Biology subjects

Rees, C.

Publications and source records attributed to Rees, C..

5 recordsLinked to original sources

bioRxiv2026

A bifunctional RNase H protein supports DNA replication and transcription in nucleus-forming Chimalliviridae phages

Jumbo bacteriophages of the Chimalliviridae family replicate their genomes within a proteinaceous phage nucleus that spatially segregates viral DNA replication and transcription from translation. While the phage nucleus protects viral DNA from host defenses, it also imposes unique constraints on genome replication and gene expression. Here we identify the phage-encoded protein Rhc (RNase H-clamp) as a central mediator of genome transactions within the phage nucleus. Using the Escherichia coli infecting bacteriophage Goslar, we show that Rhc functions as an essential component of a non-canonical three-subunit DNA polymerase complex composed of DNAPN, DNAPC and Rhc. Rhc physically interacts with the DNAPC subunit, and both the catalytic activity of Rhc's N-terminal RNase H domain and its C-terminal sliding clamp-derived DNA binding domain are required for genome replication, nuclear maturation, and productive infection. Further, Goslar Rhc also supports transcription within the phage nucleus through a function that is separable from its DNA replication function. Rhc depletion results in reduced transcription of a subset of late genes, in a pattern that suggests a role for Rhc in resolving R-loops. Imaging and cryo-electron tomography reveal that loss of Rhc arrests phage development at a nascent nuclear stage, uncoupling nuclear assembly from maturation. Together, these findings identify Rhc as a multifunctional nuclear factor that mediates both DNA replication and transcription in nucleus-forming jumbo phages, revealing how viruses repurpose a conserved enzymatic activity to coordinate gene expression within a compartmentalized replication environment.

cell biology↗

bioRxiv2026

Use of a plasmid containing a dual gene reporter system to assess the cell hydrophobicity of Listeria monocytogenes

Listeria monocytogenes causes listeriosis in humans and animals and contaminates prepared food by attaching to food processing environments. Therefore, closer monitoring of how the organism adheres to surfaces will help identify ways to prevent it from colonising food-processing environments. To develop new attachment assays, clinical and environmental strains of L. monocytogenes were transformed by inserting a plasmid containing lux, gfp reporter genes and an erythromycin-resistant gene into the parent cells. Transformed cells were grown for 48 hours on brain heart infusion agar plates containing 1-5{micro}g/ml of erythromycin, after which the cells were viewed under a molecular light imager and luminometer. Fluorescent cells containing the gfp, lux, and erythromycin-resistant genes were visible, whereas control cells without the plasmid were not. Transformation efficiency was highest with the environmental strains, and subsequent growth and hydrophobicity tests carried out with the transformed cells in different growth conditions showed that they were able to attach well to solvents when compared to the parent cells. However, the growth rate of the transformed cells was poor, indicating a disruption of cell metabolism. Results show the possibility of real-time monitoring of how cells attach to different surfaces and could lead to a better understanding of the initial colonisation of a surface by the organism.

genetics↗

bioRxiv2025

Mechanistic insights and clinical implications of cross-reactive anti-prophage antibodies and bacterial heteroresistance on phage therapeutic failure

Phage therapy is an exciting strategy against antimicrobial-resistant bacterial infections, but critical knowledge gaps regarding its clinical application persist. Studying a patient with a life-threatening, chronic bacterial infection who failed phage therapy, we uncovered important biological concepts with direct translational impact. Using longitudinal clinical samples, we found that patients can harbour pre-existing antibodies against active prophages induced from the genome of the causative pathogen. Notably, these antibodies can contribute to clinical failure by cross-reacting with and effectively neutralising therapeutic phage. We also uncovered bacterial heteroresistance, characterised by bacterial subpopulations from the initial infection with reduced phage susceptibility, as a further contributor to treatment failure. These findings highlight the intricate interplay between host immunology, bacterial genetic diversity and phage biology, bearing broad significance for clinical phage therapy. Future phage therapy patients, especially those with chronic infections, should be screened for antiphage immunity and bacterial heteroresistance prior to phage treatment.

microbiology↗

bioRxiv2023

Probing the Evolution of Genes Associated With DNA Methylation in Listeria monocytogenes

In the last decade, there have been increased reports of atypical Listeria and the discovery of new species. There are public health concerns that new strains may come with increased pathogenicity. Hence, this study aimed to establish the prevalence, evolutionary lineage and ancestry of a Listeria monocytogenes collection that includes isolates that harbour a unique set of methylase genes. The addition of methyl groups to DNA can interfere with transcription. Allelic-specific lineage analysis and ribotyping with southern hybridization were carried out after which further phylogenetic analysis was performed in silico. Results show that all the methylase strains belonged to Lineage I and were serotypes 4b or 4d. All designated ancestral strains also belonged to Lineage 1. A Listeria monocytogenes plasmid from a serotype 1/2a (Lineage II) contained sequences homologous to that of Lineage I isolates. The methylase nucleotide sequence in the strains studied appears to be highly conserved in Listeria monocytogenes and not yet orthologous among other bacterial genera. It is of epidemiological interest and public benefit if wider or continuous surveillance is carried out to ascertain if these rare strains are linked with increased pathogenesis, food type or geographical region.

genetics↗

bioRxiv2018

Variability and compensation of cardiomycoyte ionic conductances at the population level

Conductances of ion channels and transporters controlling cardiac excitation may vary in a population of subjects with different cardiac gene expression patterns. However, the amount of variability and its origin are not quantitatively known. We propose a new computational method to predict this variability that consists of finding combinations of conductances generating a normal intracellular Ca2+ transient without any constraint on the action potential. Furthermore, we validate experimentally its predictions using the Hybrid Mouse Diversity Panel, a model system of genetically diverse mouse strains that allows us to quantify inter-subject versus intra-subject variability. The method predicts that conductances of inward Ca2+ and outward K+ currents compensate each other to generate a normal Ca2+ transient in good quantitative agreement with current measurements in ventricular myocytes from hearts of different isogenic strains. Our results suggest that a feedback mechanism sensing the aggregate Ca2+ transient of the heart suffices to regulate ionic conductances.

biophysics↗