Biology subjects

White, M. R. H.

Publications and source records attributed to White, M. R. H..

4 recordsLinked to original sources

bioRxiv2024

Domain Specific AI Segmentation of IMPDH2 Rod/Ring Structures in Mouse Embryonic Stem Cells

BackgroundInosine monophosphate dehydrogenase 2 (IMPDH2) is an enzyme that catalyses the rate limiting step of guanine nucleotides. In mouse embryonic stem cells (ESCs) IMPDH2 is held as large multi-protein complexes known as rod-ring (RR) structures that dissociate when ESCs differentiate. Manual analysis of RR structures from confocal microscopy images, although possible, is not feasible on a large scale due to the quantity of RR structures present in each field of view. To address this analysis bottleneck, we have created a fully automatic RR image classification pipeline to segment, characterise and measure feature distributions of these structures in ESCs. ResultsWe find that this model can automatically segment images with a Dice score of over 80% for both rods and rings for in-domain images compared to expert annotation, with a slight drop to 70% for datasets out of domain. Important feature measurements derived from these segmentations show high agreement with the measurements derived from expert annotation, achieving an R2 score of over 90% for counting the number of rings and rods over the dataset. ConclusionsWe have established for the first time a quantitative baseline for RR distribution in pluripotent ESCs and have made a pipeline available for training to be applied to other models in which RR remain an open topic of study.

bioinformatics↗

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↗

bioRxiv2019

Multiplexing information flow through dynamic signalling systems

We consider how a signalling system can act as an information hub by multiplexing information arising from multiple signals. We formally define multiplexing, mathematically characterise which systems can multiplex and how well they can do it. While the results of this paper are theoretical, to motivate the idea of multiplexing, we provide experimental evidence that tentatively suggests that the NF-{kappa}B transcription factor can multiplex information about changes in multiple signals. We believe that our theoretical results may resolve the apparent paradox of how a system like NF-{kappa}B that regulates cell fate and inflammatory signalling in response to diverse stimuli can appear to have the low information carrying capacity suggested by recent studies on scalar signals. In carrying out our study, we introduce new methods for the analysis of large, nonlinear stochastic dynamic models, and develop computational algorithms that facilitate the calculation of fundamental constructs of information theory such as Kullback-Leibler divergences and sensitivity matrices, and link these methods to new theory about multiplexing information. We show that many current models such as those of the NF-{kappa}B system cannot multiplex effectively and provide models that overcome this limitation using post-transcriptional modifications.

systems biology↗

bioRxiv2019

ER stress-linked autophagy stabilizes apoptosis effector PERP and triggers its co-localization with SERCA2b at ER-plasma membrane junctions

Specific molecular interactions that underpin the switch between ER stress-triggered autophagy-mediated cellular repair and cellular death by apoptosis are not characterized. This study reports the unexpected interaction elicited by ER stress between the plasma membrane (PM)-localized apoptosis effector PERP and the ER Ca2+ pump SERCA2b. We show that the p53 effector PERP, which specifically induces apoptosis when expressed above a threshold level, has a heterogeneous distribution across the PM of un-stressed cells and is actively turned over by the lysosome. PERP is upregulated following sustained starvation-induced autophagy, which precedes the onset of apoptosis indicating that PERP protein levels are controlled by a lysosomal pathway that is sensitive to cellular physiological state. Furthermore, ER stress stabilizes PERP at the PM and induces its increasing co-localization with SERCA2b at ER-PM junctions. The findings highlight a novel crosstalk between pro-survival autophagy and pro-death apoptosis pathways and identify, for the first time, accumulation of an apoptosis effector to ER-PM junctions in response to ER stress.

molecular biology↗