268 research outputs found

    Exploring the biological basis of residual feed intake in beef cattle using multi-Omics analysis.

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    Efficient feed utilization is critical for both economic sustainability and environmental responsibility in modern livestock production. While there has been extensive research, the multifaceted nature of feed efficiency remains complex, with many biological factors still unexplored. This dissertation examines the physiological foundations of feed efficiency by elucidating some of the complex biological mechanisms associated with residual feed intake (RFI) phenotype in beef cattle, using a range of Omics approaches. We hypothesized that metabolites related to amino acids, carbohydrates, and fatty acids could act as potential biomarkers for RFI. Through a chemical group-based metabolomics method, we identified enriched pathways in feed-efficient steers, notably in arginine biosynthesis and histidine metabolism. This led to the identification of five potential metabolite biomarkers mainly linked to amino acid metabolism, emphasizing a relationship between blood amino acid profiles and RFI. This led us to investigate the expression of genes and associated pathways related to nutrient and energy metabolism, especially in liver tissue, where hepatic metabolism is driven by transcriptional regulation. Low-RFI steers showed upregulation of genes involved in fatty acid transport, β-oxidation, and mitochondrial ATP production. In contrast, a crucial gene in amino acid metabolism responsible for aminoadipate aminotransferase activity exhibited a significant decrease in expression in low-RFI steers. These results indicate that alteration in expression of hepatic genes regulating lipid and amino acid metabolism, and mitochondrial ATP generation is associated with RFI phenotype. We also investigated potential differences in the rumen microbiome and immune gene expression of beef steers with low or high RFI. We observed increased mRNA expression of immune-related genes in both blood and liver tissues of low-RFI beef steers, especially those linked to pathogen detection and phagocytosis. Low-RFI steers also displayed variation in the relative abundance of microbial taxa compared to high-RFI. Lastly, detailed statistical analysis indicated that plasma amino acids such as tyrosine, glycine, and dimethyl sulfone may be promising economic prospects as cost-efficient predictors of RFI in beef cattle. In conclusion, this dissertation provides invaluable insights into some of the intricate biological processes associated with RFI in crossbred beef cattle, enhancing our grasp of the involved biological mechanisms and laying the groundwork for refining feed utilization in the beef cattle sector of livestock production

    Establishment of recombinant antibody technologies allowing for the generation of SNAP-tag fusion proteins

