Biology subjects

England, H.

Publications and source records attributed to England, H..

4 recordsLinked to original sources

bioRxiv2023

Overexpression of IkappaBalpha modulates NF-kappaB activation of inflammatory target gene expression

Cells respond to inflammatory stimuli such as cytokines by activation of the nuclear factor-{kappa}B (NF-{kappa}B) signalling pathway, resulting in oscillatory translocation of the transcription factor p65 between nucleus and cytoplasm to mediate immune response. We investigate the relationship between p65 and inhibitor-{kappa}B (I{kappa}B) protein levels and dynamic properties of the system, and how this interaction impacts on the expression of key inflammatory genes. Using bacterial artificial chromosomes, we developed new cell models of I{kappa}B-eGFP protein overexpression in a native genomic context. We find that cells with high levels of the negative regulator I{kappa}B remain responsive to inflammatory stimuli and maintain dynamics for both p65 and I{kappa}B. In contrast, canonical target gene expression is dramatically reduced by overexpression of I{kappa}B, but can be partially rescued by overexpression of p65. Treatment with leptomycin B to promote nuclear accumulation of I{kappa}B also suppresses canonical target gene expression, suggesting a mechanism in which nuclear I{kappa}B accumulation prevents productive p65 interaction with promoter binding sites. This causes reduced target promoter binding and gene transcription, which we validate by chromatin immune precipitation and in primary cells. Overall, we show how inflammatory gene transcription is modulated by the expression levels of both I{kappa}B and p65, and that transcription can be partially decoupled from p65 protein dynamics. This results in an anti-inflammatory effect on transcription, demonstrating a broad mechanism to modulate the strength of inflammatory response.

cell biology↗

bioRxiv2021

Colour Vision in Stomatopod Crustaceans: more questions than answers

Stomatopod crustaceans, or mantis shrimps, are known for their extensive range of spectral sensitivities but relatively poor spectral discrimination. Instead of the colour-opponent mechanism of other colour vision systems, the 12 narrow-band colour channels they possess may underlie a different method of colour processing. We investigated one hypothesis, in which the photoreceptors are proposed to act as individual wave-band detectors, interpreting colour as a parallel pattern of photoreceptor activation, rather than a ratiometric comparison of individual signals. This different form of colour detection has been used to explain previous behavioural tests in which low saturation blue was not discriminated from grey potentially because of similar activation patterns. Results here, however, indicate that the stomatopod, Haptosquilla trispinosa was able to easily distinguish several colours, including blue of both high and low saturation, from greys. The animals did show a decrease in performance over time in an artificially lit environment, indicating plasticity in colour discrimination ability. This rapid plasticity, most likely the result of a change in opsin (visual pigment) expression, has now been noted in several animal lineages (both invertebrate and vertebrate) and is a factor we suggest needing care and potential re-examination in any colour-based behavioural tests. As for stomatopods, it remains unclear why they achieve poor colour discrimination using the most comprehensive set of spectral sensitivities in the animal kingdom and also what form of colour processing they may utilise.

animal behavior and cognition↗

bioRxiv2019

Macrophage-specific NF-kappa B activation dynamics can segregate inflammatory bowel disease patients

The heterogeneous nature of inflammatory bowel disease (IBD) presents challenges, particularly when choosing therapy. Activation of the NF-{kappa}B transcription factor is a highly-regulated, dynamic event in IBD pathogenesis. We expressed the human NF-{kappa}B/p65 subunit in blood-derived macrophages, using lentivirus. Confocal imaging of p65 activation revealed that a higher proportion of macrophages from Crohns patients responded to lipid-A compared to controls. In contrast, cells from ulcerative colitis (UC) patients exhibited a shorter duration of p65 nuclear localisation compared to healthy controls and Crohns donors. Using a similar lentivirus approach, NF-{kappa}B-regulated luciferase was expressed in patient macrophages, isolated from frozen peripheral blood mononuclear cell samples. Following activation, samples could be segregated into three clusters based on the NF-{kappa}B-regulated luciferase response. The majority of UC samples appeared in hypo-responsive cluster 1, with Crohns patients representing the majority of hyper-responsive cluster 3. A positive correlation was seen between NF-{kappa}B-induced luciferase activity and cytokine levels released to medium from stimulated macrophages, but not in serum or biopsy. Analysis of macrophage cytokine responses and patient metadata revealed a strong correlation between Crohns patients who smoked and hyper-activation of p65. These in vitro dynamic assays of NF-{kappa}B activation in blood-derived macrophages segregate IBD patients into groups with different phenotypes and therefore may help determine response to therapy. Significance statementThis manuscript describes two dynamic assays of NF-{kappa}B activation in blood-derived macrophages that can segregate IBD patients into groups with different phenotypes. For the first time we introduce the use of dynamic measurements of a transcription factor activation as a method to stratify patients and we are confident that our approach will lead in future to early patient stratification and prediction of treatment outcome.

immunology↗

bioRxiv2019

Identification of a novel therapeutic agent for Inflammatory Bowel Disease guided by systems medicine

ObjectiveInflammatory bowel diseases cause significant morbidity and mortality. Aberrant NF-{kappa}B signalling is strongly associated with these conditions, and several established drugs influence the NF-{kappa}B signalling network to exert their effect. This study aimed to identify drugs which alter NF-{kappa}B signalling and may be repositioned for use in inflammatory bowel disease. DesignThe SysmedIBD consortium established a novel drug-repurposing pipeline based on a combination of in-silico drug discovery and biological assays targeted at demonstrating an impact on NF-kappaB signalling, and a murine model of IBD. ResultsThe drug discovery algorithm identified several drugs already established in IBD, including corticosteroids. The highest-ranked drug was the macrolide antibiotic Clarithromycin, which has previously been reported to have anti-inflammatory effects in aseptic conditions. Clarithromycins effects were validated in several experiments: it influenced NF-{kappa}B mediated transcription in murine peritoneal macrophages and intestinal enteroids; it suppressed NF-{kappa}B protein shuttling in murine reporter enteroids; it suppressed NF-{kappa}B (p65) DNA binding in the small intestine of mice exposed to LPS, and it reduced the severity of dextran sulphate sodium-induced colitis in C57BL/6 mice. Clarithromycin also suppressed NF-{kappa}B (p65) nuclear translocation in human intestinal enteroids. ConclusionsThese findings demonstrate that in-silico drug repositioning algorithms can viably be allied to laboratory validation assays in the context of inflammatory bowel disease; and that further clinical assessment of clarithromycin in the management of inflammatory bowel disease is required.

systems biology↗