Current Research and Scholarly Interests
Endocytic Pathogens as Tools and Targets
Endocytic pathogens such as protein aggregates, viruses, protein toxins, and bacteria have evolved remarkable ways to enter the cell, disrupt homeostasis, and cause cell death. We use these agents both as probes to understand normal cellular trafficking and signaling events, and to find key targets for therapy.
As an example of this work, we have used high-coverage shRNA libraries and genetic interaction maps to explore the biology of the retrograde-trafficking toxin ricin. We have identified new proteins involved in retrograde trafficking, new components and functions for the vesicle tethering TRAPP complex, and a new role for the small ribosomal subunit RPS25, all of which are under current study.
Stress Signaling to the Cell Death Machinery
Cells have elaborate mechanisms of sensing diverse stresses (oxidative damage, nutrient deprivation, DNA breaks, etc), and must either repair damage or induce cell death. Misregulation of these pathways results in diseases such as cancer and Alzheimer’s. We would like to understand how these signals connect to the death pathway in health and disease in order to improve therapies.
Technology Development and Genetic Interaction Maps
Much of the work we do utilizes genetic screens enabled by novel high-coverage shRNA libraries (~25 shRNAs/gene) we have developed. The high coverage greatly reduces false positive and false negative results that have plagued traditional RNAi strategies. We use a pooled format that can be rapidly screened in large bioreactors, and analyze screens by deep sequencing to quantify changes in shRNA abundance.
Importantly, our library design allows us to knock down pairs of genes, and has facilitated the first systematic genetic interaction maps in mammalian cells. Using these maps, we can understand coordinated gene functions, predict new functions for uncharacterized genes, and identify drug targets. They also allow us to quickly identify synergistic interactions under stress conditions that we hope to exploit for combination therapies. Together with a broad network of collaborators, we are continuing to use this platform to develop new technologies for disrupting and interrogating gene function on a genomic scale.