Thesis (M.S.)--Rochester Institute of Technology, 1984. / Typescript. Includes bibliographical references (leaves 32-35).

Thesis (Ph. D.)--University of Texas at Austin, 2004. / Supervisor: Christine Schmidt. Vita. Includes bibliographical references.

Thesis (Ph. D.)--University of Texas at Austin, 2005. / Vita. Includes bibliographical references.

Thesis (Ph.D.) - University of Queensland, 2007. / Includes bibliography.

No description available.

Aquaculture is known as a major food-producing industry and Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss) are the major cultured species in Scotland. However, disease outbreaks in aquaculture have been reported and are commonly associated with intensive fish farming, which results in a tremendous cost in the industry. Hence, understanding what immune-related genes and cells are present, their responses, mechanisms and functions in these farmed animals is a first requirement for potent vaccine design, selection of disease-resistant breeds and disease outbreak prevention. The innate immune system is the first line of defence against microbes which use germline-encoded pattern recognition receptors (PRRs) to recognise specific, conserved and constitutive products of invading pathogens, called pathogen-associated molecular patterns (PAMPs) that are important for survival of the microorganism and are thus hard for the microorganism to change. This thesis focuses on the identification and characterisation of a family of PRR called toll-like receptors (TLRs) and a TLR accessory protein UNC93B1 using different approaches. In Chapters 2, 3 and 5, eleven TLR genes and UNC93B1 were identified from Atlantic salmon whole-genome shotgun (WGS) contigs. These genes were cloned and sequenced and their putative domain structure, gene synteny and homology to other genes were determined by bioinformatics analysis. In addition, the constitutive expression profile of these genes was examined in different tissues from healthy salmon using real-time PCR. The potential modulation of these genes was examined in different in vitro and in vivo models which provide information to help understand the role(s) of these genes during inflammation or in the immune responses against pathogens. Several of these TLRs are so-called non-mammalian TLRs (TLR19, TLR20a and TLR20d) and are therefore particularly interesting to study. The sub-cellular localization was also investigated in TLR-GFP expression plasmid transfected Salmon Head Kidney-1 (SHK-1) cells. Lastly, attempts were made to develop a Human Embryonic Kidney (HEK) 293T cell line based platform to study TLR signalling and ligand specificity (Chapter 4).

No description available.

Roundabout (Robo) receptors were first identified in neurons as guidance molecules, however growing evidence suggests that they also play a role in other cells. The aim of this thesis was to characterise the expression and function of a novel endothelial specific member of this family, Robo4. This study revealed that Robo4 is expressed primarily in vessels but also differentially expressed in tumour vessels. Interestingly Robo4 was primarily located within cytoplasmic vesicles coated with clathrin, suggesting that the presence of Robo4 on the cell surface is being tightly regulated. Overexpression of Robo4 induced filopodia and pseudopodia formation and actin re-organisation into stress fibres. It co-localised with actin and tubulin suggesting an important interaction between Robo4 and the cytoskeleton. Robo4's function in endothelial cells was directly investigated using two approaches, overexpression using adenovirus and knockdown using small interfering RNA. Functional cell-based assays revealed that disrupting Robo4's level of expression negatively affects endothelial cell functions that are required during angiogenesis, such as proliferation, migration and tubulogenesis. Overexpression of a truncated version of Robo4, which lacks the C-terminus, provided clues regarding Robo4's function. The intracellular domain is critical for Robo4's localisation and its association with the cytoskeleton. It is also required for pseudopodia formation. Other findings include possible cleavage of Robo4 and Robo4 homodimerisation and heterodimerisation with Robo1. Taken together, the findings presented in this study strongly suggest a role for Robo4 in endothelial cell guidance. Cell guidance during angiogenesis is poorly understood therefore the identification of a new molecule potentially involved in this mechanism will hopefully help elucidate the process.

Abstract not available

In addition to the well-characterized genomic mechanism of steroid action that uses intracellular receptors, steroid hormones also signal through nongenomic processes that use membrane receptors. A membrane receptor for corticosterone (CORT) has been described in brains of the roughskin newt (Taricha granulosa). This receptor is believed to be a G-protein coupled receptor because corticosterone binding is inhibited by guanyl nucleotides and enhanced by Mg�����. The studies described in this thesis use biochemical, pharmacological and molecular techniques to characterize the newt neuronal membrane glucocorticoid receptor (mGR) in pursuit of its molecular identification. The mGR was successfully solubilized from newt neuronal membranes and conditions were defined that maintained corticosterone binding activity for further study. The solubilized receptor was partially purified using standard chromatographic techniques and an immobilized ligand affinity resin (CORT-Sepharose). These chromatographic studies were combined with the use of a novel photoaffinity ligand (azido-CORT) to biochemically characterize the mGR protein, finding that it is an acidic glycoprotein with an apparent molecular weight of 63 kDa and an isoelectric point of approximately 5.0. Pharmacological studies with mGR showed that a subset of kappa opioid ligands displaced corticosterone from the receptor binding site with K[subscript i] values in the nanomolar to low-micromolar range. The interaction of mGR with kappa opioid ligands was specific because no mu-, delta-, or orphanin-specific opioid ligands were effective at displacing corticosterone from the receptor. These data suggest that the newt neuronal mGR may be a kappa-opioid like receptor. Finally, molecular studies were used to clone a novel newt brain protein, neuronal axonal protein 22 (NAP-22), that was identified in a protein differential display strategy designed to identify mGR. Studies with the cloned and expressed NAP-22 protein suggest that it is not the mGR but, instead, may be a mGR-associated protein. These studies provided new information about the biochemical and pharmacological properties of mGR, and may have discovered a protein that is associated with the newt neuronal mGR. / Graduation date: 1999