hwangl

Dr. Ling Chin Hwang

Room: E10
0114 222 2847
l.hwang@sheffield.ac.uk

General

Career History

  • 2015 - present: Lecturer, Dept. of Molecular Biology and Microbiology, University of Sheffield, UK
  • 2010 - 2015: Postdoctoral Fellow, National Institutes of Health, Bethesda, MD, USA
  • 2006 - 2009: Postdoctoral Researcher, Dept. of Physics, University of Oxford, UK
  • 2002 - 2006: PhD., Chemistry, National University of Singapore
  • 1995 - 2000: BSc. (Honors), Chemistry, National University of Singapore

Research Keywords

Biophysics, biochemistry, single-molecule fluorescence microscopy, bacterial chromosome segregation and cell division

Related Links

Research

We use multidisciplinary techniques such as single-molecule imaging, synthetic biology, biochemistry and microfluidics to study the molecular mechanisms of spatial organization in bacteria using cell-free systems. We are interested in understanding how bacteria, despite their small size and lack of obvious cytoskeleton, are able to spatially organize genomic DNA and proteins to specific locations in the cell. These bacterial positioning systems form distinctive patterns within the cells, and are involved in important cellular functions such as DNA segregation, cell division and motility.
“What I cannot create, I do not understand”, as Richard Feynman once wrote. Our lab works on this basis coming from a reductionist approach, piecing together minimal components from a cell to recreate the patterns seen inside the cell.


Chromosome and plasmid segregation

How do you ensure that each daughter cell inherits a copy of genomic DNA for its survival? Most bacterial chromosomes and low copy number plasmids encode an active segregation machinery called ParABS to partition replicated DNA prior to cell division. It was observed that low copy number plasmids are precisely localized to opposite cell halves upon replication (top left panel) but the mechanism underlying this positioning is not well understood. We reconstituted the plasmid partition system in vitro and coated a flow cell with a ‘DNA carpet’ to act as a biomimetic of the bacterial nucleoid (bottom panel). Purified proteins and plasmids that were fluorescently-labeled were flowed in and we directly watched their dynamics using Total Internal Reflection Fluorescence Microscopy (TIRFM). We found that plasmid dynamics are driven by transient protein gradients or patterns that coat the DNA carpet (top right panel). Future studies will be carried out on understanding how similar protein patterns drive chromosome segregation in bacteria.


fig1Figure 1: P1 plasmid partitioning in E. coli from Hatano and Niki et al. (top left panel). Reconstituted P1 plasmid dynamics (top right panel) imaged with TIRF microscopy (bottom panel) from Hwang et al.



Cell division dynamics

In E. coli, the cell division machinery MinDE oscillates from pole-to-pole to position the cell division septum in the middle (top panel). We are interested in the spatial patterning on a molecular level and have reconstituted the Min oscillations on a supported lipid bilayer that mimics the E. coli cell membrane (bottom panel). These traveling waves and spirals are reminiscent of patterns found in nature (Turing patterns) such as zebra stripes and leopard spots, which are based on similar reaction-diffusion mechanism. These in vitro Min patterns will allow us to explore the characteristics of self-organization in the cell.


fig2Figure 2: MinD oscillations in E. coli taken from Raskin et al. 1999 (top panel). Reconstituted Min waves on supported lipid bilayer (bottom panel).



Fluorescence microscopy

Single-molecule fluorescence microscopy allows us to watch the movement and interactions of individual DNA and protein molecules, which is inaccessible with conventional biochemical approaches. Our lab uses TIRF microscopy to image surface-immobilized molecules and surface-mediated processes to directly observe single-molecule to mesoscale dynamics. We are interested in developing and applying single-molecule and super-resolution microscopy to answer biological questions as part of the Imagine-Imaging Life program.

Teaching

Level 3 Modules

PhD Opportunities

I welcome applications from self-funded prospective home and international PhD students; see examples of a possible project below.

You can apply for a PhD position in MBB here.

Contact me at l.hwang@sheffield.ac.uk for further information.



Single-molecule fluorescence study of bacterial chromosome segregation dynamics

Cell division is a process of fundamental importance in all forms of life. During cell division, DNA must be properly segregated into each daughter cell. The aim of this project is to gain insight into the molecular mechanisms underpinning bacterial chromosome segregation.
The process of division in bacterial chromosomes is regulated by the Par (partition) proteins, which bind to DNA. Recent advances in fluorescence microscopy have revealed that bacterial chromosomes are highly organized and segregate with distinctive patterns in the cell; low copy number plasmid partitioning is driven by protein patterns. In this project, model systems consisting of nanopatterned arrays of biomolecules will be formed using lithographic techniques developed in Sheffield, and used as “workbenches” on which to study the spatiotempral dynamics of the partition proteins of the Par system from Vibrio cholerae and how they are involved in driving chromosome movement and positioning. The project combines multidisciplinary approaches including biochemistry, single-molecule fluorescence microscopy and bionanotechnology to visualize the chromosome segregation proteins on synthetic surfaces in a cell-free system.
The project is especially well suited to Chemical Biology graduates, but applications are invited from graduates in Chemistry, Biochemistry and Biophysics.

Further Information









































Selected Publications

Journal articles