Integrins are heterodimers that mediate cell adhesion in many physiological processes. Binding of integrins to ligands provides anchorage and signals for the cell. However, how force regulates integrin/ligand dissociation is unclear. Atomic force microscopy was used to measure the force dependence of lifetimes of single bonds between a FN fragment and integrin α₅β₁.
First, lifetime-force relationships demonstrated that force prolonged bond lifetimes in the 10-30 pN range, a behavior called catch bonds. Changing divalent cations from Ca²⁺/Mg²⁺ to Mg²⁺/EGTA and to Mn²⁺ caused more pronounced catch bonds. A truncated α₅β₁ construct containing the headpiece but not the legs (trα₅β₁-Fc) formed much longer-lived catch bonds in the same force range. Bindings of two activating mAbs, 12G10 and TS2/16, left shift the catch bond and converted catch bonds to slip bonds, respectively. Catch bonds may provide a mechanical mechanism for the cell to regulate adhesion by applying different forces.
Second, FNIII₇₋₁₀/α₅β₁-Fc/GG-7 bond was stretched to ~ 30 pN and then relaxed to ~ 7 pN at which the bond's lifetime was measured. The strong bond state induced by the 30 pN stretching stayed stable even after the force was reduced to 7 pN. In other words, lower the force would not weaken FNIII₇₋₁₀/α₅β₁-Fc bond once it had been stretched. Similar behaviors were observed for FNIII₇₋₁₀/trα₅β₁-Fc and FNIII₇₋₁₀/mα₅β₁interactions. In addition, the efficiency of the force to induce such a strong bond state for FNIII₇₋₁₀/α₅β₁-Fc interaction in 2 mM Mg²⁺/EGTA condition was characterized. The probability of force to induce the strong bond state increased as force increased and when the force reached 26 pN, all bonds were transit to the strong state.
Moreover, reversible unbending of α₅β₁binding with FNIII₇₋₁₀ under mechanical force were observed, which proved that integrin bending and unbending was dynamic. Importantly, integrin could restore bent conformation even when engaged with its ligand, providing a mechanism for mechanotransduction.
Third, structural changes of α₅β₁under force were observed. The structural changes did not change the trend of lifetime-force relationships of FNIII₇₋₁₀/α₅β₁/GG-7 bond. Moreover, the lifetime for the structural changes to occur and molecular length changes caused by them were characterized.
| Identifer | oai:union.ndltd.org:GATECH/oai:smartech.gatech.edu:1853/31705 |
| Date | 17 November 2008 |
| Creators | Kong, Fang |
| Publisher | Georgia Institute of Technology |
| Source Sets | Georgia Tech Electronic Thesis and Dissertation Archive |
| Detected Language | English |
| Type | Dissertation |
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