[Editors] Force, not light, gives MIT images of cell receptors

[Elizabeth Thomson]

MIT News Office
Massachusetts Institute of Technology
Room 11-400
77 Massachusetts Avenue
Cambridge, MA  02139-4307
Phone: 617-253-2700
http://web.mit.edu/newsoffice/www

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Force, not light, gives MIT researchers images of cell receptors

--Technique could assist in the design and testing of new drug molecules

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For Immediate Release
TUESDAY, JUNE 12, 2007
Contact: Elizabeth A. Thomson, MIT News Office
Phone: 617-258-5402
Email: thomson at mit.edu

PHOTO AVAILABLE

CAMBRIDGE, Mass.--MIT researchers have found a way to glimpse 
interactions between molecules on the surface of a cell.

By measuring the force generated by these cell surface interactions, 
the MIT team was able to image and measure the rate at which 
individual molecules join and separate from receptors on the cell 
surface. These interactions are not visible with traditional light 
microscopy.

"We were able to measure regions of strong intermolecular binding on 
the cell surfaces, which enabled us to map the locations of the 
receptors," said Sunyoung Lee, a graduate student in the Department 
of Materials Science and Engineering and lead author of a paper on 
the work in the June 5 issue of the Proceedings of the National 
Academy of Sciences.

The technique, known as functionalized force imaging, could allow 
researchers to better understand the strength and rates of 
interactions between molecular ligands outside the cell and the 
molecular receptors on the cell surface. These interactions play a 
critical role in cell growth, proliferation and differentiation. It 
could also assist in the design and testing of new drug molecules 
that bind strongly or quickly to the target cell.

Receptors on the cell surface allow the cell to maintain constant 
communication with its environment-they bind to molecules that convey 
information about the environment and instructions telling them what 
functions to carry out. In this study, the researchers looked at a 
receptor called vascular endothelial growth factor receptor-2 
(VEGFR2), which is important for the proliferation, migration and 
differentiation of the vascular endothelial cells that line blood 
vessels.

Researchers in the lab of Krystyn Van Vliet, senior author of the 
PNAS study, are working to understand the kinetics of cell-molecule 
interactions and how a cell responds to mechanical and chemical 
changes in its environment. These changes in function can be 
evidenced by the number and type of receptors displayed on their 
surfaces.

"You can ask specific questions about how the mechanical and chemical 
stimuli outside the cell generate changes in cell surfaces and 
structures within the cell," said Van Vliet, the Thomas Lord 
Assistant Professor of Materials Science and Engineering.

With traditional light-based (optical) microscopy, you can see large 
cell structures like the nucleus and cytoskeleton, but not tiny 
molecules such as individual receptors on the living cell surface. To 
achieve the nanometer-scale spatial resolution required to see these 
molecules on the cell surface, the researchers used mechanical force, 
rather than light.

To pull a bound molecule from its target receptor, a very small force 
of about 100 piconewtons is required. The researchers measured that 
force by attaching anti-VEGFR2 antibody molecules to the end of a 
cantilevered probe in a scanning probe microscope. The cantilever 
oscillates in a regular pattern as it scans along the cell surface, 
and whenever the pattern is disturbed, the researchers can infer that 
the antibody on the probe has bound or "stuck" to its target 
receptor, VEGFR2.

By mapping those reversible interactions at every point on the cell 
surface, the researchers can determine where the receptors are 
located with respect to other cell structures. More importantly, said 
Van Vliet, they can follow the molecular interactions on the cell 
over time, allowing them to determine the binding kinetics, or the 
rate at which molecules join and separate from the cell surface.

The force-based imaging also allows for visualization of the stiff 
cytoskeleton underneath the cell surface, which provides internal 
structure for the cell. By overlapping images of the cytoskeleton and 
the VEGF receptors, the researchers found that most of the receptors 
were located near the cytoskeleton. Such correlations support the 
current hypothesis that VEGF receptor function is linked to that of 
other cell surface proteins including integrins, which transmit 
mechanical forces from the outside to the inside of the cell, Van 
Vliet said.

Other researchers have used this approach to measure binding forces 
in isolated proteins, but the MIT team shows that these tiny forces 
can also be used to visualize binding kinetics on chemically fixed 
(nonliving) cells and living cells.

"It's challenging to do this on cells because their surfaces are made 
up of many different kinds of molecules, which makes them 
topographically rough, chemically diverse, and mechanically 
compliant," said Van Vliet.

Van Vliet and Lee outlined several possible applications of 
functionalized force imaging on endothelial, cancer and stem cells, 
including identification of target receptors for cell-specific drugs; 
comparison of kinetics for individual and clustered receptors; and 
visualizing how the mechanical stiffness of extracellular materials 
alters cell function and cell surface receptor activity over time.

The research was funded by the National Science Foundation Nanoscale 
Exploratory Research, the Center for the Integration of Medicine and 
Innovative Technology, the Hugh Hampton Young Memorial Foundation and 
the Beckman Foundation Young Investigators Program.

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Written by Anne Trafton, MIT News Office