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    Triple negative breast cancer (TNBC's) is a highly aggressive and invasive subtype of breast cancer, typically characterised by the lack of estrogen receptor (ER), progesterone receptor (PR) and Human epidermal growth factor receptor 2 (HER2) with an inexplicable partiality towards African women. The acute heterogenicity and complexity of TNBC tumours, together with a lack of well-defined molecular targets, complicates prognosis of the diseases resulting in patient reliance on traditional therapies, like chemotherapy, radiotherapy, and surgery, which are associated with elevated incidence of adverse effects and relapse. A major contributor to the heterogenicity of TNBCs is the tumour microenvironment which is composed of tumour infiltrating lymphocytes (TILs), tumour cells, healthy cells, and tumour vasculature. TILs have commonly been used as a prognostic marker and show robust predictive value for TNBC. In-depth analysis of the TIL composition within TNBC tumours may provide greatly beneficial information for the development of newer tumour microenvironment changing therapies and could assist doctors in understanding what therapies a particular patient maybe susceptible to. Thus, the diagnosis and therapy of this disease may greatly benefit from improved molecular profiling and patient stratification. Precision medicine seeks to provide such a solution, by dividing patients into subpopulations based on disease-specific profiles. The identification of new molecular targets would provide the basis for development of novel therapies. To this end, one of the major aims of this thesis was to develop a phage display based screening technique which could be utilised to isolate novel TNBC specific cancer antibodies. Once selected these antibodies could be used to generate TNBC specific therapies. Specific monoclonal antibodies (mAbs) and derivatives thereof, have already been established as a revolutionary tool for drug delivery to cancerous cells. Such antibodies have been conjugated to cytotoxic drugs to form antibody-drug conjugates, which may exhibit multiple advantages over their unconjugated counterparts, but their general use in clinical application has been restricted due to developmental deliberations. Historical conjugation strategies used for the generation of ADCs commonly resulted in heterogeneous mixtures of ADC species, with varying drug-to-antibody ratios resulting in unpredictable pharmacologic characteristics and safety profiles. In more recent time, self-labelling tags such as Snaptag have provided a means of developing homogenised recombinant immunotherapeutics. Snaptag is a modified version of a human DNA repair enzyme, O6 - alkylguanine-DNA-alkyltransferase (AGT) which naturally removes alkyl residues from damaged DNA. The enzyme reacts specifically with benzylguanine (BG) derivatives via irreversible transfer of alkyl groups to cysteine residues forming stable end products. In this thesis, Snaptag technology, together with other antibody discovery and manipulation tools was used to develop a methodology allowing for the generation of disease specific fusion proteins. Specifically, these fusion proteins consist of single-chain antibody fragments genetically fused to snaptag, allowing for the generation of recombinant ADCs that could be used as a drug delivery system carrying any BG-modified drug to a disease specific targets. In addition, snaptag interacts with BG in a 1:1 stoichiometry giving rise to homogenised combination products which when fused to a scFv provides a fail-safe target-specific therapeutic option. In addition to antibody conjugates, one of the most promising of all mAb based therapies currently used, are checkpoint inhibitors. In a balanced immune response, immune activation is counteracted with immunoregulatory pathways such as checkpoint inhibition. These negative regulatory pathways are necessary for maintaining tolerance and preventing hyperactivation, and are governed by cell surface, inhibitory receptors known as ‘'checkpoint inhibitors''. Blocking of checkpoint pathways during chronic infections and cancer has been shown to improve T-cell functions leading to reduced viral load and tumour burden. These findings have been translated into clinical application where checkpoint inhibitors, which are monoclonal antibodies targeting CTLA-4, PD1, PD-L1 or other inhibitory ligands, have been used to block these inhibitory interactions. The main intention of this research was to develop a methodology which could be used to generate snaptag based recombinant fusion proteins with potential diagnostic and therapeutic applications. Several snaptag based fusion proteins were developed using the recommended methodology these included fusion proteins targeting breast cancer specific antigen BCK1, checkpoint inhibitors PDL1, B7.1/CD80 (interacts with CTLA-4),and TIL characterising markers CD3, CD4, CD8, CD19 and CD20. In addition, to demonstrate the versatility and robustness of this methodology we sought to develop a snaptag based fusion protein not targeting breast cancer related antigens. Zika virus, an emerging infectious disease, currently lacking specific therapies was chosen for this purpose. An scFv derived from antibodies targeting the the Zika-DIII envelop protein, which is essential to the viral infection cycle was used in the snap fusion protein. The resulting ZIKA-DII-snap fusion protein demonstrated specific binding to zika virus membrane fractions. This research demonstrates the feasibility of using snaptag technology as a state-of-the-art conjugation strategy capable of bypassing the challenges previously associated with using antibodies as an effective delivery system for therapeutic drugs. By combining the applicability of snaptag technology with other antibody isolation and manipulation tools we were able to generate several functional snaptag based recombinant fusion proteins. Establishment of this methodology represents an important first step in generating medically necessary, pharmaceutically acceptable immunoconjugates that is instrumental in shifting general therapy towards a more personalized precision medicine approach

    Endocrine and Neuroendocrine Cancers

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    Endocrine and neuroendocrine tumors originate from endocrine cells but are heterogeneous in terms of clinical presentation, disease outcome, and treatments available. This Special Issue addresses specific aspects of pre-clinical, clinical, and translational research and clinical management of these diseases with the aim of providing novel insights, addressing current unmet needs, and discussing future treatment perspectives

    Polarity development by asymmetric protein-cluster distributions in response to cortical flows in C. elegans zygotes

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    Asymmetric cell-division in the one-cell embryo is a key step in embryonic development. The initially homogeneous zygote establishes an anterior-posterior axis within the cell, allowing for the unequal distribution of cell fate determinants and subsequent cell differentiation. Therefore, polarity development is a fundamental procedure that defines the information template from which all future cell processes derive their cues. Many molecular players in polarity formation in C. elegans have been identified, but the design principles that underpin their interactions and how this contributes to successful polarisation remain unclear. This thesis focuses on the role of the clustering species PAR-3 and how its integration in the governing biochemical network promotes robust polarisation. To correctly proceed to the two-cell stage, the foundational step of polarity formation must be responsive to the polarising cue and maintain established domains ready for downstream cell-cycle processes. We characterise global flows along the polarity axis and investigate coupling between the cortical flows and PAR-3 cluster sizes. We find there is no dynamic advantage for larger clusters and instead conclude all clusters flow with the same efficiency. Alternatively, we investigate whether enhancement of clusters is a response to mechanical forces within the cortex during the period of flow but find little direct evidence of this relationship. Rodriguez et al proposed kinase cycling between inactive (advective) and inactive (diffuse) state. We model two distinct reaction pathways through reaction-advection-diffusion simulation and assess their viability by implementation of Approximate Bayesian Computation. We find that direct binding through a flow-sensing, inactive state, followed by switching to a diffuse active state yields a network that is unviable and sensitive to perturbation. Sensitivity is alleviated when the advective species serves only to enhance independent binding of the active species. Therefore, we propose kinase cycling through this network motif as a mechanism towards enhanced robust polarisation.Open Acces

    Virus host shifts in Drosophila: The influences of virus genotype and coinfection on susceptibility within and across host species

