Biology subjects

Lopez, O.

Publications and source records attributed to Lopez, O..

7 recordsLinked to original sources

bioRxiv2026

Integrator governs NF-κB dwell time on chromatin to drive inflammatory transcription

NF-{kappa}B must remain bound to chromatin long enough to activate innate immune genes, but what stabilizes this transcriptionally productive state is unknown. Here, we identify the Integrator complex as an essential determinant of NF-{kappa}B chromatin residence in mouse and human cells. Integrator associated with NF-{kappa}B and co-occupied its genomic targets. Single-molecule tracking revealed that Integrator markedly prolonged the chromatin residence of RELA molecules in a low-mobility state. Catalytically inactive INTS11-E203Q fully restored NF-{kappa}B-dependent transcription, separating this chromatin-retention function from RNA cleavage. Conditional Ints11 deletion abolished TLR4-dependent dendritic-cell activation in adult mice and impaired LPS-induced transcription in mouse embryonic fibroblasts. Loss of Integrator in human cancer cells similarly suppressed TNF-induced inflammatory gene expression. These findings uncover a noncatalytic function for Integrator that stabilizes NF-{kappa}B at enhancers, complementing its established catalytic control of RNA polymerase II pause release. By coupling signal recognition at enhancers to productive elongation, Integrator provides a molecular solution to a central challenge of multicellularity: converting transient extracellular cues into coherent gene-expression programs across complex chromatin landscapes.

molecular biology↗

bioRxiv2026

Linking coronary microvascular structure and function in preclinical models of coronary microvascular disease

Despite growing awareness of the importance of the coronary microvasculature in cardiac health and disease, Coronary Microvascular Disease (CMVD) remains poorly understood, underdiagnosed, and without targeted therapies. Preclinical models and quantitative tools to measure CMVD are needed. Here, we use two novel quantitative methods to assess coronary microvascular structure (multi-fractal spectrum analysis) and function (Single Photon Emission Computed Tomography (SPECT)-based intramyocardial blood volume (IMBV) imaging) in mouse models of CMVD. Change in IMBV ({bigtriangleup}IMBV), which serves as a quantitative measure of vasodilatory capacity or microvascular function, was reduced with aging, driven by a significant decrease in female mice. Male mice on ApoE-/- background, fed a high-fat diet (HFD) for 6 months, or both had significantly reduced {bigtriangleup}IMBV. We used immunofluorescence to assess both traditional capillary density and global branching structure and vessel heterogeneity using multifractal spectrum analysis. Both were significantly reduced in all groups, and linear regression modeling showed that they were independently associated with {bigtriangleup}IMBV. Finally, we used {bigtriangleup}IMBV to assess the effects of widely used control Adeno-associated viral (AAV) vectors on coronary microvascular function. AAV-overexpression of GFP did not affect function, but Cre-recombinase compromised coronary microvascular structure and function by 16 weeks. In summary, quantitative assessments of coronary microvascular structure and function highlight changes consistent with CMVD seen with aging, female sex, and metabolic insults. Functional changes are partially driven, but not fully defined, by changes in the underlying structure, highlighting the important and incomplete link between structure and function.

physiology↗

bioRxiv2025

The histone variant H2A.Z and its chaperone SRCAP are required for RNA polymerase II to access HBV chromatin

BackgroundChronic hepatitis B virus (HBV) infection remains a major global health burden due to the persistence of covalently closed circular DNA (cccDNA), a stable episomal viral reservoir resistant to current antiviral therapies. The transcriptional activity of cccDNA depends on its chromatin organization, yet the contribution of histone variants to this regulation remains poorly understood. MethodsUsing mass spectrometry-based proteomics on native cccDNA purified from infected primary human hepatocytes, we identified the histone variant H2A.Z and its chaperone SRCAP as cccDNA-associated proteins. Functional analyses combining shRNA-mediated silencing, chromatin immunoprecipitation (ChIP), and ATAC-seq were performed in HBV-infected HepG2-NTCP cells and primary human hepatocytes to characterize their roles in cccDNA formation and transcription. ResultsDepletion of H2A.Z.1, H2A.Z.2, or SRCAP reduced both HBV RNA levels and cccDNA formation. H2A.Z recruitment to cccDNA correlated with the establishment of active chromatin marks (H3K4me3) and enhanced RNA polymerase II loading, promoting an open and transcriptionally active chromatin state. In addition, we identified BRD2, an H2A.Z-associated transcriptional co-activator, as a positive regulator of HBV transcription. shRNA mediated depletion and pharmacological degradation of BRD2 using the PROTAC ARV-771 reduced HBV RNA levels, supporting its potential as an antiviral target. ConclusionOur findings uncover a critical role of the H2A.Z variant and SRCAP complex in cccDNA formation and transcriptional activation. By facilitating chromatin accessibility and RNA polymerase II recruitment, H2A.Z establishes an epigenetic environment favorable to HBV persistence. Targeting H2A.Z-associated co-activators such as BRD2 may represent a promising strategy to silence cccDNA transcription and achieve functional HBV cure.

