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Antioxid Redox Signal. 2014 Feb 20;20(6):986-99. doi: 10.1089/ars.2013.5708. Epub 2014 Jan 15.

Fluorescence-based force/tension sensors: a novel tool to visualize mechanical forces in structural proteins in live cells.

Author information

1
1 Department of Biochemistry, Nanjing Medical University , Nanjing, People's Republic of China .

Abstract

SIGNIFICANCE:

Three signaling systems, chemical, electrical, and mechanical, ubiquitously contribute to cellular activities. There is limited information on the mechanical signaling system because of a lack of tools to measure stress in specific proteins. Although significant advances in methodologies such as atomic force microscopy and laser tweezers have achieved great success in single molecules and measuring the mean properties of cells and tissues, they cannot deal with specific proteins in live cells.

RECENT ADVANCES:

To remedy the situation, we developed a family of genetically encoded optical force sensors to measure the stress in structural proteins in living cells. The sensors can be incorporated into specific proteins and are not harmful in transgenic animals. The chimeric proteins distribute and function as their wild-type counterparts, and local stress can be read out from changes in Förster resonance energy transfer (FRET).

CRITICAL ISSUES:

Our original sensor used two mutant green fluorescence proteins linked by an alpha helix that served as a linking spring. Ever since, we have improved the probe design in a number of ways. For example, we replaced the helical linker with more common elastic protein domains to better match the compliance of the wild-type hosts. We greatly improved sensitivity by using the angular dependence of FRET rather than the distance dependence as the transduction mechanism, because that has nearly 100% efficiency at rest and nearly zero when stretched.

FUTURE DIRECTIONS:

These probes enable researchers to investigate the roles of mechanical force in cellular activities at the level of single molecules, cells, tissues, and whole animals.

PMID:
24205787
PMCID:
PMC3924807
DOI:
10.1089/ars.2013.5708
[Indexed for MEDLINE]
Free PMC Article

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