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    Virus host shifts are a major source of outbreaks and emerging infectious diseases, and continue to cause considerable damage to public health, society, and the global economy. Predicting and preventing future virus host shifts has become a primary goal of infectious disease research, and multiple tools and approaches are being developed to work towards this goal. In this thesis, I examine three key aspects of infection that have implications for our wider understanding of virus host shifts and their predictability in natural systems: whether the outcome of infections across species is correlated between related viruses, whether the presence of a coinfecting virus can alter the outcomes of cross-species transmission, and the influence of host genetics and immunity on the outcomes of coinfection. These experiments make use of a large and evolutionarily diverse panel of Drosphilidae host species, and infections with two insect Cripaviruses: Drosophila C virus (DCV) and Cricket Paralysis virus (CrPV), with the outcomes of infection quantified throughout as viral loads via qRT-PCR. In Chapter Two, phylogenetic generalised linear mixed models are applied to data on the outcome of single infections with three isolates of DCV (DCV-C, DCV-EB, DCV-M) and one isolate of CrPV, to look for correlations in viral load across host species. Strong positive corrections were found between DCV isolates and weaker positive correlations between DCV and CrPV, with evidence of host species by virus interactions on the outcome of infection. Of the four viruses tested, the most closely related isolates tended to be the most strongly correlated, with correlation strength deteriorating with the evolutionary distance between isolates, although we lacked the diversity or sample size of viruses to properly determine any effect of evolutionary distance on correlation strength. Together, this suggests that hosts susceptible to one virus are also susceptible to closely related viruses, and that knowledge of one virus may be extrapolated to closely related viruses, at least within the range of evolutionary divergence tested here. In the remainder of this thesis, I examine the outcome of coinfection with DCV-C and CrPV across host species (Chapter Three) and across genotypes and immune mutants of Drosophila melanogaster (Chapter Four). These chapters aim to assess the potential for coinfection to alter the outcomes of cross-species transmission – and so interfere with predictions of virus host shifts – and the potential influence of host genetics and immunity on the outcome of coinfection. Chapter Three finds little evidence of systematic changes in the outcome of single and coinfection for both viruses across species, suggesting that coinfection may not be a required consideration in predictive models of every host-virus system. Effects of coinfection were found in a subset of species but were not recapitulated in a follow-up experiment looking at tissue tropism during coinfection on a subset of host species. Together, this suggests that any effects of coinfection across species with DCV and CrPV are due to stochastic effects within individual hosts. Chapter Four finds small but credible effects of coinfection across genotypes of D. melanogaster, but these effects showed little host genetic basis or effect on the genetic basis of susceptibility to each virus separately. Mutations in several immune genes caused virus-specific changes in viral load between single and coinfection, suggesting that coinfection interactions between viruses can be moderated by the host immune response. This thesis has aimed to explore several fundamental features of cross-species transmission that are relevant to our understanding – and ability to predict – virus host shifts. Both the finding that correlations exist between viruses and the approach used to characterise coinfection across and within host species would now benefit from an increased diversity of experimental pathogens, to better investigate the influence of virus evolutionary relationships on the outcomes of virus host shifts and present a broader understanding of the potential impact of coinfection on the outcomes of cross-species transmission.Natural Environment Research Council (NERC

    Hallmarks of Parkinson’s Disease

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    This book gathers a collection of review articles aiming to provide an update of the current knowledge on PD pathogenesis. It notably includes an overview of several key cellular dysfunctions underlying the etiology of Parkinson’s disease, including ER stress, mitophagy and alpha-synuclein-linked pathology

    RNA, the Epicenter of Genetic Information

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    The origin story and emergence of molecular biology is muddled. The early triumphs in bacterial genetics and the complexity of animal and plant genomes complicate an intricate history. This book documents the many advances, as well as the prejudices and founder fallacies. It highlights the premature relegation of RNA to simply an intermediate between gene and protein, the underestimation of the amount of information required to program the development of multicellular organisms, and the dawning realization that RNA is the cornerstone of cell biology, development, brain function and probably evolution itself. Key personalities, their hubris as well as prescient predictions are richly illustrated with quotes, archival material, photographs, diagrams and references to bring the people, ideas and discoveries to life, from the conceptual cradles of molecular biology to the current revolution in the understanding of genetic information. Key Features Documents the confused early history of DNA, RNA and proteins - a transformative history of molecular biology like no other. Integrates the influences of biochemistry and genetics on the landscape of molecular biology. Chronicles the important discoveries, preconceptions and misconceptions that retarded or misdirected progress. Highlights major pioneers and contributors to molecular biology, with a focus on RNA and noncoding DNA. Summarizes the mounting evidence for the central roles of non-protein-coding RNA in cell and developmental biology. Provides a thought-provoking retrospective and forward-looking perspective for advanced students and professional researchers
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