http://theses.ncl.ac.uk/jspui/handle/10443/5256 2026-05-19T10:55:46Z 2026-05-19T10:55:46Z Lim, Jing Xuan http://theses.ncl.ac.uk/jspui/handle/10443/6785 2026-05-15T11:38:12Z 2025-01-01T00:00:00Z

Title: Determining Previously Undefined Immune Regulatory Networks in Regulatory T Cells and Innate Lymphoid Cells within the Tumour Microenvironment Authors: Lim, Jing Xuan Abstract: Regulatory T cells (Tregs) are essential for immune suppression, employing multiple mechanisms, including co-inhibitory receptor expression. However, the role of the programmed cell death-1 receptor (PD-1) in Treg function remains controversial. Here, we identify PD-1 as a key checkpoint in Tregs, orchestrating a unique co-inhibitory receptor network that shapes their function in tumour immunity. We demonstrate that PD-1 regulates the expression and activity of CD30, a central driver of Treg suppressive function within the tumour microenvironment (TME). Mechanistically, PD-1 deficiency amplifies STAT5 signalling in Tregs, leading to upregulated CD30 expression and thus suppressive capacity. Consistently, in stage IV metastatic melanoma patients, anti-PD-1-resistant individuals exhibit increased CD30 expression on Tregs compared to responders. Thus, we uncover a novel role for PD-1 in restraining CD30 expression, thereby modulating Treg-mediated immunosuppression. These insights provide a rationale for targeting PD-1 and CD30 in combination therapies to improve cancer treatment outcomes. Innate lymphoid cells (ILCs) also play a crucial role in tissue immunity and are influenced by co-receptor signalling. Here, we define a subset of tumour-infiltrating ILCs characterised as Tbet⁺NK1.1⁻, which express PD-1 within the TME. We demonstrate that PD-1 profoundly regulates the function of Tbet⁺NK1.1⁻ ILCs in murine B16 melanoma. PD-1-deficient Tbet⁺NK1.1⁻ ILCs exhibit significantly increased production of IFN-γ, granzyme B, and granzyme K, correlating with diminished tumour growth. These findings highlight a novel regulatory axis through which PD 1 modulates anti-tumour responses in ILCs, suggesting potential therapeutic avenues to enhance immune-mediated tumour clearance. Description: Ph. D. Thesis.

2025-01-01T00:00:00Z Dimitriou, Anastasia http://theses.ncl.ac.uk/jspui/handle/10443/6769 2026-05-13T07:57:36Z 2025-01-01T00:00:00Z

Title: The impact of alpha (α)-synuclein pathology on prefrontal cortex networks in a mouse model of Dementia with Lewy Bodies Authors: Dimitriou, Anastasia Abstract: Dementia with Lewy Bodies (DLB) is caused by aggregated insoluble alpha(α) synuclein (α-syn) in neurons and patients exhibit cognitive impairment involving the anterior cingulate cortex (ACC). Cortical network hyperexcitability has also been observed in DLB patients. Parvalbumin interneurons (PVIs) are critical for controlling excitability and normal cognitive function and are often surrounded by a specialised extracellular matrix, the perineuronal nets (PNNs). Loss of PVIs and PNNs occurs in Alzheimer’s Disease but their role in DLB remains unclear. Neuroinflammatory changes may play an early role in DLB. In this thesis, I aimed to investigate hyperexcitability in the early stages of α-syn disease pathology in the ACC using hA30P transgenic mice expressing human α-syn. Changes in PVIs, PNNs, and glial cells in different disease stages were examined. In the A30P mice, I found hyperexcitability in ACC in vitro including frequent seizure-like events associated with PV activity. Additionally, immunofluorescence was conducted to examine the impact of hα-syn pathology on PVIs, PNNs, microglia, and reactive astrocytes in the ACC in young (2-4 months) and aged (10-12 months) A30P and control mice. A trend towards a decrease in the PVI number was revealed in young and aged A30P mice. Additionally, a significantly greater proportion of PNNs surrounding non-PV neurons was observed in young A30P animals. PV somas and PNNs contained hα-syn in young A30P mice and this expression increased with age. Neuroinflammation also increased in all aged animals showing a significant increase in the %area of GFAP+ astrocytes and reactive microglia. This thesis provided evidence of hyperexcitability potentially related to changes in PV neurons in the ACC and a shift in PNN localisation to surround presumed pyramidal neurons in young A30P mice which might be protective against hα-syn pathology. Additionally, the data suggested increased neuroinflammation correlated with age in the ACC in hA30P and control mice. Description: PhD Thesis

