Biology subjects

Dybas, J. M.

Publications and source records attributed to Dybas, J. M..

7 recordsLinked to original sources

bioRxiv2026

Differential assembly of RNP granules via activation of distinct dsRNA sensors by adenovirus mutants

Recognition of dsRNA triggers antiviral defense mediated by PKR and OAS3/RNase L pathways through translational arrest and RNA decay. This is accompanied by assembly of distinct cytoplasmic ribonucleoprotein (RNP) condensates termed stress granules (SGs) and RNase L-dependent bodies (RLBs). Here we show that adenovirus infection differentially modulates dsRNA sensors and RNP granule assembly. Infection with splicing-defective {Delta}E4 mutant leads to dsRNA accumulation and activation of both PKR and OAS3/RNase L, promoting formation of RLB-like granules. In contrast, mutants lacking virus-associated (VA) RNAs trigger PKR activation and assembly of SGs despite absence of detectable dsRNA. Proteomic analysis revealed distinct protein compositions of canonical SGs and RLBs, which were reflected in virus-induced granules. While {Delta}VA-induced granules were PKR-dependent, {Delta}E4 mutants induced RLB-like granules independently of PKR and RNase L. In these cells, granule assembly coincided with translational arrest independent of eIF2 phosphorylation, indicating additional pathways linking nuclear dsRNA sensing to translational control and RNP granule assembly during viral infection. These findings provide novel insights into how distinct dsRNA sensors modulate translation and RNP condensates in response to stress.

cell biology↗

bioRxiv2024

The splicing modulator CLK1 is a candidate oncogenic dependency in pediatric high-grade gliomas

BackgroundPediatric brain tumors are the leading cause of disease-related mortality in children, yet many aggressive tumors lack effective therapies. RNA splicing is a hallmark of cancer, but it has not yet been systematically studied in pediatric brain tumors. MethodsWe analyzed 729 pediatric brain tumors spanning histologies and molecular subtypes to quantify differential tumor splicing. We developed the Splicing Burden Index (SBI) to enable cross-sample comparisons and performed hierarchical clustering of highly variable splice events to define splicing-informed tumor groups. These were integrated with clinical outcomes, pathway activity, and proteogenomic data. Recurrent splice events were prioritized for predicted functional impact, and in vitro perturbation studies were performed targeting the splicing kinase CDC-like kinase 1 (CLK1). ResultsSBI revealed substantial inter- and intra-histology heterogeneity. Clusters were enriched for histologies and molecular subtypes, several of which were independently associated with survival beyond histology and clinical covariates. Spliceosome pathway activity varied across clusters and was associated with worse survival, yet was not correlated with SBI, indicating distinct dimensions of splicing dysregulation. Functional prioritization identified a recurrent in CLK1 exon 4, required for canonical kinase activity. CLK1 exon 4 inclusion followed an oncofetal pattern and showed context-dependent associations with outcome distinct from total CLK1 expression. Pharmacologic inhibition and exon 4-specific perturbation of CLK1 reduced tumor cell viability and disrupted cancer-relevant splicing and transcriptional programs. ConclusionsThis study systematically characterizes splicing in pediatric brain tumors, identifies splicing-informed subgroups, and prioritizes CLK1 exon 4 as an oncofetal tumor-specific event, motivating further preclinical exploration. Key Points[bullet] Splicing analysis of 729 pediatric CNS tumors identifies splicing-defined clusters. [bullet]CLK1 exon 4 inclusion is widespread and developmentally regulated. [bullet]Exon-level CLK1 regulation shows context-dependent links to prognosis in aggressive CNS tumors.

genomics↗

bioRxiv2024

The Open Pediatric Cancer Project

BackgroundIn 2019, the Open Pediatric Brain Tumor Atlas (OpenPBTA) was created as a global, collaborative open-science initiative to genomically characterize 1,074 pediatric brain tumors and 22 patient-derived cell lines. Here, we present an extension of the OpenPBTA called the Open Pediatric Cancer (OpenPedCan) Project, a harmonized open-source multi-omic dataset from 6,112 pediatric cancer patients with 7,096 tumor events across more than 100 histologies. Combined with RNA-Seq from the Genotype-Tissue Expression (GTEx) and The Cancer Genome Atlas (TCGA), OpenPedCan contains nearly 48,000 total biospecimens (24,002 tumor and 23,893 normal specimens). FindingsWe utilized Gabriella Miller Kids First (GMKF) workflows to harmonize WGS, WXS, RNA-seq, and Targeted Sequencing datasets to include somatic SNVs, InDels, CNVs, SVs, RNA expression, fusions, and splice variants. We integrated summarized CPTAC whole cell proteomics and phospho-proteomics data, miRNA-Seq data, and have developed a methylation array harmonization workflow to include m-values, beta-vales, and copy number calls. OpenPedCan contains reproducible, dockerized workflows in GitHub, CAVATICA, and Amazon Web Services (AWS) to deliver harmonized and processed data from over 60 scalable modules which can be leveraged both locally and on AWS. The processed data are released in a versioned manner and accessible through CAVATICA or AWS S3 download (from GitHub), and queryable through PedcBioPortal and the NCIs pediatric Molecular Targets Platform. Notably, we have expanded PBTA molecular subtyping to include methylation information to align with the WHO 2021 Central Nervous System Tumor classifications, allowing us to create research-grade integrated diagnoses for these tumors. ConclusionsOpenPedCan data and its reproducible analysis module framework are openly available and can be utilized and/or adapted by researchers to accelerate discovery, validation, and clinical translation.