microbiology↗

bioRxiv2025

ddOTs: a multiplexed quantitative ddPCR approach for resolving overlapping hepatitis B virus transcripts to decipher cccDNA-driven transcription

Chronic hepatitis B virus infection persists as a global health crisis, driving life-threatening liver pathologies such as hepatocellular carcinoma. Central to HBVs resilience is the covalently closed circular DNA (cccDNA), a viral minichromosome that orchestrates viral transcription and sustains infection. Dissecting cccDNA-driven transcription remains a formidable challenge due to the dense overlap of viral open reading frames, which generates RNA transcripts with shared sequences, complicating precise quantification and hindering efforts to unravel HBVs transcriptional regulation. To address this bottleneck, we developed a multiplexed assay named ddPCR for Overlapping Transcripts (ddOTs) capable of simultaneously quantifying all major HBV RNA species, including splice variants, with unprecedented specificity and sensitivity. This method overcomes current limitations by leveraging the high-resolution power of ddPCR to deconvolute overlapping transcripts, enabling direct interrogation of individual promoter/enhancer activities and RNA stability. By offering a cost-effective, scalable solution for precise RNA profiling in rare biological samples, this breakthrough tool unlocks new avenues for exploring cccDNA biology and accelerating antiviral drug development.

microbiology↗

bioRxiv2025

Evaluating the antiviral activity of Termin-8 and Finio against a surrogate ASFV-like algal virus

IntroductionAnitox has developed and markets Termin-8 (a formaldehyde-based product) and Finio (non-formaldehyde solution) for the control microbial contamination in feed. To date, both Finio and Termin-8 have not been tested for their potential antiviral activity against megaviruses, such African swine fever virus (ASFV) and its surrogate algal virus, Emiliania huxleyi virus (EhV). Given the limited access and great expense for routine chemical mitigation testing with ASFV, we focused on using EhV as a safe and effective surrogate to evaluate the antiviral activity of these Anitox products. The specific objective of the current study was to evaluate the time course of incubation from hours to days to mimic possible field relevant exposure times for the potential preventative mitigation of megaviruses using Termin-8 and Finio. MethodsEmiliania huxleyi virus was treated with the Anitox recommended concentrations of Termin-8 (0.1% to 0.3% final) and Finio (0.05% to 0.2% final) in biological triplicate experiments. Both viability qPCR (V-qPCR) and standard PCR (S-qPCR) were conducted for EhV copies at 1 hr, 5 hrs, 24 hrs and day 7 post-inoculation. ResultsWe observed that both Termin-8 and Finio, at their highest treatment concentrations, showed the greatest log reduction of 2 and 4.5 log10 units, respectively, at the earliest 1 hr post-inoculation time point. Although Termin-8 efficacy did not improve with time, treatment with Finio showed 100% viable viral inactivation (>5 log10 reduction units) at the lowest concentration after 7 days of exposure. In addition, Finio showed negligible viral DNA removal post-treatment as observed via S-qPCR, but use of Termin-8 resulted in a reduction of viral DNA (S-qPCR) equivalent to that observed using the V-qPCR assay. DiscussionOur results demonstrate for the first time that both Termin-8 and Finio can be used as effective chemical mitigants against megaviruses such as EhV and ASFV. Moreover, the mechanism of chemical mitigation involves degradation of the virus particle itself. However, although Termin-8 efficacy appeared to be less than that of Finio, this is likely not the case because Termin-8 cross-links protein viral capsids and fixes the viral particle to make it non-infectious. With the threat of ASFV introduction into the North American swine industry and other non-infected regions around the world, the use of both Termin-8 and Finio as an effective preventive or mitigation strategy to prevent the transmission of ASFV by reducing particle viability in contaminated feed. Additional research is warranted with ASFV, in the presence of various types of feed ingredients and complete feeds and use bioassays to evaluate infectivity because results from V-qPCR revealed that intact viable particles remain post-treatment.

