Role of DNA adenine methylase in regulation of bacterial gene expression, virulence, and the elicitation of immune responses

The threat of emerging infectious diseases and bio-warfare agents has underscored the need for development of safe and effective anti-microbial therapies. One viable approach to address this issue is through protective vaccination, which second to water sanitation, is the principal method for reduction of morbidity and mortality worldwide. This dissertation describes the role of the DNA adenine methylase (dam) as a global regulator of bacterial virulence and the utility of Dam-based technology for the development of live-attenuated vaccines that elicit potent states of immunity against a variety of infectious agents. Dam is a highly conserved enzyme found in many pathogens, and is involved in a variety of cellular processes including the regulation of bacterial virulence and elicitation of protective immune responses. Work presented in this dissertation has further evaluated the role of Dam in regulation of several virulence factors in Salmonella typhimurium and Yersinia pseudotuberculosis. Studies in Yersinia revealed that Dam overproduction alters the stringent regulation of many virulence factors, including a principal Yersinia immunogen, LcrV, and an actin cytotoxin, YopE. Dysregulation of these virulence factors may be the mechanism by which dam mutants elicit protective immunity against Yersinia infections in vaccinated hosts. In Salmonella, Dam regulates the expression of several virulence genes as well. Moreover, expression of these genes was differentially affected by altered Dam levels in closely-related pathogenic and non-pathogenic strains of Salmonella. These data suggest that differential gene regulation, rather than gross genomic differences contributes to the virulence disparities observed between these pathogenic and non-pathogenic isolates. Finally, by expressing reovirus antigens in Salmonella, the possibility of using Dam-based vaccines as carriers of foreign antigens was investigated. Effective live-attenuated carrier vaccines could be used to protect hosts from a variety of diseases for which no therapy is currently available. Investigating the role of Dam in regulation of virulence factors in pathogens such as Salmonella and Yersinia will not only help decipher its role in regulation of virulence in other pathogens, but will also help design new vaccines that provide potent immunity against a wide range of infectious diseases that threaten the safety of public health worldwide.