I am a Simons Postdoctoral Fellow in the laboratory of Feng Zhang at the Broad Institute and the Departments of Brain and Cognitive Sciences and Bioengineering at MIT. I received my PhD at MIT in Sebastian Seung's lab and I completed my undergraduate education at Stanford University.
My scientific interests include genome engineering, functional genomic screening using programmable nucleases, autism genetics and synaptic pathophysiology, and the regulation of RNA editing in neurons. Please click here for my online CV.
functional genomics with CRISPR-Cas9
Many targeted loss-of-function genetic screens, such as RNA interference, work at the transcript level. Recently, we developed a new tool for performing functional genetic screens on the genome itself. Taking advantage of the easy programmability of the prokaryotic CRISPR-Cas9 nuclease, we created genome-wide pooled libraries to interrogate loss-of-function mutations in a massively parallel fashion. When compared to shRNA knock-down, individual CRISPR reagents are much more consistent in their ability to target and knock-out genes, resulting in a greater number of validated hits from the screen. With GeCKO, we have conducted genome-scale positive and negative selection screens using human cancer cells and human pluripotent stem cells. Through our work, we have been able to discover and validate several new loss-of-function mutations that confer resistance to vemurafenib, a FDA-approved BRAF-inhibitor commonly prescribed for malignant melanoma.
genome engineering toolbox with TALEs
We are now able to sequence entire genomes easily and cheaply. But what about our ability to write DNA? How can we make targeted changes to genomes? What kinds of new experiments and therapeutics will these technologies enable? In this work, we developed a protocol to rapidly clone and test TALE nucleases and transcriptional activators. Currently, I use modular programmable DNA binding proteins to test the causal role of putative genetic disease variants in human cells. My goal is to expand the available tools for genome engineering while applying these technologies to understand and treat neuropsychiatric diseases, such as autism and schizophrenia.
activity-dependent RNA editing in neurons
Can neural activity modify RNA after transcription? In RNA editing, adenosines are converted to inosine (A-to-I editing) by a deaminase after transcription. At the ribosome, inosine is read as guanosine and thus can alter the amino acid translation of the edited transcript. There is a growing number of newly identified A-to-I RNA editing sites in the genome but editing levels of the best characterized sites seem to be fixed after early development. We screened all known coding, non-synonymous editing sites in the rat genome to understand how neural activity can influence RNA editing. We found that activity can dynamically alter editing in several functionally relevant transcripts and that this editing depends crucially on Ca2+ entry into the cell.
long-term time-lapse imaging of growing cortical axons
What can the movements of axon growth cones tell us about how nervous systems are wired up? Applying stochastic modeling techniques to time-series of growth cone positions, we find that axons have distinct stereotyped behaviors during early outgrowth. Our goal is to use these models of "cellular behavior" to discover the underlying biophysical mechanisms in the axon growth cone. On the experimental side, we have developed an integrated software-hardware platform and primary neuron prep for long-term (weeks) unattended time-lapse microscopy of genetically labeled axons and dendrites in culture.
microfluidics for diffusable growth factor gradients
How can we create target-derived, diffusible (vs. substrate-bound, see inkjet project below) chemotropic signals to neurons in vitro? We developed a simple microfluidic system to use for time-lapse imaging of axon outgrowth from rat dorsal root ganglia neurons. The microfluidic chamber was designed to produce a linear gradient of a neurotrophic factor for guiding growing axons.
inkjet substrate micropatterning for neurons
For precise micropatterning of substrate-bound neural adhesion and guidance molecules, we developed a custom inkjet printer and flexible surface chemistry. We have created viable, healthy cultures of primary hippocampal neurons and glia that adhere to specific patterns for weeks in vitro. An example of a patterned culture is shown in the image to the left: Each letter in the MIT logo contains an isolated micronetwork of ~50-200 neurons.