2025-01-01T00:00:00Z Norris, Jonathan http://theses.ncl.ac.uk/jspui/handle/10443/6766 2026-05-08T14:35:49Z 2025-01-01T00:00:00Z

Title: Understanding the role of transcription in organisation of the bacterial chromosome Authors: Norris, Jonathan Abstract: Bacterial chromosomes are organised by various proteins, types of supercoiling and other cellular processes. One such process, transcription, massively impacts the chromosomal structure from the local level up to overall organisation of the nucleoid. Uniquely to bacterial transcription, the process can be physically coupled with translation since they occur in the same cellular compartment. The processes and associated proteins can, thus, happen simultaneously and physically interact. This coupling can have further impact of the overall structure of the nucleoid. While transcription-translation coupling is well documented in E. coli, some work suggests that it does not happen in other bacteria, including the Gram-positive model bacterium Bacillus subtilis. To study transcription-dependent chromosome organisation at a single cell level in B. subtilis, I fluorescently labelled DNA in the vicinity of the promoter of an inducible gene coding for a transmembrane protein and followed the localisation of the gene locus using fluorescence microscopy. We found that, upon induction, the gene migrates from a central position in the cell towards the membrane, and back towards the nucleoid when induction is removed. This movement was further confirmed by monitoring the fluorescently labelled locus in vertically immobilised cells (Vertical Cell Imaging by Nanostructured Immobilisation), which provides a better optical viewing angle for the observed process. Inhibiting either transcription and translation, via antibiotics and mutations in respective initiation regions, abolished the movement of fluorescently labelled locus towards the cell periphery. This loss of gene movement indicates the involvement of both transcription and translation in the process. Our results are fully consistent with transertion; a postulated process in which transmembrane proteins are inserted in the membrane co-translationally and cotranscriptionally thereby pulling the gene locus from the nucleoid core to the periphery of the cell and provide the first direct experimental evidence for transertion in Gram-positive bacteria. Furthermore, these findings demonstrate that translation and transcription can indeed be coupled in B. subtilis, alongside translocation, at least for genes encoding for membrane proteins. Description: PhD Thesis

2025-01-01T00:00:00Z Kerridge, Scott Thomas http://theses.ncl.ac.uk/jspui/handle/10443/6752 2026-05-01T09:32:16Z 2025-01-01T00:00:00Z

Title: Cyclin B1 regulation and APC/C processivity in mouse oocyte meiosis I Authors: Kerridge, Scott Thomas Abstract: Chromosome alignment is orchestrated by the activity of CDK1 bound to its coactivator cyclin B1. Equally important, chromosome segregation is orchestrated by termination of CDK1 activity, driven by cyclin B1 destruction. Both events must be precisely timed by APC/C (Anaphase Promoting Complex/Cyclosome) activity, an E3 ligase that targets cell cycle proteins for destruction via the ubiquitin-proteasome pathway. In mitosis, this system is rapid, robust, and well-studied. Critically, through oocyte chromosome alignment, CDK1 and APC/C activities must be regulated differently, over substantially longer time frames. Specifically, the APC/C must first be dampened to prevent cyclin B1 destruction and CDK1 activity loss as chromosomes begin aligning. This is followed by a period of non-CDK1-bound cyclin B1 destruction during final alignment stages, followed by CDK1-bound-cyclin B1 destruction once fully aligned. This ordering is vital to minimise errors that otherwise cause the cell to arrest (infertility) or produce an embryo that is incompatible with life (miscarriage). However, little is published regarding the molecular mechanism of cyclin B1 targeting in oocytes, or indeed the wider landscape of any APC/C substrate ordering. To address this, I have expressed fluorescent versions of cyclin B1 and APC/C substrates in live mouse oocytes, utilising time-lapse fluorescence microscopy to map their levels and movements through meiosis I. Additionally, I mutated short linear motifs within these substrates, as well as APC/C subunit knockdowns to obtain mechanistic insight into substrate targeting. I reveal that cyclin B1 harbours a motif which boosts its affinity for a specific form of the APC/C during chromosome alignment. This form of the APC/C does not exist globally across the cell, but likely in high priority areas across this spindle zone. This and other insights, describe novel aspects of cell cycle regulation that are critical to produce healthy eggs Description: Ph. D. Thesis.

2025-01-01T00:00:00Z