Neville picture I am currently 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.

projects

functional genomics with CRISPR-Cas9

genome-scale crispr-cas9 knock out screening logo Recently, we developed a Genome-scale CRISPR Knock-Out (GeCKO) library. Taking advantage of the easy programmability of the CRISPR-Cas9 nuclease, GeCKO enables pooled functional genomics screens 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 (not knock-down) 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.

More information | GeCKO online resources and MIT News article

genome engineering toolbox with TALEs

transcription activator-like effector nuclease and transcription factor diagrams 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.

More information | TAL Effectors online resources & engineered stem cells for autism (Simons, NPR)

activity-dependent RNA editing in neurons

chromatogram example 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 (base pairs with cytosine) 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.

More information | Publication and journal cover in Genetics

long-term time-lapse imaging of growing cortical axons

time-lapse cultured neurons 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.

More information | 5 day time-lapse of neural network formation (Olympus BioScapes Gallery)

microfluidics for diffusable growth factor gradients

microfluidic mixer for diffusible gradient 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

ink-jet patterning of text MIT in 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.

More information | Neuro-Inkjet web tutorial and MIT News article