Biology subjects

Pham, E.

Publications and source records attributed to Pham, E..

5 recordsLinked to original sources

bioRxiv2023

Genomes of the Autonomous Parvovirus Minute Virus of Mice Induce Replication Stress Through RPA Exhaustion

The oncolytic autonomous parvovirus Minute Virus of Mice (MVM) establishes infection in the nuclear environment by usurping host DNA Damage Response (DDRs) in the vicinity of cellular DNA break sites. MVM replication induces a global cellular DDR that is dependent on signaling by the ATM kinase and inactivates the cellular ATR-kinase pathway. However, the mechanism of how MVM generates cellular DNA breaks remains unknown. Using single molecule DNA Fiber Analysis, we have discovered that MVM infection leads to a shortening of host replication forks as infection progresses, as well as induction of replication stress prior to the initiation of virus replication. Ectopically expressed viral non-structural proteins NS1 and NS2 are sufficient to cause host-cell replication stress, as is the presence of UV-inactivated non-replicative MVM genomes. The host single-stranded DNA binding protein Replication Protein A (RPA) associates with the UV-inactivated MVM genomes, suggesting MVM genomes might serve as a sink for cellular stores of RPA. Overexpressing RPA in host cells prior to UV-MVM infection rescues DNA fiber lengths and increases MVM replication, confirming that MVM genomes deplete RPA stores to cause replication stress. Together, these results indicate that the presence of ssDNA in the nucleus generated by MVM genomes and viral proteins induces replication stress in the host cell through RPA exhaustion, rendering the host genome vulnerable to additional DNA breaks. AUTHOR SUMMARYParvoviruses are used in the clinic to design recombinant gene therapy vectors and as oncolytic agents. The autonomous parvovirus MVM utilizes the host cells DNA damage response machinery to replicate in host cells and cause additional DNA breaks. However, the mechanism of MVM-induced DNA damage remains unknown. We have discovered that MVM sequesters the host DNA repair protein RPA, which normally associates with single stranded DNA in the nucleus, rendering the host genome susceptible to replication stress. Our study provides insights into the mechanisms utilized by single-stranded DNA viruses to amplify host-cell DNA damage.

microbiology↗

bioRxiv2022

In BRAFV600E-mutated colorectal cancer cells, fascin1 restructures adherens junctions to drive Wnt activation

Fascin1 expression in colorectal carcinomas (CRCs) is linked to a clinically aggressive disease with poor prognosis. Despite that fascin1s role in the etiology of CRCs has not been directly investigated. We show fascin1 expression in one-third of all CRCs underscoring the critical need to identify fascin1s function in colorectal carcinogenesis. Here, we identify for the first time, fascin1s role as a mechanosensor that modulates CRC cell adherens junction (AJ) plasticity to induce tumor growth and metastasis. We show that fascin1 expression drives protein sorting to transform AJ mechanotransduction and we reveal how these force-sensitive pathways activate oncogenic signaling in CRC cells. We made the novel finding that AJ remodeling by fascin1 also controls "collective plasticity" and bidirectional cell migration. Few studies have examined AJ plasticity in cancer cells, which remains poorly understood and has not been therapeutically targeted. Our findings could have widespread implications for understanding and treating metastatic carcinomas.

cancer biology↗

bioRxiv2021

Dysfunctional temporal stages of eye-gaze perception in adults with ADHD: a high-density EEG study

ADHD have been associated with social cognitive impairments across the lifespan, but no studies have specifically addressed the presence of abnormalities in eye-gaze processing in the adult brain. This study investigated the neural basis of eye-gaze perception in adults with ADHD using event-related potentials (ERP). Twenty-three ADHD and 23 controls performed a delayed face-matching task with neutral faces that had either direct or averted gaze. ERPs were classified using microstate analyses. ADHD and controls displayed similar P100 and N170 microstates. ADHD was associated with cluster abnormalities in the attention-sensitive P200 to direct gaze, and in the N250 related to facial recognition. For direct gaze, source localization revealed reduced activity in ADHD for the P200 in the left/midline cerebellum, as well as in a cingulate-occipital network at the N250. These results suggest brain impairments involving eye-gaze decoding in adults with ADHD, suggesting that neural deficits persist across the lifespan.

neuroscience↗

bioRxiv2020

Genetics of white color and iridophoroma in "Lemon Frost" leopard geckos

The squamates (lizards and snakes) are close relatives of birds and mammals, with more than 10,000 described species that display extensive variation in a number of important biological traits, including coloration, venom production, and regeneration. Due to a lack of genomic tools, few genetic studies in squamates have been carried out. The leopard gecko1, Eublepharis macularius, is a popular companion animal, and displays a variety of coloration patterns. We took advantage of a large breeding colony and used linkage analysis, synteny, and homozygosity mapping to investigate a spontaneous semi-dominant mutation, "Lemon Frost", that produces white coloration and causes skin tumors (iridophoroma). We localized the mutation to a single locus which contains a strong candidate gene, SPINT12,3, a tumor suppressor implicated in human skin cutaneous melanoma (SKCM) and over-proliferation of epithelial cells in mice and zebrafish4-16. Our work establishes the leopard gecko as a tractable genetic system and suggests that a tumor suppressor in melanocytes in humans can also suppress tumor development in iridophores in lizards.

genetics↗

bioRxiv2018

Laboratory Validation of a Clinical Metagenomic Sequencing Assay for Pathogen Detection in Cerebrospinal Fluid

Metagenomic next-generation sequencing (mNGS) for pan-pathogen detection has been successfully tested in proof-of-concept case studies in patients with acute illness of unknown etiology, but to date has been largely confined to research settings. Here we developed and validated an mNGS assay for diagnosis of infectious causes of meningitis and encephalitis from cerebrospinal fluid (CSF) in a licensed clinical laboratory. A clinical bioinformatics pipeline, SURPI+, was developed to rapidly analyze mNGS data, automatically report detected pathogens, and provide a graphical user interface for evaluating and interpreting results. We established quality metrics, threshold values, and limits of detection of between 0.16 - 313 genomic copies or colony forming units per milliliter for each representative organism type. Gross hemolysis and excess host nucleic acid reduced assay sensitivity; however, a spiked phage used as an internal control was a reliable indicator of sensitivity loss. Diagnostic test accuracy was evaluated by blinded mNGS testing of 95 patient samples, revealing 73% sensitivity and 99% specificity compared to original clinical test results, with 81% positive percent agreement and 99% negative percent agreement after discrepancy analysis. Subsequent mNGS challenge testing of 20 positive CSF samples prospectively collected from a cohort of pediatric patients hospitalized with meningitis, myelitis, and/or encephalitis showed 92% sensitivity and 96% specificity relative to conventional microbiological testing of CSF in identifying the causative pathogen. These results demonstrate the analytic performance of a laboratory-validated mNGS assay for pan-pathogen detection, to be used clinically for diagnosis of neurological infections from CSF.

genomics↗