DISRUPTION OF THE TALIN GENE COMPROMISES FOCAL ADHESION ASSEMBLY IN UNDIFFERENTIATED BUT NOT DIFFERENTIATED EMBRYONIC STEM-CELLS

We have used gene disruption to isolate two talin (-/-) ES cell mutant s that contain no intact talin. The undifferentiated cells (a) were un able to spread on gelatin or laminin and grew as rounded colonies, alt hough they were able to spread on fibronectin (b) showed reduced adhes ion to laminin, but not fibronectin (c) expressed much reduced levels of beta 1 integrin, although levels of alpha 5 and alpha V were wild-t ype (d) were less polarized with increased membrane protrusions compar ed with a vinculin (-/-) ES cell mutant (e) were unable to assemble vi nculin or paxillin-containing focal adhesions or actin stress fibers o n fibronectin, whereas adhesions or actin stress fibers on fibronectin , whereas vinculin (-/-) ES cells were able to assemble talin-containi ng focal adhesions. Both talin (-/-) ES cell mutants formed embryoid b odies, but differentiation was restricted to two morphologically disti nct cell types. Interestingly, these differentiated talin (-/-) ES cel ls were able to spread and form focal adhesion-like structures contain ing vinculin and paxillin on fibronectin. Moreover, the levels of the beta 1 integrin subunit were comparable to those in wild-type ES cells . We conclude that talin is essential for beta 1 integrin expression a nd focal adhesion assembly in undifferentiated ES cells, but that a su bset of differentiated cells are talin independent for both characteri stics.