Biology subjects

Dunning, M.

Publications and source records attributed to Dunning, M..

5 recordsLinked to original sources

bioRxiv2025

TWIST1 drives endothelial-to-mesenchymal-transition to stabilize atherosclerotic plaques

Atherosclerosis progresses from fatty streaks to complex plaques, with rupture leading to life-threatening complications. Endothelial-to-mesenchymal transition (EndMT) is associated with advanced atherosclerotic plaques, but its role in plaque progression remains unclear. To investigate this, we analyzed the role of TWIST1, a key EndMT-driving transcription factor, in plaque development. Using single-cell RNA sequencing of atherosclerotic plaques from hypercholesterolemic mice with inducible deletion of Twist1 from endothelial cells (Twist1ECKO ApoE-/-) we demonstrate that Twist1 regulates endothelial cell heterogeneity by promoting cell states expressing EndMT markers. Cell tracking confirmed that Twist1 induces EndMT in advanced plaques. Mechanistically, we found that TWIST1 contributes to EndMT by promoting endothelial migration and proliferation through the transcriptional coactivator PELP1. Additionally, TWIST1 induces AEBP1 transcription, which upregulates COL4A1 to drive endothelial proliferation. Analyses of murine brachiocephalic plaques show that endothelial Twist1 promotes plaque growth, collagen deposition, and ACTA2-positive cell accumulation, hallmarks of plaque stability, while reducing necrosis and macrophage infiltration, features of plaque instability. Moreover, TWIST1 expression was associated with asymptomatic human carotid atherosclerosis and predicts favourable clinical outcomes. These findings challenge the prevailing view that EndMT destabilizes plaques, by suggesting that TWIST1-driven EndMT can promote plaque stability, offering new insights into atherosclerosis pathophysiology and therapeutic potential.

pathology↗

bioRxiv2024

Cooperation between Intrinsically Disordered Regions regulates CBP condensate behaviour.

Intrinsically disordered regions (IDRs) have emerged as crucial regulators of protein function, allowing proteins to sense and respond to their environment. Creb binding protein (CBP) and EP300 (p300) are transcription coactivators that regulate gene expression in multicellular organisms, following their recruitment to cis-regulatory elements. CBP and p300 contain large IDRs, however little is known about how these different IDRs work together to regulate CBP function. Here, we show that CBP-IDRs cooperate to control different aspects of CBP behaviour in the nucleus, by regulating the properties of fluid-like condensates formed by endogenous CBP. We show how IDRs with different sequence properties make unique contributions to CBP behaviour by establishing a balance between positive and negative regulation of CBP condensates. These conflicting interactions are functionally important, shaping CBPs response to factors such as lysine acetylation, that influence condensate formation. When disrupted, regulatory CBP-IDRs change how CBP interacts with chromatin, alter patterns of CBP-dependent histone acetylation and change gene expression. Together, our work highlights how IDRs with different sequences, spatially segregated in the same protein, can cooperate to shape protein function.

molecular biology↗

bioRxiv2022

SpheroidAnalyseR - an online platform for analysing data from 3D spheroids or organoids grown in 96-well plates

Spheroids and organoids are increasingly popular three-dimensional (3D) cell culture models. Spheroid models are more physiologically relevant to a tumour compared to two-dimensional (2D) cultures and organoids are a simplified version of an organ with similar composition. Both can be used in cell migration studies, disease modelling and drug discovery. A drawback of these models is, however, the lack of appropriate analytical tools for high throughput imaging and analysis over a time course. To address this, we have developed an R Shiny app called SpheroidAnalyseR: a simple, fast, effective open-source app that allows the analysis of spheroid or organoid size data generated in a 96-well format. SpheroidAnalyseR processes and analyses datasets of image measurements that can be obtained via a bespoke software, described herein, that automates spheroid imaging and quantification using the Nikon A1R Confocal Laser Scanning Microscope. However, templates are provided to enable users to input spheroid image measurements obtained by user-preferred methods. SpheroidAnalyseR facilitates outlier identification and removal followed by graphical visualisation of spheroid measurements across multiple predefined parameters such as time, cell-type and treatment(s). Spheroid imaging and analysis can, thus, be reduced from hours to minutes, removing the requirement for substantial manual data manipulation in a spreadsheet application. The combination of spheroid generation in 96-well ultra-low attachment microplates, imaging using our bespoke software, and analysis using SpheroidAnalyseR toolkit allows high throughput, longitudinal quantification of 3D spheroid growth whilst minimising user input and significantly improving the efficiency and reproducibility of data analysis. Our bespoke imaging software is available from https://github.com/GliomaGenomics. SpheroidAnalyseR is available at https://spheroidanalyser7.azurewebsites.net, and the source code found at https://github.com/GliomaGenomics. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=96 SRC="FIGDIR/small/481039v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@1f779fborg.highwire.dtl.DTLVardef@698103org.highwire.dtl.DTLVardef@94f252org.highwire.dtl.DTLVardef@b1180e_HPS_FORMAT_FIGEXP M_FIG Graphical abstract - SpheroidAnalyseR main steps. A summary of the process followed within the SpheroidAnalyseR app. Dotted boxes are the steps required to produce the input data for SpheroidAnalyseR. Solid boxes show the 5 main steps of SpheroidAnalyseR: Data input, Preview, Outlier removal, Merging and Plotting. C_FIG

cell biology↗

bioRxiv2020

JAG1-NOTCH4 MECHANOSENSING DRIVES ATHEROSCLEROSIS

Endothelial cell (EC) sensing of fluid shear stress regulates atherosclerosis, a disease of arteries that causes heart attack and stroke. Atherosclerosis preferentially develops at regions of arteries exposed to low oscillatory shear stress (LOSS), whereas high shear regions are protected. We show using inducible EC-specific genetic deletion in hyperlipidaemic mice that the Notch ligands JAG1 and DLL4 have opposing roles in atherosclerosis. While endothelial Jag1 promoted atherosclerosis at sites of LOSS, endothelial Dll4 was atheroprotective. Analysis of porcine and murine arteries and cultured human coronary artery EC exposed to experimental flow revealed that JAG1 and its receptor NOTCH4 are strongly upregulated by LOSS. Functional studies in cultured cells and in mice with EC-specific deletion of Jag1 show that JAG1-NOTCH4 signalling drives vascular dysfunction by repressing endothelial repair. These data demonstrate a fundamental role for JAG1-NOTCH4 in sensing LOSS during disease, and suggest therapeutic targeting of this pathway to treat atherosclerosis.Competing Interest StatementThe authors have declared no competing interest.View Full Text

pathology↗

bioRxiv2019

Failure to clear developmental apoptosis contributes to the pathology of RNASET2-deficient leukoencephalopathy.

The contribution of microglia in neurological disorders is emerging as a leading driver rather than a consequence of pathology. RNAseT2-deficient leukoencephalopathy is a severe childhood white matter disorder affecting patients in their first year of life and mimics a cytomegalovirus brain infection. The early onset and resemblance of the symptoms to an immune response suggest an inflammatory and embryonic origin of the pathology. In this study, we identify deficient microglia as an early marker of pathology. Using the ex utero development and the optical transparency of an rnaset2-deficient zebrafish model, we found that dysfunctional microglia fail to clear apoptotic neurons during brain development. This was associated with increased number of apoptotic cells and behavioural defects lasting into adulthood. This zebrafish model recapitulates all aspect of the human disease to be used as a robust preclinical model. Using microglia-specific depletion and rescue experiments, we identified microglia as potential drivers of the pathology and highlight tissue-specific approaches as future therapeutic avenues.

neuroscience↗