C. glabrata is an important opportunistic pathogen causing mucosal infection in healthy individuals and disseminated infection in immunocompromised, primarily hospitalized, patients, responsible for approximately 20% of all candidemias. The Cormack lab is broadly interested in understanding the virulence of this important pathogen. Over the last several years, the Cormack lab has focused on understanding the EPA family of cell wall localized adhesins important in virulence. The EPA genes are normally transcriptionally-silenced and are derepressed during infection in host environments limiting for vitamin precursors of NAD+. Building on their previous analysis, I want to carry out two studies: 1) The lab has previously shown, primarily using whole cell binding assays, that the Epa proteins are lectins, and has analyzed the glycan specificity for a subset of the Epa proteins. I wish to determine the structural basis of Epa-mediated binding to host glycoconjugates, This will be done by X ray crystallographic analysis of the lectin domain of one or more Epa proteins. In these studies, I will be co-mentored by Dan Leahy, who occupies a lab adjacent to the Cormack lab. Understanding the structural basis for lectin-glycan interaction in the Epa family may allow modeling of specificity for all Epa family members, as well as development of small molecule therapeutics that disrupt Epa-mediated colonization. 2) I want to understand the role of NAD+ limitation in the general regulation of virulence in C. glabrata. The lab previously showed that NAD+ limitation increases EPA gene expression, and has unpublished data that NAD+ limitation results in a general increase in virulence of C. glabrata. The transcriptional pathways controlling this response to NAD+ limitation are not known. I propose to identify transcription factor(s) important for regulation of virulence in response to NAD+ limitation and analyze the role of downstream targets in regulating infection. In particular, I am interested in understanding why genes that are regulated by the Bast and Bas2 transcription factors in S. cerevisiae, are upregulated in response to NAD+ limitation, and will use genetic and molecular biological techniques, including microarray analysis and Chromatin Immunoprecipitation approaches to delineate the transcriptional response to NAD+ limitation. These two studies, both related to the core studies of the lab on function and regulation of the Epa adhesion family, will yield important information in understanding the virulence of this important pathogen and in addition will help develop very diverse skill sets in molecular biology, biochemistry, and biophysics that will be important for my future career.
