A Single-molecule Study of Molecular Mechanisms and Cellular Pathways of Viral Infection
The major interest of Professor Zhuang and her group is to study complex biological processes at the single molecule (or single working unit) level. We are also interested in developing new imaging techniques to image biological molecules and cells.
Single molecule studies of complex biological processes
Understanding fundamental molecular mechanisms underlying biological processes is one of the major goals in modern biology. As we enter the post-genomic era and biology becomes more quantitative, this goal is becoming more and more accessible. However, roadblocks still exist on the way to quantitative and mechanistic understandings of complex biological processes. Among those, the challenge of characterizing complex dynamics is a critical one. The existence of multiple kinetic paths and transient intermediate states often makes biological processes difficult to dissect by ensemble methods, as individual reaction steps of a multi-step process are usually not synchronized among molecules. To tackle this problem, my research group is developing biophysical techniques to monitor the behavior of individual biological molecules and particles and thus to elucidate complex dynamics beyond the limit of ensemble methods. Currently we are exploiting these techniques in four areas.
1) Fundamental understanding of viral infection
We are developing state-of-the-art instrumentation to image the behavior of individual viral proteins, viral RNAs and viruses in cells and to elucidate molecular mechanisms and cellular pathways of viral infection. Three specific areas are being studied. 1) We are tracking the behavior of individual viruses to elucidate individual steps of the endocytic pathway. 2) We are investigating the conformational dynamics of viral fusion protein at the single-molecule level to illuminate the mechanism by which fusion proteins catalyze viral membrane fusion. 3) We are tracking the behavior of single viral genes to explore the molecular mechanisms underlying the nuclear traffic of viral genes.
These experiments allow us to visualize directly the infection process in real time, dissect individual stages of the viral entry pathway, and obtain a better understanding of the molecular mechanisms governing the influenza infection.
2) Structural dynamics and function RNA and ribonucleoprotein (RNP) enzymes
We are studying the folding and assembly dynamics of these enzymes at the single-molecule level using fluorescence spectroscopy and exploring the correlation between their structural dynamics and function. Three specific areas are being studied. 1) We are investigating small RNA enzymes to uncover basic molecular mechanisms of RNA structural formation. 2) We are investigating large multi-domain RNA enzymes to explore the capabilities and limitations of RNA as functional enzymes. 3) We are investigating RNP enzymes to elucidate the effect of proteins on the structural dynamics and functional capabilities of RNA.
These single-molecule experiments can detect non-accumulative intermediate states that are extremely difficult if not impossible to detect by ensemble methods and thus can greatly improve our ability to characterize complex RNA structural dynamics. These experiments will provide new and critical knowledge about RNA-RNA and RNA-protein interactions, and thus uncover the physical and chemical principles that determine the structure and behavior of RNA and RNP.
3) RNA interference and micro RNAs
One of the most exciting discoveries in biology in the past few years is that small RNA molecules can control the cell's gene expression level. Two such classes are discovered, small interfering RNA (siRNA) and micro RNA (miRNA). The former controls gene expression by selectively degrading messenger RNAs and the latter by inhibiting translation. The biochemical pathways via which these small RNAs control gene expression are far from being completely understood. We are using single-molecule fluorescence spectroscopy to study the interaction of key enzymes on these pathways, such as Dicers and RISC complexes, with the small RNAs to elucidate the molecule mechanism of gene control by siRNA and miRNA.
4) Nano-electronic devices for sensing bio-molecules and bio-pathogens at the single-unit level
Despite the tremendous potential that single-molecule approaches have in exploring complex biological systems, current single-molecule techniques all have limitations and new single-molecule detection technology is in great demand. We are developing nano-electronic FRET devices for sensing biological molecules and biological pathogens at the single-unit level. These ultra-sensitive semiconductor nanowires based sensors will strongly enhance our ability to detect and characterize a large variety of microscopic biological entities at the single-unit level, opening a new realm of single-molecule biophysics and biological sensing technology.