molecular biology↗

bioRxiv2022

Temporary knockdown of p53 during focal limb irradiation increases the development of sarcomas

BackgroundApproximately half of all cancer patients receive radiotherapy and, as cancer survivorship rates increase with more effective therapies, the very low rate of radiation-associated sarcomas is rising. Radiation-associated sarcomas are life-threatening cancers, and radiation exposure is a primary risk factor for sarcoma development. During radiotherapy or other genotoxic cancer therapy for p53 mutant cancers, pharmacological inhibition of p53 has been proposed to ameliorate acute injury of normal tissues. However, enhancing the survival of normal cells that sustain DNA damage by temporarily inhibiting p53 has the potential to increase the risk of cancer development. Here, we use in vivo shRNA technology to examine the consequences of temporarily reducing p53 expression on radiation-induced sarcoma development. MethodsWe utilized a mouse model of radiation-induced sarcoma where mice express a doxycycline (dox)-inducible p53 shRNA to temporarily and reversibly reduce p53 expression. Mice were placed on a dox diet 10 days prior to receiving 30 or 40 Gy hind limb irradiation in a single fraction and then returned to normal chow. Mice were examined weekly for sarcoma development and scored for radiation-induced normal tissue injuries. Radiation-induced sarcomas were harvested and subjected to RNA sequencing. ResultsFollowing single high-dose irradiation, 21% of temporary p53 knockdown animals developed a sarcoma in the radiation field compared to 2% of control animals. Mice with more severe acute injuries in the first 3 months after irradiation had a significantly increased risk of developing late persistent wounds in the soft tissue and bone. Chronic radiation-induced wounds were associated with sarcomagenesis. Examination of muscle stem cells by flow cytometry following hind limb irradiation indicated p53 knockdown preserves muscle stem cells in the irradiated limb, supporting the notion that temporary p53 knockdown at the time of irradiation reduces death of cells with DNA damage which may then persist to develop into a sarcoma. We performed RNA sequencing on 16 radiation-induced sarcomas compared to normal muscle controls. Gene set enrichment analysis revealed upregulation in the sarcomas of genes related to translation, epithelial mesenchymal transition (EMT), inflammation, and the cell cycle versus downregulation of genes related to myogenesis and tumor metabolism. Furthermore, genes with increased copy number such as Met and Cdk4 were overexpressed in tumors. ConclusionsTemporary reduction of p53 during high-dose irradiation increases late effects including tissue injuries and sarcoma development.

cancer biology↗

bioRxiv2018

Genetic Determinants of Cortical Structure (Thickness, Surface Area and Volumes) among Disease Free Adults in the CHARGE Consortium

Cortical thickness, surface area and volumes (MRI cortical measures) vary with age and cognitive function, and in neurological and psychiatric diseases. We examined heritability, genetic correlations and genome-wide associations of cortical measures across the whole cortex, and in 34 anatomically predefined regions. Our discovery sample comprised 22,822 individuals from 20 cohorts within the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) consortium and the United Kingdom Biobank. Significant associations were replicated in the Enhancing Neuroimaging Genetics through Meta-analysis (ENIGMA) consortium, and their biological implications explored using bioinformatic annotation and pathway analyses. We identified genetic heterogeneity between cortical measures and brain regions, and 161 genome-wide significant associations pointing to wnt/{beta}-catenin, TGF-{beta} and sonic hedgehog pathways. There was enrichment for genes involved in anthropometric traits, hindbrain development, vascular and neurodegenerative disease and psychiatric conditions. These data are a rich resource for studies of the biological mechanisms behind cortical development and aging.

genetics↗