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Mechano-coupling and regulation of contractility by the vinculin tail domain.

Mierke CT, Kollmannsberger P, Zitterbart DP, Smith J, Fabry B, Goldmann WH - Biophys. J. (2007)

Bottom Line: Compared to wild-type, cell stiffness was reduced in vinculin -/- and head cells and was restored in tail and rescue cells.The power-law exponent of the creep modulus was force-independent and did not differ between cell lines.Furthermore, the regulatory function resides in the tail domain of vinculin containing the paxillin-binding site.

View Article: PubMed Central - PubMed

Affiliation: Center for Medical Physics and Technology, Department of Physics, Biophysics, University of Erlangen-Nuremberg, Erlangen, Germany. claudia.mierke@t-online.de

ABSTRACT
Vinculin binds to multiple focal adhesion and cytoskeletal proteins and has been implicated in transmitting mechanical forces between the actin cytoskeleton and integrins or cadherins. It remains unclear to what extent the mechano-coupling function of vinculin also involves signaling mechanisms. We report the effect of vinculin and its head and tail domains on force transfer across cell adhesions and the generation of contractile forces. The creep modulus and the adhesion forces of F9 mouse embryonic carcinoma cells (wild-type), vinculin knock-out cells (vinculin -/-), and vinculin -/- cells expressing either the vinculin head domain, tail domain, or full-length vinculin (rescue) were measured using magnetic tweezers on fibronectin-coated super-paramagnetic beads. Forces of up to 10 nN were applied to the beads. Vinculin -/- cells and tail cells showed a slightly higher incidence of bead detachment at large forces. Compared to wild-type, cell stiffness was reduced in vinculin -/- and head cells and was restored in tail and rescue cells. In all cell lines, the cell stiffness increased by a factor of 1.3 for each doubling in force. The power-law exponent of the creep modulus was force-independent and did not differ between cell lines. Importantly, cell tractions due to contractile forces were suppressed markedly in vinculin -/- and head cells, whereas tail cells generated tractions similar to the wild-type and rescue cells. These data demonstrate that vinculin contributes to the mechanical stability under large external forces by regulating contractile stress generation. Furthermore, the regulatory function resides in the tail domain of vinculin containing the paxillin-binding site.

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Stiffness (top row) and power-law exponent (bottom row) in F9 wild-type and vinculin mutant cell lines obtained from the fit of Eq. 1 to the creep response to a 0.5 nN force step. Between 60 and 86 cells from each cell line were measured. (*p < 0.05.)
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fig4: Stiffness (top row) and power-law exponent (bottom row) in F9 wild-type and vinculin mutant cell lines obtained from the fit of Eq. 1 to the creep response to a 0.5 nN force step. Between 60 and 86 cells from each cell line were measured. (*p < 0.05.)

Mentions: Equation 1 was fitted to the displacement of each bead in response to a 0.5 nN force step. From the fit, one value for cell stiffness (1/a) and one value for the power-law exponent (b) were obtained. For the vast majority of the beads, the quality of the fit was good (Fig. 3). The median deviation between the fit and the measured creep response was 3.7% (median correlation coefficient, r2 = 0.97). Within any given cell line, the stiffness values of individual cells showed approximately a log normal distribution, and the power-law exponent showed a normal distribution (data not shown). To obtain the average response of a given cell line, the geometric mean of stiffness and the arithmetic mean of the power-law exponent, averaged over all cells, were computed. F9 wild-type, rescue, and tail cells all displayed nearly identical stiffness values, whereas the stiffness of vinculin −/− and head cells were 33% and 24% lower, respectively (Fig. 4). These differences were statistically significant (p < 0.05). However, the power-law exponent did not differ significantly between F9 wild-type and the four vinculin mutant cell lines (Fig. 4).


Mechano-coupling and regulation of contractility by the vinculin tail domain.

Mierke CT, Kollmannsberger P, Zitterbart DP, Smith J, Fabry B, Goldmann WH - Biophys. J. (2007)

Stiffness (top row) and power-law exponent (bottom row) in F9 wild-type and vinculin mutant cell lines obtained from the fit of Eq. 1 to the creep response to a 0.5 nN force step. Between 60 and 86 cells from each cell line were measured. (*p < 0.05.)
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2481521&req=5

fig4: Stiffness (top row) and power-law exponent (bottom row) in F9 wild-type and vinculin mutant cell lines obtained from the fit of Eq. 1 to the creep response to a 0.5 nN force step. Between 60 and 86 cells from each cell line were measured. (*p < 0.05.)
Mentions: Equation 1 was fitted to the displacement of each bead in response to a 0.5 nN force step. From the fit, one value for cell stiffness (1/a) and one value for the power-law exponent (b) were obtained. For the vast majority of the beads, the quality of the fit was good (Fig. 3). The median deviation between the fit and the measured creep response was 3.7% (median correlation coefficient, r2 = 0.97). Within any given cell line, the stiffness values of individual cells showed approximately a log normal distribution, and the power-law exponent showed a normal distribution (data not shown). To obtain the average response of a given cell line, the geometric mean of stiffness and the arithmetic mean of the power-law exponent, averaged over all cells, were computed. F9 wild-type, rescue, and tail cells all displayed nearly identical stiffness values, whereas the stiffness of vinculin −/− and head cells were 33% and 24% lower, respectively (Fig. 4). These differences were statistically significant (p < 0.05). However, the power-law exponent did not differ significantly between F9 wild-type and the four vinculin mutant cell lines (Fig. 4).

Bottom Line: Compared to wild-type, cell stiffness was reduced in vinculin -/- and head cells and was restored in tail and rescue cells.The power-law exponent of the creep modulus was force-independent and did not differ between cell lines.Furthermore, the regulatory function resides in the tail domain of vinculin containing the paxillin-binding site.

View Article: PubMed Central - PubMed

Affiliation: Center for Medical Physics and Technology, Department of Physics, Biophysics, University of Erlangen-Nuremberg, Erlangen, Germany. claudia.mierke@t-online.de

ABSTRACT
Vinculin binds to multiple focal adhesion and cytoskeletal proteins and has been implicated in transmitting mechanical forces between the actin cytoskeleton and integrins or cadherins. It remains unclear to what extent the mechano-coupling function of vinculin also involves signaling mechanisms. We report the effect of vinculin and its head and tail domains on force transfer across cell adhesions and the generation of contractile forces. The creep modulus and the adhesion forces of F9 mouse embryonic carcinoma cells (wild-type), vinculin knock-out cells (vinculin -/-), and vinculin -/- cells expressing either the vinculin head domain, tail domain, or full-length vinculin (rescue) were measured using magnetic tweezers on fibronectin-coated super-paramagnetic beads. Forces of up to 10 nN were applied to the beads. Vinculin -/- cells and tail cells showed a slightly higher incidence of bead detachment at large forces. Compared to wild-type, cell stiffness was reduced in vinculin -/- and head cells and was restored in tail and rescue cells. In all cell lines, the cell stiffness increased by a factor of 1.3 for each doubling in force. The power-law exponent of the creep modulus was force-independent and did not differ between cell lines. Importantly, cell tractions due to contractile forces were suppressed markedly in vinculin -/- and head cells, whereas tail cells generated tractions similar to the wild-type and rescue cells. These data demonstrate that vinculin contributes to the mechanical stability under large external forces by regulating contractile stress generation. Furthermore, the regulatory function resides in the tail domain of vinculin containing the paxillin-binding site.

Show MeSH
Related in: MedlinePlus