genomics↗

bioRxiv2023

LETHAL COVID-19 ASSOCIATES WITH RAAS-INDUCED INFLAMMATION FOR MULTIPLE ORGAN DAMAGE INCLUDING MEDIASTINAL LYMPH NODES

Lethal COVID-19 outcomes are most often attributed to classic cytokine storm and attendant excessive immune signaling. We re-visit this question using RNA sequencing in nasopharyngeal and 40 autopsy samples from COVID-19-positive and negative individuals. In nasal swabs, the top 100 genes which significantly correlated with COVID-19 viral load, include many canonical innate immune genes. However, 22 much less studied "non-canonical" genes are found and despite the absence of viral transcripts, subsets of these are upregulated in heart, lung, kidney, and liver, but not mediastinal lymph nodes. An important regulatory potential emerges for the non-canonical genes for over-activating the renin-angiotensin-activation-system (RAAS) pathway, resembling this phenomenon in hereditary angioedema (HAE) and its overlapping multiple features with lethal COVID-19 infections. Specifically, RAAS overactivation links increased fibrin deposition, leaky vessels, thrombotic tendency, and initiating the PANoptosis death pathway, as suggested in heart, lung, and especially mediastinal lymph nodes, with a tightly associated mitochondrial dysfunction linked to immune responses. For mediastinal lymph nodes, immunohistochemistry studies validate the transcriptomic findings showing abnormal architecture, excess fibrin and collagen deposition, and pathogenic fibroblasts. Further, our findings overlap findings in SARS-CoV-2 infected hamsters, C57BL/6 and BALB/c mouse models, and importantly peripheral blood mononuclear cell (PBMC) and whole blood samples from COVID-19 patients infected with early variants and later SARS-CoV-2 strains. We thus present cytokine storm in lethal COVID-19 disease as an interplay between upstream immune gene signaling producing downstream RAAS overactivation with resultant severe organ damage, especially compromising mediastinal lymph node function.

immunology↗

bioRxiv2022

Targeted Down Regulation Of Core Mitochondrial Genes During SARS-CoV-2 Infection

Defects in mitochondrial oxidative phosphorylation (OXPHOS) have been reported in COVID-19 patients, but the timing and organs affected vary among reports. Here, we reveal the dynamics of COVID-19 through transcription profiles in nasopharyngeal and autopsy samples from patients and infected rodent models. While mitochondrial bioenergetics is repressed in the viral nasopharyngeal portal of entry, it is up regulated in autopsy lung tissues from deceased patients. In most disease stages and organs, discrete OXPHOS functions are blocked by the virus, and this is countered by the host broadly up regulating unblocked OXPHOS functions. No such rebound is seen in autopsy heart, results in severe repression of genes across all OXPHOS modules. Hence, targeted enhancement of mitochondrial gene expression may mitigate the pathogenesis of COVID-19. One-Sentence SummaryCovid-19 is associated with targeted inhibition of mitochondrial gene transcription.

molecular biology↗

bioRxiv2021

The Great Deceiver: miR-2392's Hidden Role in Driving SARS-CoV-2 Infection

MicroRNAs (miRNAs) are small non-coding RNAs involved in post-transcriptional gene regulation that have a major impact on many diseases and provides an exciting avenue towards antiviral therapeutics. From patient transcriptomic data, we have discovered a circulating miRNA, miR-2392, that is directly involved with SARS-CoV-2 machinery during host infection. Specifically, we show that miR-2392 is key in driving downstream suppression of mitochondrial gene expression, increasing inflammation, glycolysis, and hypoxia as well as promoting many symptoms associated with COVID-19 infection. We demonstrate miR-2392 is present in the blood and urine of COVID-19 positive patients, but not detected in COVID-19 negative patients. These findings indicate the potential for developing a novel, minimally invasive, COVID-19 detection method. Lastly, using in vitro human and in vivo hamster models, we have developed a novel miRNA-based antiviral therapeutic that targets miR-2392, significantly reduces SARS-CoV-2 viability in hamsters and may potentially inhibit a COVID-19 disease state in humans.

systems biology↗

bioRxiv2020

Viral-mediated ubiquitination impacts interactions of host proteins with viral RNA and promotes viral RNA processing

Viruses promote infection by hijacking host ubiquitin machinery to counteract or redirect cellular processes. Adenovirus encodes two early proteins, E1B55K and E4orf6, that together co-opt a cellular ubiquitin ligase complex to overcome host defenses and promote virus production. Adenovirus mutants lacking E1B55K or E4orf6 display defects in viral RNA processing and protein production, but previously identified substrates of the redirected ligase do not explain these phenotypes. Here we used a quantitative proteomics approach to identify substrates of E1B55K/E4orf6-mediated ubiquitination that facilitate RNA processing. While all currently known cellular substrates of E1B55K/E4orf6 are degraded by the proteasome, we uncovered RNA-binding proteins (RBPs) as high-confidence substrates which are not decreased in overall abundance. We focused on two RBPs, RALY and hnRNP-C, which we confirm are ubiquitinated without degradation. Knockdown of RALY and hnRNP-C increased levels of viral RNA splicing, protein abundance, and progeny production during infection with E1B55K-deleted virus. Furthermore, infection with virus deleted for E1B55K resulted in increased interaction of hnRNP-C with viral RNA, and attenuation of viral RNA processing. These data suggest viral-mediated ubiquitination of RALY and hnRNP-C relieves a restriction on viral RNA processing, revealing an unexpected role for non-degradative ubiquitination in manipulation of cellular processes during virus infection.

microbiology↗