Molecular characterisation of the mechanisms of compatible solute accumulation in Listeria monocytogenes

The ability of the Gram-positive foodborne pathogen Listeria monocytogenes to survive and grow in environments of elevated osmolarity can be attributed, at least in part, to the accumulation of a restricted range of low molecular mass solutes compatible with cellular function. Accumulated to high internal concentrations in hyper-saline environments, compatible solutes, either transported into the cell or synthesized de novo, play a dual role: helping to stabilize protein structure and function while also counterbalancing external osmotic strength, thus preventing water loss from the cell and plasmolysis. While previous physiological investigations identified glycine betaine, carnitine, and proline as the principal compatible solutes in the listerial osmostress response, genetic alanysis of the uptake/synthesis systems governing the accumulation of these compounds has, until now, remained largely unexplored. Representing the first genetic analysis of compatible solute accumulation in L. monocytogenes, this thesis describes the molecular characterization of BetL; a highly specific secondary glycine betaine transport system, OpuC; a multicomponent carnitine/glycine betaine transporter, and finally proBA; a two-gene operon encoding the first two enzymes of the listerial proline piosynthesis pathway. In addition to their role in osmotolerance, the potential of each system in contributing to listerial pathogenesis was investigated. While mutations in each gene cluster exhibited dramatic reductions in listerial osmotolerance, OpuC- mutants were additionally shown to exhibit reduced virulence when admisistered via the oral route. This represents the first direct link between the salt stress response and virulence in L. monocytogenes.

The architecture of influence: Paul Rudolph's Art and Architecture Building at Yale

Focussing on Paul Rudolph’s Art & Architecture Building at Yale, this thesis demonstrates how the building synthesises the architect’s attitude to architectural education, urbanism and materiality. It tracks the evolution of the building from its origins – which bear a relationship to Rudolph’s pedagogical ideas – to later moments when its occupants and others reacted to it in a series of ways that could never have been foreseen. The A&A became the epicentre of the university’s counter culture movement before it was ravaged by a fire of undetermined origins. Arguably, it represents the last of its kind in American architecture, a turning point at the threshold of postmodernism. Using an archive that was only made available to researchers in 2009, this is the first study to draw extensively on the research files of the late architectural writer and educator, C. Ray Smith. Smith’s 1981 manuscript about the A&A entitled “The Biography of a Building,” was never published. The associated research files and transcripts of discussions with some thirty interviewees, including Rudolph, provide a previously unavailable wealth of information. Following Smith’s methodology, meetings were recorded with those involved in the A&A including, where possible, some of Smith’s original interviewees. When placed within other significant contexts – the physicality of the building itself as well as the literature which surrounds it – these previously untold accounts provide new perspectives and details, which deepen the understanding of the building and its place within architectural discourse. Issues revealed include the importance of the influence of Louis Kahn’s Yale Art Gallery and Yale’s Collegiate Gothic Campus on the building’s design. Following a tumultuous first fifty years, the A&A remains an integral part of the architectural education of Yale students and, furthermore, constitutes an important didactic tool for all students of architecture.