Text Box: Research My research interests include single molecule biophysics, cell biology and nanotechnology. My near-future research plan is 1) to unravel the nuclear transport mechanism; 2) to investigate the nuclear envelope disassembly mechanism and 3) to apply the quantum dots (QDs) as bio-probes in the biological systems. My long-term career goal is to set up an advanced biophysical research program which includes four subdivisions: single molecule imaging, cell biology, advanced optics and nanocrystal materials fabrication and application. The individual subdivisions can be developed independently and can be combined to study challenging research projects. The following are the brief introduction for each project:      

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Text Box: Pante, N. Curr. Opin. 1996, Cell Bio. 8:397-406 Text Box: Nucleocytoplasmic Transport Mechanism In eukaryotic cells, nuclear pore complexes mediate bidirectional transport of proteins, RNAs, and ribonucleoprotein complexes across the double-membrane nuclear envelope.  The functional bidirectional traffic is vital for the health and viability of cells. Dysfunction of transport or mutation of nuclear porins can result in numerous human diseases including leukemia, cancers, and primary biliary cirrhosis. However, the transport mechanism is still poorly understood though numerous models have been postulated. Single molecule methods have been proven to be powerful way to elucidate problems unresolved by ensemble average methods. We developed narrow-field epi-fluorescence single molecule microscopy in Dr. Musser's lab to successfully image and track single molecule transitions through single NPCs for the first time. The interaction time between transiting cargos and the pore is ~ 9 ms and cargos mainly interact with the center of the pore for our model protein NLS-2xGFP (labeled with 4 organic dyes). We also found that transport time and efficiency can be altered when pores are occupied by Importin beta, one of the main transport cofactors. I now seek to expand these single molecule investigations. My goal is to both improve the experimental techniques and to seek answers for new important questions: where is the dominant binding sites within the pore for transiting molecules? Do signal-dependent and signal-independent cargos follow different transport paths? How do large molecules go through the pore? Whether or not the same story happens in lower eukaryotic cells? The answers to these questions could result in fundamental advances in the unraveling of nuclear transport mechanism.  

Text Box: Nuclear Envelope Disassembly Mechanism During mitosis, a single nucleus gives rise to two nuclei that are identical to the parent nucleus. Mitosis consists of a continuous sequence of events that must be carried out once and only once. Two such important events are the disassembly of the nuclear envelope (also known as nuclear envelope breakdown (NEBD)) during the first stages of mitosis, and its accurate reassembly during the last stages of mitosis. NEBD mechanism is still controversial and has been described by two different models: the phosphorylation model and the microtubule-dependent model. The former model suggests that NEBD is initiated when maturation-promoting factor (MPF) actively imports into the nucleus and starts phosphorylating NPCs and nuclear lamina proteins, which last until 10 minutes before NEBD. In contrast, the latter model argues that the beginning of NEBD starts with spindle microtubules causing NE folds and invaginations, up to 1 hour before NEBD, which would predict that MPF passively enters the nucleus much earlier. Surprisingly the permeability of NE is not lost completely even at the last stage of NE disassembly.  I suspect there are multiple steps during the NEBD in which some steps occurred very briefly which easily could be undistinguished by average ensemble methods. Therefore I intend to study the mechanism at both the bulk and single molecule levels to test current models or to postulate new models. It is well known that the interactions between the thousands of phenylalanine-glycine (FG) repeats of the Nups and cargos are responsible for the selectivity and the permeability of nuclear pore. Numerous studies show that the sequential dissociation of Nups from NPCs occurs before the NE totally beaks down. However, how the individual Nups affects nuclear transport is still unknown. The transport dependence of kinase molecules on a number of individual Nups could also supply more direct evidences for the understanding of NEBD.   D ng  

Text Box: Application of QDs as Bio-probes Highly photostable fluorescent bio-probes can make a revolutionary progress for drug-delivery study and biomedical imaging. Quantum dots are such promising candidates which have many advantages over organic dyes people used: 1) cross-section of excitation is much larger; 2) nearly no photobleaching; 3) narrow spectral width makes the separation of different wavelengths much easier and avoids cross-talk from the emission spectral tail of organic dyes; 4) multiple color fluorescent probes are available from single element QDs merely by changing their sizes; 5) the QDs can be detected by electron microscopy (EM) which is an independent or a good complementary method to fluorescent studies. Due to these unique characteristics, the applications of QDs as bio-probes have been intensively studied. So far II/VI semiconductor QDs have been mostly used as luminescence probes in the previous reports of bioimaging. I plan to introduce the QDs as bio-probes in the research of nuclear transport and NEBD which is expected to produce higher S/N ratio for single molecules imaging and to obtain the EM data simultaneously. Microinjection of different size QDs with solubilization layer into the living cells, decoration of the solubilization layer of QDs with the nuclear localization signal (NLS) or the nuclear export signal (NES) or attaching QDs to other cargo molecules can be the alternative applications.