We previously described that Trask is physically associated with Src kinases, in particular with Yes. The presence of this interaction was determined for all the Trask mutants. The M4, M5, and M8 constructs that contain ICD regions and phosphorylated tyrosines interact with Yes. The mutants that lack the ICD or the three intracellular tyrosines fail to interact with Yes. The YDF mutant also fails to interact with Yes. Conclusions cannot be drawn regarding any quantitative differences among the mutants with regards to tyrosine phosphorylation or with regards to interaction with Yes. This is because the constructs have BI-9564 different expression characteristics, likely due to different protein half lives, precluding direct comparative analyses of their phosphorylation or interaction levels. Next we determined the effect of each of the Trask mutants on the anti-adhesive function of Trask. When overexpressed, Trask is constitutively phosphorylated and inhibits cell spreading and adhesion. The effects on cell adhesion were qualitatively and quantitatively assessed following doxycycline induction. The M4, M5, and M8 mutant constructs retain the ability to inhibit cell adhesion similar to wild type Trask. After doxycycline induction, these cells have the number of adherent cells seen in their uninduced Methimazole controls. On the other hand the M7, M9, and YDF mutants fail to inhibit cell adhesion showing cell adhesion after doxycycline their uninduced controls. These results are consistent among four different clones of each transfectant and therefore are not due to clonal variation. Data obtained from additional clones, with microscopic images from several additional fields and magnifications, and quantitative adhesion assays from additional clones are shown in Figure S2. When doxycycline induction is initiated in detached cells at the time of plating, cells fail to adhere. When the induction is initiated in fully adherent cells, there is also an inhibition of cell adhesion, although the loss of cell adhesion occurs more gradually. The induction of Trask overexpression results in the dephosphorylation of focal adhesion kinase, consistent with the disruption of cell adhesion and dismantling of focal adhesions.