Overview Cell Structures Cell Migration Cell Division  

Mechanosensitive Protein Fluxes at Focal Adhesions

Guo and Wang, Mol. Biol. Cell. 18:4519-4527 (2007)

Although focal adhesion is recognized as a likely structure for sensing adhesion-mediated mechanical signals, the mechanism remains unclear. Zyxin, with its variable association at focal adhesions, may represent an important component of the mechanosensitive mechanism. This possibility was investigated by examining the distribution of GFP/RFP-tagged zyxin and actin at focal adhesions under different mechanical conditions.

Zyxin Tails at Focal Adhesions in Cells Immobilized on Stiff Substrates
While few zyxin tails are present in migrating 3T3 fibroblasts expressing RFP-zyxin, prominent tails form when these cells are immobilized on adhesive islands generated by microcontact printing on stiff polyacrylamide. Time-lapse recording indicates that these tails are highly active structures. Recording time, 128 min.

Lack of Zyxin Tails in Cells Immobilized on Soft Substrates
Few long zyxin tails are found when 3T3 fibroblasts are immobolized on soft substrates, indicating that the flux is sensitive to mechanical signals. Recording time, 90 min.

Flux of Zyxin along the Tail As Detected by Photobleaching

A dark spot placed at a zyxin tail by photobleaching (arrowhead) moves away from the edge indicating that there is a flux of molecules toward the cell interior. An identical flux was found with actin, suggesting that the two move as a complex. Recording time, 5 min.

Concentration of Phosphotyrosine at Focal Adhesions but Not Along the Zyxin Tail
The distribution of phosphotyrosine relative to zyxin was examined in NRK cells co--expressing RFP-zyxin (upper panel) and YFP-dSH2 (lower panel; a probe for phosphotyrosine). Phosphotyrosine is concentrated at focal adhesions but not along zyxin tails. Recording time, 62 min.

Requirement of Tyrosine Kinase Activities for the Maintenance of Zyxin Tails
Upon treatment with the tyrosine kinase inhibitor genistein, zyxin tails detach from focal adhesions and disappear into the the cytoplasm without affecting the association of zyxin at focal adhesions. Therefore, tyrosine kinase activities at focal adhesions are required for generating the flux and/or maintaining the association with the actin tail. Recording time, 132 min.