Biology subjects

Harris, F.

Publications and source records attributed to Harris, F..

3 recordsLinked to original sources

bioRxiv2023

RNAP promoter search and transcription kinetics in live E. coli cells

Bacterial transcription has been studied extensively in vitro, which has provided indepth insight regarding transcription mechanisms. However, the live cell environment may impose different rules on transcription than the homogenous and simplified in vitro environment. How an RNA polymerase (RNAP) molecule searches rapidly through the vast nonspecific chromosomal DNA in the three-dimensional nucleoid space and binds a specific promoter sequence remains elusive. The kinetics of transcription in vivo could also be impacted by specific cellular environments including nucleoid organization and nutrient availability. In this work, we investigated the promoter search dynamics and transcription kinetics of RNAP in live E. coli cells. Using single-molecule tracking (SMT) and fluorescence recovery after photobleaching (FRAP) and combining with different genetic, drug inhibition, and growth conditions, we observed that RNAPs promoter search is facilitated by nonspecific DNA interactions and largely independent of nucleoid organization, growth condition, transcription activity, or promoter classes. RNAPs transcription kinetics, however, is sensitive to these conditions and mainly modulated at the levels of actively engaged RNAP and the promoter escape rate. Our work establishes a foundation for further mechanistic studies of bacterial transcription in live cells.

biophysics↗

bioRxiv2022

Spatial transcriptomic analysis of childhood ependymoma implicates unresolved wound healing as a driver of tumor progression

Childhood ependymoma (EPN) is a brain tumor that has seen limited improvements in outcome over past decades. The underlying cellular components of EPN have recently been revealed by single cell RNA-sequencing (scRNAseq), providing biological insights. Here we use spatial transcriptomics to comprehensively chart gene expression across the cellular landscape of posterior fossa subgroup A (PFA) EPN, the commonest and most deadly EPN variant, providing novel resolution of cellular heterogeneity and cellular interaction. We reveal that PFA are comprised of epithelial and mesenchymal histological zones each containing a diversity of cellular states. These include co-existing and spatially distinct undifferentiated progenitor-like clusters - a quiescent mesenchymal zone population, and a second highly mitotic progenitor population that is restricted to hypercellular epithelial zones. We show that myeloid cell interaction is the leading cause of mesenchymal transition in PFA, occurring in zones spatially distinct from hypoxia-induced mesenchymal transition, and these distinct EMT-initiating processes were replicated in in-vitro models of PFA. Collectively, our transcriptomic and functional analyses mirror the processes of normal wound healing where PFA mesenchymal and epithelial zones interact with immune subpopulations in cycle of persistent tissue damage response and mitogenic re-epithelialization signals. Spatial transcriptomics advances our understanding of PFA biology, implicating a "wound that will not heal" process as a driver of tumor progression, a new concept that could provide novel targets for effective therapeutic intervention. SignificanceSpatial transcriptomic analysis of the ependymoma tumor microenvironment identifies novel cell populations and interactions, implicating unresolved wound healing as a clinically actionable driver of tumor progression in this refractory childhood brain tumor.

cancer biology↗

bioRxiv2018

Effect of chronic stimulation and stimulus level on temporal processing by cochlear implant listeners

A series of experiments investigated potential changes in temporal processing during the months following activation of a cochlear implant (CI) and as a function of stimulus level. Experiment 1 tested patients on the day of implant activation and two and six months later. All stimuli were presented using direct stimulation of a single apical electrode. The dependent variables were rate discrimination ratios (RDRs) for pulse trains with rates centred on 120 pulses per second (pps), obtained using an adaptive procedure, and a measure of the upper limit of temporal pitch, obtained using a pitch-ranking procedure.\n\nAll stimuli were presented at their most comfortable level (MCL). RDRs decreased from 1.23 to 1.16 and the upper limit increased from 357 to 485 pps from 0 to 2 months post-activation, with no overall change from 2 to 6 months. Because MCLs and hence the testing level increased across sessions, two further experiments investigated whether the performance changes observed across sessions could be due to level differences. Experiment 2 re-tested a subset of subjects at 9 months post-activation, using current levels similar to those used at 0 months. Although the stimuli sounded softer, some subjects showed lower RDRs and/or higher upper limits at this re-test. Experiment 3 measured RDRs and the upper limit for a separate group of subjects at levels equal to 60%, 80%, and 100% of the dynamic range. RDRs decreased with increasing level. The upper limit increased with increasing level for most subjects, with two notable exceptions. Implications of the results for temporal plasticity are discussed, along with possible influences of the effects of level and of across-session learning.

neuroscience↗