Stanford University Department of Applied Physics

William Greenleaf

(Acting) Assistant Professor of Genetics (Sept. 2011)

Research areas:
Biophysics, Nano Sci/Eng, Precision Measurement

Description

Biophysics

The positioning and composition of nucleosomes provides an epigenetic layer of information to the genome itself, creating a molecular memory that can help to perpetuate cell-specific gene expression patterns through generations. Indeed it is becoming apparent that the physical positioning and modifications of histone particles encode a molecular "state machine" of the cell. To investigate the chromatin of single-cells at the level of histone positioning and composition, we strive to adapt microfluidic cell sorting and laser-based manipulation methods to probe the chemical and mechanical characteristics of chromatin. This analysis will help lift the veil on cell-to-cell epigenetic variability and the modifications that likely determine cell fate and begin to cast light on the mechanism of heritability of this component of the epigenetic code. The dynamic mechanical processes whereby nucleosomes are assembled, ejected, exchanged, and moved by molecular motors to make genetic information available to the machinery of expression is at the heart of gene regulation. Recent technical advances in the areas of single molecule biophysics create a tremendous opportunity to explore the mechanics of chromatin dynamics. We are also interested in the mechanical characterization of histone-DNA interactions and chemomechanical investigation of the molecular motors required to slide nucleosomes, with the aim of providing a detailed, dynamic physical picture of histone assembly and remodeling, complementing traditional biochemical techniques.

Condensed Matter Physics

The cell faces a spectacular topological challenge in packing meters of chromosomal DNA inside a ~5 micron nucleus. The solution to this challenge is the hierarchical folding of genomic DNA into regulated structures, from DNA-wrapped histone particles called nucleosomes to the large-scale structures that comprise the metaphase chromosome. However, the topological structure of the metaphase chromosome, one of the most recognizable and interesting biological structures, remains obscure. Because chromatin is not crystalline, the topological structure of condensed chromatin cannot be solved with standard crystallographic techniques -- instead, single molecule methods are called for. We hope to help unravel this structure by developing innovative new single molecule manipulation methods based on "trappable" magnetic micro particles and optical tweezers. These magnetic methods in combination with optical trapping methods, will allow for both intracellular and extracellular mechanical manipulation of chromatin, and will help shed light on the enduring mystery of higher-order chromatin structure.

Nanoscience and Quantum Engineering

As a natural extension of our interest in individual cells and molecules, we are motivated to adapt and develop methods that allow manipulation, observation, and quantification of very small amounts of biomaterials through the use of microfluidic devices. We have also developed a methods of generating minuscule resealable reactions chambers in polydimethylsiloxane. Standard microfluidic devices are constructed on a length scale of 10s or 100s of microns and involve volumes on the order of nanoliters or picoliters. We are interested in reaction volumes approximately 10,000 times smaller, allowing generation of tens of millions or more isolated reaction chambers from a single microliter of liquid. Using these microreactors, we can image molecules as they freely diffuse, opening the door to high-throughput single molecule fluorescence correlation spectroscopies and analysis of DNA binding proteins and transcription factors in a highly parallel, yet single molecule, manner. These resealable femtoreactors also have application to digital PCR and next-generation sample preparation for DNA sequencing.