Cells generate defined forces on integrin-matrix contacts and respond to the forces that those contacts exert on them. Sensing matrix rigidity and generating the correct force are critical functions for cells in development, wound healing and diseases such as cancer. A major hallmark of transformed cells is their ability to grow on soft agar, i.e. in the absence of force generated on matrix contacts. We have developed new methods to measure force-dependent changes in matrix-integrin-cytoskeleton links and to measure the forces that cells generate on submicron areas. We observed force-dependent reinforcement of integrin-cytoskeleton linkages in proportion to the force applied. Surprisingly, matrix-liganded fibronectin receptor-cytoskeleton linkages are reinforced but unliganded integrins are not. Linkage formation and dynamics are tyrosine phosphatase/kinase dependent. Tyrosine phosphatase inhibitors block reinforcement and the tyrosine kinase, Src, inhibits reinforcement but only of vitronectin-avb3-cytoskeleton linkages. We now propose to measure reinforcement in cells missing tyrosine phosphatases (Shp2-/-, RPTP alpha-/-, and PTEN-/- cells) that are altered in motility on fibronectin or vitronectin. The first two phosphatases are strongly implicated in fibronectin and vitronectin-dependent motility, respectively, whereas PTEN functions primarily as a lipid phosphatase and normally suppresses motility. In addition, we are studying primary Pyk2-/-, FAK-/+/Pyk2-/-, Src-/-/Pyk2-/-, Fyn-/-, Fyn-/-/Src-/- and related control cells to examine how tyrosine kinases modulate the dynamics of force-dependent linkages. To understand how cellular forces are generated and controlled, we have developed force sensors in silicon chips and have improved the laser tweezers measurements of isometric traction forces. Our studies show that traction forces are rearward in the front of all cells tested and switch to forward direction in the nuclear region. Further, forces of the same level are generated on both dorsal and ventral surfaces, which validates the measurements of traction forces from the dorsal surface. We will now measure how force changes when actin dynamics are altered and when cells are stimulated to modify contractility or myosin phosphorylation. These studies will enable us to define the molecular bases of force sensing and force generation, which is relevant to a deeper understanding of metastasis and development.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
Project #
Application #
Study Section
Cell Development and Function Integrated Review Group (CDF)
Program Officer
Deatherage, James F
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Columbia University (N.Y.)
Other Domestic Higher Education
New York
United States
Zip Code
Sheetz, Michael P (2012) Following nature's challenges. Nat Med 18:1483-5
Moore, Simon W; Zhang, Xian; Lynch, Christopher D et al. (2012) Netrin-1 attracts axons through FAK-dependent mechanotransduction. J Neurosci 32:11574-85
Wombacher, Richard; Heidbreder, Meike; van de Linde, Sebastian et al. (2010) Live-cell super-resolution imaging with trimethoprim conjugates. Nat Methods 7:717-9
Cai, Yunfei; Rossier, Olivier; Gauthier, Nils C et al. (2010) Cytoskeletal coherence requires myosin-IIA contractility. J Cell Sci 123:413-23
Zhang, Xian; Jiang, Guoying; Cai, Yunfei et al. (2008) Talin depletion reveals independence of initial cell spreading from integrin activation and traction. Nat Cell Biol 10:1062-8
Boldogh, Istvan R; Pon, Liza A; Sheetz, Michael P et al. (2007) Cell-free assays for mitochondria-cytoskeleton interactions. Methods Cell Biol 80:683-706
Cai, Yunfei; Biais, Nicolas; Giannone, Gregory et al. (2006) Nonmuscle myosin IIA-dependent force inhibits cell spreading and drives F-actin flow. Biophys J 91:3907-20
Kundaje, Anshul; Middendorf, Manuel; Shah, Mihir et al. (2006) A classification-based framework for predicting and analyzing gene regulatory response. BMC Bioinformatics 7 Suppl 1:S5
Kundaje, Anshul; Middendorf, Manuel; Gao, Feng et al. (2005) Combining sequence and time series expression data to learn transcriptional modules. IEEE/ACM Trans Comput Biol Bioinform 2:194-202
Middendorf, Manuel; Ziv, Etay; Wiggins, Chris H (2005) Inferring network mechanisms: the Drosophila melanogaster protein interaction network. Proc Natl Acad Sci U S A 102:3192-7

Showing the most recent 10 out of 43 publications