Biology subjects

Barker, S.

Publications and source records attributed to Barker, S..

6 recordsLinked to original sources

bioRxiv2026

Inhibition of CDK7, CDK9 or CDK12 affects phosphorylation of human SPT5.

SPT5, complexed with SPT4 in the DRB-sensitivity-inducing factor, DSIF, is a critical regulator of transcription elongation that acts negatively in the unphosphorylated form and positively when phosphorylated. In vivo, phosphorylation of multiple sites in the largely unstructured linker and C-terminal regions of the protein facilitates release from an early elongation checkpoint and processive transcription elongation. Transcription elongation factors, including the PAF complex (PAF1C), interact with phosphorylated SPT5 and phosphorylation of residues S666 and T806 are associated with the positive elongation functions of SPT5. Here, using in vitro phosphorylation of regions of the SPT5 C-terminus followed by GST-mediated pull-down we confirm that the RTF1 subunit of PAF1C interacts with residues 750-850 only when phosphorylated. Antibodies we raised to phospho-S666 or phospho-T806 are specific for these sites phosphorylated in vitro and in vivo. Western analysis of endogenous SPT5 in HEK293 cells using these antibodies before and after inhibition of analog-sensitive versions of CDK7, CDK9 and CDK12 indicate that all three kinases play roles in the phosphorylation of SPT5, although inhibition of CDK9 has the most marked effect. CDK9 inhibition also inhibits the association of PAF1C subunits with SPT5 in vivo and affects phosphorylation of chromatin-associated SPT5 as measured by chromatin immunoprecipitation (ChIP).

molecular biology↗

bioRxiv2026

Time-Resolved Profiling: Metabolic Adaptation & Morphology-Specific Drug Response in MCF-7 spheroids

Three-dimensional (3D) tumor models exhibit drug responses that differ from conventional 2D cultures. However, how cellular metabolism dynamically evolves across culture models and during drug treatment remains poorly understood. We compared the responses of MCF-7 cells in 2D and 3D environments to 5-fluorouracil (5-FU), combining {superscript 1}H NMR cellular metabolomics with viable cell counts, the GLUT1-positive population, gene expression, ATP activity, and morphology. Principal component analysis revealed that culture dimensionality, rather than 5-FU treatment, was the primary driver of metabolic flux variation. 3D spheroids exhibited higher glycolytic flux at 72h. Importantly, this elevated glycolysis reflected a higher per-cell flux in larger spheroids rather than an increased cell number. We further observed a higher proportion of GLUT1-positive cells and increased HK2 expression in 3D culture, together with an epithelial phenotype characterized by increased CDH1 and decreased VIM expression. Functionally, 3D displayed maintaining higher cell viability, ATP activity following treatment. Together, these findings suggest that 3D architecture promotes a metabolically defensive phenotype and cellular metabolic behaviors is associated with morphology, which may inform future drug screening model selection. BlurbTime-resolved NMR metabolomics reveals that 3D culture architecture, rather than 5-FU treatment, defines the metabolic phenotype of MCF-7 cells, linking spheroid morphology with per-cell glycolytic activity, ATP preservation, and reduced chemotherapy sensitivity. Synopsis O_FIG O_LINKSMALLFIG WIDTH=197 HEIGHT=200 SRC="FIGDIR/small/740505v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@15cfa5corg.highwire.dtl.DTLVardef@138ca69org.highwire.dtl.DTLVardef@170d0d8org.highwire.dtl.DTLVardef@19f3572_HPS_FORMAT_FIGEXP M_FIG C_FIG Graphical abstract -- Time-resolved metabolic and phenotypic responses of 2D and 3D MCF-7 cultures to 5-fluorouracil. Bullet pointsO_LITime-resolved {superscript 1}H NMR cellular metabolomics combined with multilevel phenotypic readouts reveals that culture dimensionality, rather than 5-FU chemotherapy, is the dominant determinant of metabolic phenotype in MCF-7 breast cancer cells. C_LIO_LI3D spheroids develop a glycolysis-dominant metabolic state with markedly elevated GLUT1 populations and HK2 transcript, accompanied by substantially reduced sensitivity to 5-FU compared with 2D monolayers. C_LIO_LISpheroid morphology correlates with cellular metabolic flux: larger and more elongated spheroids exhibit higher glycolytic activity on a per cell, independent of spheroid cell number. C_LIO_LIThe 3D-defined metabolic state buffers ATP under 5-FU stress and reinforces an epithelial transcriptional program (CDH1{uparrow}, VIM{downarrow}), arguing that culture architecture must be considered when interpreting preclinical drug responses. C_LI

cancer biology↗

bioRxiv2025

Distinct Virome and Bacteriome Profiles of Term and Preterm Placentas from African, Asian and European Women

