Valerie A. Ray – Microbiology and Immunology
Mentor – Dr. Karen L. Visick
Loyola University Medical Center, Maywood, IL
Explanation of project:
My research in the Visick lab at Loyola University Medical Center focuses on understanding bacteria-host interactions. Specifically, we are interested in how bacteria transition from living as single cells to living as part of a community, or biofilm, and how biofilm formation impacts host colonization. Since many disease-causing bacteria form biofilms, it is important to understand how biofilm formation is regulated. To do this, I study biofilm formation using a well-characterized model system, in which the bacterium Vibrio fischeri enters into a symbiotic relationship with the Hawaiian bobtail squid Euprymna scolopes. During the initial stages of colonization, biofilm formation by V. fischeri is a critical process. Furthermore, we know that biofilm formation depends upon a set of genes, syp, predicted to be involved in the regulation, production, and transport of an important biofilm component. Since biofilm formation is a tightly regulated process, and is typically limited to the appropriate conditions, we are particularly interested in how the syp genes are regulated under different environmental conditions. Currently, we have evidence to suggest that loss of molecules predicted to be involved in the biosynthesis of the amino acid cysteine leads to a loss of biofilm formation. One readily testable hypothesis is that cysteine is required for normal expression of the syp genes. Thus, to begin to understand how cysteine regulates biofilm formation, we will start by examining syp gene expression under a variety of different conditions, and if we observe an effect, we will probe the regulatory level at which the effect occurs. Overall, understanding how cysteine regulates biofilm formation will provide insights into the environmental control of biofilm formation by V. fischeri during colonization of the squid host.
Description of Summer Undergraduate Work:
The project goal is to understand how cysteine regulates biofilm formation. Thus, the student will begin by examining the impact of cysteine of gene expression (transcription), focusing on the syp genes, which are known to play a critical role in biofilm formation. To do this, the student will make use of genetic “reporter” tools that permit an assessment of transcription using a colorimetric assay called a -galactosidase assay. This assay allows us to examine the relative level of transcription from a syp reporter strain in various V. fischeri mutants (e.g., cysteine biosynthesis mutants) and under various conditions, such as in the presence or absence of cysteine. The results of these experiments will dictate future work. With my help, the student will develop and test specific hypotheses to further understand the impact of cysteine on biofilm formation. The research will take the following form: the summer student will spend the first couple of weeks learning the literature and background for the project as well as learning standard laboratory practices and, in particular, how to perform the -galactosidase assay. Once reproducible data have been obtained, additional permutations of the assay may be completed, such as using additional mutant strains or different amounts of cysteine. With the results in hand, and with my help, the student will develop specific testable hypothesis and likely, generate additional particular strains of interest. Possible experiments include assessing the production of proteins by an immunologic approach called the Western blot, the generation of other amino acid biosynthesis mutants, and/or a genetic approach to search for additional members of a genetic pathway for cysteine-mediated control of biofilm formation. The student should expect to be in lab ~7-8 hours a day Monday-Friday to generate useful data; the more you put into lab, the more you get out of it. Overall, the student will gain a general understanding of standard laboratory practices and importantly, learn how a scientist follows the results of their data to develop and test specific hypotheses, as well as how to think critically.