Stanley Fields develops biological assays to analyze the function of proteins, often using yeast as a model for assays that can be applied to proteins from any organism. In one approach, his laboratory characterizes the activity of each of thousands of variants of a single protein to infer fundamental properties and to assess the effects of human genetic variation. Other efforts focus on genome engineering to optimize metabolic pathways in yeast.
Mutagenesis of Yeast Drug Export Proteins To Induce Drug Tolerance
Drug resistance is a critical feature of many human diseases and is an important consideration in the production of small molecules by engineered microbes. For example, cancer cells can become drug resistant by increasing the activity of the cellular detoxification machinery. Hyperactive cellular exporters can complicate efforts to use microbial cells to produce and export specific small molecules. Much of the research on cellular detoxification has been performed in the genetically tractable yeast Saccharomyces cerevisiae, whose multidrug transporters and pleiotropic drug response are conserved from bacteria to higher eukaryotes. Therefore, disabled or hyperactive yeast transporters can reveal insights that have implications for therapeutic-resistant cancers and fungal infections as well as for the industrial production of small molecules.
In this project, the student will create a large library of variants (on the order of 1 million) of a yeast drug transporter and use this library to select for yeast cells that become highly resistant to a drug. High-throughput DNA sequencing will be used to identify the causative mutations in the protein, and drug tolerance and specificity will be assayed. These data should help to elucidate the function of drug transporters and inform the study of industrially—and medically—relevant hyperactive transporters. The student will become familiar with genetic and molecular biology techniques and high-throughput DNA sequencing.