While the human placental microbiome has been extensively studied, information regarding the virome remains limited. The association between subclinical placental viral infections and spontaneous preterm birth (PTB) is poorly understood. Hence, we examined fresh placenta samples from women in the UK, Bangladesh, and South Africa to elucidate the virome profiles and association with spontaneous PTB. We employed shotgun metagenomics, real-time PCR, and Gram staining, complemented by bioinformatics, to detect both viruses and bacteria in the decidua and chorionic villous tissues. We did not identify any known pathogenic viruses in 236 placental samples, except for one preterm placenta from the Bangladeshi cohort, where human herpesvirus 6 was detected. The majority of the viruses were bacteriophages present at very low abundance and low frequencies across the three cohorts. Lambdavirus lambda was detected in 14% of UK and 11% of Bangladeshi samples but was absent in South African samples. Papiine betaherpesvirus 3 was identified in 62.5% of South African placentas. Punavirus (29.1%) and Streptococcus phages (16.67%) were also prevalent in South African samples. A variety of bacterial species including Moraxella osloensis, Delftia lacustris, Cutibacterium acnes, Escherichia coli and Staphylococcus epidermidis were detected in samples across the three cohorts in variable frequencies but without any notable tissue immune responses. None of the viruses or bacteria exhibited associations with PTB, except D. lacustris, which was significantly more abundant in preterm compared to term placentas in the Bangladeshi cohort. We conclude that the human placenta harbours a virome predominated by bacteriophages and that subclinical viral infection in spontaneous PTB is rare.

microbiology↗

bioRxiv2025

Metagenomic resolution of spotted-fever group Rickettsia tasmanensis and novel DNA viruses in Australian wildlife ticks, with spatial modelling of Rickettsia exposure zones.

Australia's spotted fever group (SFG) rickettsiae cause significant illness, yet microbial diversity in wildlife ticks remains incompletely understood. Metagenomic sequencing of nine tick pools representing three Ixodidae species, Ixodes tasmani, I. holocyclus, and Haemaphysalis bancrofti, from Australian wildlife revealed a genome of a novel SFG Rickettsia species from I. tasmani ticks collected from koalas in New South Wales. Phylogenomic analysis confirmed this as a distinct species closely related to Rickettsia tasmaniensis fragments previously reported from Tasmanian devils. Additionally, we discovered four novel DNA Anellovirus species forming a new genus 'Sintorquevirus' and one Circovirus from likely vertebrate blood meals, revealing wildlife viral diversity. Species distribution modelling of I. tasmani for assessing Rickettsia risk revealed suitable habitat along Australia's eastern coastlines, with overlap between vector distribution, marsupial hosts, and population centres. This identified potential Candidatus R. tasmanensis exposure zones in coastal regions where positive samples originated, highlighting areas for surveillance.

microbiology↗

bioRxiv2019

Drug screening to identify compounds to act as co-therapies for the treatment of pathogenic Burkholderia

Burkholderia pseudomallei is a soil-dwelling organism present throughout the tropics, and is the causative agent of melioidosis, a disease that is believed to kill 89,000 people per year. It is naturally resistant to most currently available antibiotics. The most efficacious treatment for melioidosis requires at least two weeks of intravenous treatment with ceftazidime or meropenem. This places a large treatment burden on the predominantly middle income nations where the majority of disease occurs. We have established a high-throughput assay for compounds that could be used as a co-therapy to potentiate the effect of ceftazidime, using the related non-pathogenic bacterium Burkholderia thailandensis as a surrogate. Optimization of the assay gave a Z factor of 0.68. We screened a library of 61,250 compounds, and identified 29 compounds with a pIC50 (-log10(IC50)) greater than five. Detailed investigation allowed us to down select to six \"best in class\" compounds, which included the licensed drug chloroxine. Co-treatment of B. thailandensis with ceftazidime and chloroxine reduced culturable cell numbers by two orders of magnitude over 48 hours compared to treatment with ceftazidime alone. Hit expansion around chloroxine was performed using commercially available compounds. Minor modifications to the structure abolished activity, suggesting that chloroxine likely acts against a specific target. Finally, preliminary data also demonstrates the utility of chloroxine to act as a co-therapy to potentiate the effect of ceftazidime against B. pseudomallei. This approach successfully identified potential co-therapies for a recalcitrant Gram-negative bacterial species. Our assay could be used more widely to aid in chemotherapy against these bacteria.

microbiology↗

bioRxiv2018

Genetic dissection of assortative mating behavior

The evolution of new species is made easier when traits under divergent ecological selection are also mating cues. Such ecological mating cues are now considered more common than previously thought, but we still know little about the genetic changes underlying their evolution, or more generally about the genetic basis for assortative mating behaviors. The warning patterns of Heliconius melpomene and H. cydno are under disruptive selection due to increased predation of non-mimetic hybrids, and are used during mate recognition. We carried out a genome-wide quantitative trait locus (QTL) analysis of preference behaviors between these species and showed that divergent male preference has a simple genetic basis. Three QTLs each explain a large proportion of the differences in preference behavior observed between the parental species. Two of these QTLs are on chromosomes with major color pattern genes, including one that is tightly associated with the gene optix. Different loci influence different aspects of attraction, suggesting that behavioral isolation in Heliconius involves the evolution of independently segregating modules, similar to those for the corresponding wing pattern cues. Hybridization and subsequent sharing of wing pattern loci has played an important role during adaptation and speciation in Heliconius butterflies. The existence of large effect preference loci could similarly assist the evolution of novel behavioral phenotypes through recombination and introgression, and should facilitate rapid speciation.

evolutionary biology↗