithelial homeostasis and dysfunction of MGs, facilitating drug development and regenerative medicine for dry eye disease.

cell biology↗

bioRxiv2025

Disturbed engram network caused by NPTXs downregulation underlies aging-related memory deficits

Engram cells storing specific memories are allocated to separate neuronal ensembles, which preferentially recruit either excitatory or inhibitory inputs to drive precise memory expression. However, how these formed neuronal ensembles maintain their stability, and whether the disturbed stability contributes to aging-related memory deficits remain elusive. Here, we show that neuronal pentraxin1 (NPTX1) facilitates Kv7.2-mediated inhibition of Fos+ensemble hyperexcitability, thereby restricting its response to excitatory inputs from medial entorhinal cortex (MEC) and promoting memory expression in the fear context. Meanwhile, NPTX2 facilitates the perisomatic inhibition of the Npas4+ ensemble by parvalbumin+ (PV+) interneurons, thus preventing fear memory overgeneralization. Pharmacological activation of Kv7.2 or chemogenetic activation of PV+ interneurons repaired memory deficits caused by engram specific NPTXs depletion. Contextual fear memory precision and NPTXs expression in dentate gyrus (DG) engram cells are decreased in aged mice. Overexpression of NPTX1 in Fos+ensemble or AMPAR binding domain of NPTX2 in Npas4+ ensemble rescued memory imprecision. These findings elucidate that the coordination of NPTXs prevents engram ensembles from becoming hyperactive and provide a causal link between engram network destabilization and aging-associated memory deficits.

neuroscience↗

bioRxiv2025

Protease-Containing Nanobodies for Detecting and Manipulating Intracellular Antigens Using Antiviral Drugs

Tools to induce the formation of protein-protein interactions (PPIs) via small molecules are essential for investigating and engineering biological systems. Here, we introduce a protease-based strategy for controlling the preservation of otherwise self-cleaving nanobodies. By inserting the Hepatitis C Virus NS3 cis-protease into the nanobody scaffold, we showed that the antigen-binding ability of these chimeric nanobodies can be controlled in a dose-dependent manner using NS3 inhibitors. We demonstrated the generalizability of this approach by designing and validating drug-controllable nanobodies targeting mCherry (LaM4), eGFP (LaG16), and the ALFA peptide tag. Additionally, we showed that an NS3-containing version of a nanobody targeting the {beta}2-adrenergic receptor can control endogenous G-protein-mediated signaling activity. Overall, we introduce new chemogenetic components for controlling intracellular PPIs using clinically approved antiviral drugs.

synthetic biology↗

bioRxiv2024

EpigeneticAgePipeline: an R package for comprehensive assessment of epigenetic age metrics from methylation microarrays

Epigenetic age is a biological age estimate based on nuclear DNA methylation patterns. Epigenetic clocks measure biological age by analyzing predictable changes in DNA methylation sites associated with aging. This study introduces EpigeneticAgePipeline, an R package that streamlines the estimation of epigenetic age metrics including Horvath, Horvath skin and blood, Hannum, PhenoAge/Levine, GrimAge (V1, and V2), and DunedinPACE plus additional acceleration metrics based on all other clocks. Quality control includes detection p-value filtering (sample- and probe-level), bead-count thresholds, and Illumina quality control intensity checks. EpigeneticAgePipeline supports Illumina Infinium methylation microarrays (HumanMethylation27, HumanMethylation450, HumanMethylationEPIC/EPICv2, and Human Methylation Screening Array). It offers functionalities including data preprocessing, normalization, cell count imputation, residual generation accounting for principal components and batch effects, and extensive visualizations for improved interpretability. Validation was performed using GEO dataset GSE237561, confirming the accuracy of the pipeline. EpigeneticAgePipeline provides an integrated workflow from raw data to advanced statistical analyses and visualizations, improving usability over existing tools. In addition to the traditional clocks mentioned, the package also integrates a set of additional epigenetic age clocks (PedBE, Wu, TL, BLUP, and EN). Future updates will include emerging epigenetic age measures to maintain relevance in this evolving field.

bioinformatics↗

bioRxiv2023

Chromatin Profiling of CBFA2T3-GLIS2 AMLs Identifies Key Transcription Factor Dependencies and BRG1 Inhibition as a Novel Therapeutic Strategy

Oncogenic fusions involving transcription factors are present in the majority of pediatric leukemias; however, the context-specific mechanisms they employ to drive cancer remain poorly understood. CBFA2T3-GLIS2 (C/G) fusions occur in treatment-refractory acute myeloid leukemias and are restricted to young children. To understand how the C/G fusion drives oncogenesis we applied CUT&RUN chromatin profiling to an umbilical cord blood/endothelial cell (EC) co-culture model of C/G AML that recapitulates the biology of this malignancy. We find C/G fusion binding is mediated by its zinc finger domains. Integration of fusion binding sites in C/G- transduced cells with Polycomb Repressive Complex 2 (PRC2) sites in control cord blood cells identifies MYCN, ZFPM1, ZBTB16 and LMO2 as direct C/G targets. Transcriptomic analysis of a large pediatric AML cohort shows that these genes are upregulated in C/G patient samples. Single cell RNA-sequencing of umbilical cord blood identifies a population of megakaryocyte precursors that already express many of these genes despite lacking the fusion. By integrating CUT&RUN data with CRISPR dependency screens we identify BRG1/SMARCA4 as a vulnerability in C/G AML. BRG1 profiling in C/G patient-derived cell lines shows that the CBFA2T3 locus is a binding site, and treatment with clinically-available BRG1 inhibitors reduces fusion levels and downstream C/G targets including N-MYC, resulting in C/G leukemia cell death and extending survival in a murine xenograft model.

cancer biology↗

bioRxiv2023

EZH2 synergizes with BRD4-NUT to drive NUT carcinoma growth through silencing of key tumor suppressor genes

NUT carcinoma (NC) is an aggressive carcinoma driven by the BRD4-NUT fusion oncoprotein, which activates chromatin to promote expression of pro-growth genes. BET bromodomain inhibitors (BETi) impede BRD4-NUTs ability to activate genes and are thus a promising treatment but limited as monotherapy. The role of gene repression in NC is unknown. Here, we demonstrate that EZH2, which silences genes through establishment of repressive chromatin, is a dependency in NC. Inhibition of EZH2 with the clinical compound tazemetostat (taz) potently blocked growth of NC cells. Epigenetic and transcriptomic analysis revealed that taz reversed the EZH2-specific H3K27me3 silencing mark, and restored expression of multiple tumor suppressor genes while having no effect on key oncogenic BRD4- NUT-regulated genes. CDKN2A was identified as the only gene amongst all taz-derepressed genes to confer resistance to taz in a CRISPR-Cas9 screen. Combined EZH2 inhibition and BET inhibition synergized to downregulate cell proliferation genes resulting in more pronounced growth arrest and differentiation than either inhibitor alone. In pre-clinical models, combined taz and BETi synergistically blocked growth and prolonged survival of NC-xenografted mice, with all mice cured in one cohort. STATEMENT OF SIGNIFICANCEIdentification of EZH2 as a dependency in NC substantiates the reliance of NC tumor cells on epigenetic dysregulation of functionally opposite, yet highly complementary chromatin regulatory pathways to maintain NC growth. In particular, repression of CDKN2A expression by EZH2 provides a mechanistic rationale for combining EZH2i with BETi for the clinical treatment of NC.

cancer biology↗